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Site-specific proteolytic processing plays important roles in the regulation of cellular activities. The histone modification activity of the human trithorax group mixed-lineage leukemia (MLL) protein and the cell cycle regulatory activity of the cell proliferation factor herpes simplex virus host cell factor 1 (HCF-1) are stimulated by cleavage of precursors that generates stable heterodimeric complexes. MLL is processed by a protease called taspase 1, whereas the precise mechanisms of HCF-1 maturation are unclear, although they are known to depend on a series of sequence repeats called HCF-1PRO repeats. We demonstrate here that the Drosophila homologs of MLL and HCF-1, called Trithorax and dHCF, are both cleaved by Drosophila taspase 1. Although highly related, the human and Drosophila taspase 1 proteins display cognate species specificity. Thus, human taspase 1 preferentially cleaves MLL and Drosophila taspase 1 preferentially cleaves Trithorax, consistent with coevolution of taspase 1 and MLL/Trithorax proteins. HCF proteins display even greater species-specific divergence in processing: whereas dHCF is cleaved by the Drosophila taspase 1, human and mouse HCF-1 maturation is taspase 1 independent. Instead, human and Xenopus HCF-1PRO repeats are cleaved in vitro by a human proteolytic activity with novel properties. Thus, from insects to humans, HCF proteins have conserved proteolytic maturation but evolved different mechanisms.
Site-specific proteolysis has emerged as an important mechanism contributing to the regulation of basic cellular processes such as development (24), metabolism (3), cell cycle progression (7), and apoptosis (22). Site-specific proteolysis regulates both physiological and disease processes, often by activating latent functions.
In human cells, a number of nuclear proteins, such as the trithorax group mixed-lineage leukemia (MLL) protein (11, 21, 35) and herpes simplex virus (HSV) host cell factor 1 (HCF-1) (15, 31, 32), are synthesized as precursors that undergo proteolytic maturation to generate stable, noncovalently associated heterodimeric complexes. The importance of these proteolytic maturation processes is underscored by the finding that the Drosophila homologs of these proteins, Trithorax (Trx) and dHCF, also undergo proteolytic maturation (16, 18). Nevertheless, the cellular mechanism of proteolytic maturation is completely known only for human MLL.
MLL is cleaved by a novel endopeptidase called taspase 1 that utilizes an N-terminal threonine generated by autoproteolysis as the nucleophile for polypeptide cleavage (10). For HCF-1, autocatalytic processing of bacterially synthesized HCF-1 has been observed (28), but the mechanisms of HCF-1 maturation in human cells remain to be clarified. Curiously, in addition to sharing proteolytic maturation processes, MLL and HCF-1 bind each other (36), and both play important roles in the regulation of the cell division cycle (6, 23, 26, 30). These relationships encouraged us to compare their proteolytic maturation pathways along with those of the Drosophila Trx and dHCF homologs.
Of these four proteins, MLL is the largest, consisting of 3,969 amino acids. It was originally discovered because the gene encoding MLL is the site of chromosomal translocations in human childhood leukemias (1, 5, 8). MLL possesses histone H3 lysine 4 (H3K4) methyltransferase activity (20, 21) and is required for maintaining proper Hox and Cyclin gene expression (26, 37). MLL is cleaved by taspase 1 (10) at two sites to generate the associated amino-terminal (MLLN) and carboxy-terminal (MLLC) subunits (11, 21, 35). This proteolysis enhances the H3K4 methyltransferase activity of the MLLC subunit, which promotes Cyclin gene expression and cell cycle progression (26).
The enzyme responsible for Trx cleavage is not known, but it has been hypothesized that it is a homolog of taspase 1 because the region of cleavage (16) contains a putative taspase 1 recognition site (10, 35). The possible importance of Trx cleavage for its biological function has been indicated by the activity of a mutant Trx protein, called TrxE3, which contains a 271-amino-acid deletion (19) that spans the predicted processing site and abrogates Trx protein maturation (16). Trx is required to maintain proper expression of antennapedia and bithorax complex genes during fly development (2). Interestingly, trxE3 mutants display defective antennapedia but not bithorax complex gene expression (25), suggesting a selective role of Trx cleavage in its function.
HCF-1 is a 2,035-amino-acid chromatin-associated protein that was first discovered as a transcriptional coactivator for HSV immediate-early gene transcription, where it stimulates formation of the viral VP16-induced transcriptional regulatory complex (see reference 33 for a review). Proteolytic maturation of HCF-1 involves multiple cleavages at any one of six centrally located 26-amino-acid repeats called HCF-1proteolytic (HCF-1PRO) repeats, generating the associated amino-terminal (HCF-1N) and carboxy-terminal (HCF-1C) subunits (15, 31, 32). The HCF-1N and HCF-1C subunits play separate roles in two key phases of the cell cycle: the HCF-1N subunit promotes passage through the G1 phase, and the HCF-1C subunit is necessary for proper mitosis and cytokinesis during the M phase (14). Proteolytic processing is necessary to ensure proper HCF-1 function, as HCF-1C subunit functions are inhibited if the HCF-1 precursor cannot be processed (14).
Consistent with the importance of HCF-1 proteolytic maturation, the 1,500-amino-acid dHCF protein, although lacking HCF-1PRO repeats, also undergoes proteolytic maturation to generate associated dHCFN and dHCFC subunits (18). The dHCFN and dHCFC subunits display considerable structural similarity to the human HCF-1N and HCF-1C subunits (Fig. 1) as well as functional conservation. Thus, as for the HCF-1N subunit, the dHCFN subunit can associate with the HSV transactivator VP16 and stabilize the VP16-induced transcriptional regulatory complex (18), as well as associate with the Drosophila cell cycle regulators dE2F1 (the homolog of human E2F1) and dE2F2 (the homolog of human E2F4) (27).
Following our interest in HCF protein proteolytic maturation and in the conservation of HCF proteins in animals, we noted that dHCF, albeit not human HCF-1, contains a potential taspase 1 cleavage site within the region predicted by Mahajan et al. (18) for cleavage. This observation has led us to find that the dHCF and Trx proteins are both cleaved by the Drosophila taspase 1 homolog, whereas human HCF-1 proteolytic cleavage is independent of taspase 1. Thus, the Drosophila and human trithorax group proteins share proteolytic maturation pathways, while the Drosophila and human HCF proteins, although conserving proteolysis as a maturation process, mature via different proteolytic mechanisms.
Plasmid constructions and DNA template preparations.A full-length Drosophila taspase 1 cDNA (LD05057) was obtained from the Drosophila Genomics Resource Center, Bloomington, IN, and cloned into the pET-28a(+) His-tagged bacterial expression vector (Novagen). The His-tagged full-length human taspase 1 cDNA bacterial expression vector and the vector for in vitro transcription/translation containing wild-type and CS1/2-mutated MLL amino acids 2400 to 2900 have been described previously (10). The full-length dHCF cDNA cloned in a T7 epitope-tagged Drosophila expression vector (pACXT-dHCF) was a kind gift of Angus Wilson (New York University). For in vitro transcription/translation of the precursor, a PCR fragment encoding dHCF amino acids 733 to 1212 was amplified from pACXT-dHCF and inserted in the pNCITE vector (30). For the Trx precursor, a PCR fragment encoding Trx amino acids 1973 to 2458 was amplified by reverse transcription-PCR from SL2 cell RNA, verified by sequence analysis, and inserted directly into the pGEM-T vector (Promega). Fragments consisting of HCF-1 amino acids 686 to 1166 derived from either wild-type or mutant pCGNHCFFL (31, 32) were PCR amplified using a forward primer including the phage T7 promoter and β-globin translational initiation codon and directly used for in vitro transcription/translation. The Xenopus HCF-1 (xHCF-1) precursor, corresponding to amino acids 1358 to 1653, was PCR amplified from pCMV-SPORT6-xHCF-11358-1653 (clone ID 7653830; BioCat) as for HCF-1 and directly used for in vitro transcription/translation. In vitro transcription/translation and [35S]methionine labeling were performed using the TNT T7 Quick transcription/translation system (Promega) as recommended by the manufacturer. Mutants of the Trx CS2-like and dHCF CS1-like sites were generated by QuickChange site-directed mutagenesis (Stratagene).
Taspase 1 protein expression.His-tagged human taspase 1 and Drosophila taspase 1 were expressed in Escherichia coli BL21(DE5) cells and purified by Ni affinity chromatography (QIAGEN) as recommended by the manufacturer. Proteins eluted from the resin were dialyzed against phosphate-buffered saline, 25% glycerol.
Cell culture and extracts.HeLa cells were grown at 37°C in Dulbecco's modified Eagle's medium with 10% fetal bovine serum (FBS). Drosophila SL2 cells were grown at 25°C in Schneider's medium (GIBCO) with 10% heat-inactivated FBS. Nuclear and cytosolic extracts from HeLa and SL2 cells were prepared as previously described (4). Wild-type or taspase 1−/− mouse embryonic fibroblasts (MEFs) from day 12.5 embryos were grown as described previously (26). Extract was prepared by lysis in radioimmunoprecipitation assay buffer with Complete protease inhibitor cocktail (Roche) on ice for 30 min and clarified by centrifugation at 20,000 × g for 20 min.
In vitro cleavage assays.Taspase 1 in vitro cleavage assays were performed as previously described (10). For comparative experiments, recombinant human taspase 1 and Drosophila taspase 1 activities were titrated on the respective cognate MLL and Trx substrates and used in the amount required to cleave 50% of the substrate (corresponding to 10 ng of human taspase 1 and 50 ng of Drosophila taspase 1). For the xHCF-1 in vitro cleavage assay, 1 μg of human taspase 1 and Drosophila taspase 1 were used. Cell extract in vitro cleavage assays were performed using 22 μl of the indicated cell extract in a 30-μl reaction mixture and incubating reaction mixtures at 30°C for 8 h or the indicated time period. HeLa cell extract heat treatment was at 65°C for 20 min, and protease inhibitors were utilized at the final concentration of 0.5 mg/ml Pefabloc (Roche) and 1× Complete protease inhibitor cocktail (Roche). The reaction mixtures were resolved by 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), the resulting radioactive signals were visualized with a Typhoon Trio+ imaging system and quantified with ImageQuant (Amersham Biosciences).
RNAi and cell transfection.A 183-bp double-stranded RNA (dsRNA) (RNAi 1) and a 495-bp dsRNA (RNAi 2) against the Drosophila taspase 1 gene sequence as well as a nonspecific firefly luciferase dsRNA were used for RNA interference (RNAi). dsRNAs were synthesized with a MEGAscript T7 kit (Ambion), using as templates PCR products amplified with primer pairs (RNA: 2), CAGTCGTTCTGTGTCGGCTA and CTAGGAGCTGCAGGAAGGTG (RNA: 1) and TGGGTTGCTCCATGGTG and TCCAGCGTACAGATCCTG each containing an additional 5′ T7 promoter sequence. For dsRNA treatment, 1 × 106 SL2 cells were seeded in six-well plates in 1 ml of serum-free Schneider's medium and incubated at 25°C for 2 h in the presence of 15 μg of dsRNA before addition of 2 ml of Schneider's medium with 10% heat-inactivated FBS. dsRNA-treated cells were collected after 96 h. For transient transfection, 2 × 106 untreated or 48-h-dsRNA-treated SL2 cells were transfected with 5 μg of DNA using Cellfectin (Invitrogen) as described by the manufacturer. Cells were collected at 48 h posttransfection. For collection, SL2 cells were washed with phosphate-buffered saline, lysed in Laemmli SDS sample buffer, and subsequently analyzed by immunoblot analysis.
Immunoblot analysis.Protein lysates were resolved by SDS-PAGE, transferred to nitrocellulose membrane, and probed with T7 tag monoclonal antibody (Novagen), polyclonal HCF-1C antiserum (H12) (31), or polyclonal dHCFN antiserum, followed by appropriate IRDye800-conjugated anti-mouse or IRDye680-conjugated anti-rabbit immunoglobulin G (Rockland). The immunoreactive bands were detected by fluorescence with an Odyssey infrared imager (LI-COR). The polyclonal dHCFN antiserum was raised in rabbits against a 15-amino-acid peptide (EGSDFVDPAFSSGER) corresponding to the N terminus of dHCF and affinity purified with the peptide immunogen.
Figure 1 shows schematic representations of the human and Drosophila MLL/Trx and HCF proteins highlighting conserved sequence and structural elements. Figure 1A also shows the positions of the two known taspase 1 cleavage sites in MLL, called CS1 (QVD/GADD) and CS2 (QLD/GVDD) (10, 11, 35). By sequence similarity, Trx contains a CS2-like site (QMD/GVDD) (10, 11, 35) and dHCF contains a CS1-like site (QLD/GAGD), both of which lie in positions analogous to the cleavage sites in the respective human homologs (i.e., MLL CS1 and CS2 sites and HCF-1PRO sites). We therefore tested whether the CS2-like Trx and CS1-like dHCF taspase 1 recognition sites are substrates for taspase 1.
Trx and dHCF are cleaved by Drosophila taspase 1.In initial experiments, we tested the ability of human taspase 1 to cleave the Trx and dHCF proteins and found that the human protease showed a strong preference for the human MLL substrate (see below). Therefore, we tested the ability of the Drosophila taspase 1 homolog to cleave Trx and dHCF substrates as shown in Fig. 2. To facilitate this analysis, we generated for each protein shorter precursor cleavage substrates that contain the regions with the putative taspase 1 cleavage sites. Both wild-type and cleavage site mutant substrates were used (Fig. 2A). These precursors were synthesized in vitro in the presence of [35S]methionine and incubated with recombinant Drosophila taspase 1 purified from E. coli, and the products were separated by SDS-PAGE. Figure 2B shows that Drosophila taspase 1 can cleave the wild-type Trx precursor (lanes 1 and 2) but not the precursor carrying the mutation in the CS2-like site (lanes 3 and 4). These results suggest that, like its human homolog MLL, Trx is cleaved by taspase 1 at the CS2-like cleavage site. Similarly, as shown in Fig. 2C, Drosophila taspase 1 can effectively cleave the wild-type dHCF precursor (lanes 1 and 2) but not the CS1-like mutant precursor (lanes 3 and 4). These experiments define the first heterologous protease for HCF protein maturation, the Trx protease Drosophila taspase 1.
To characterize this dHCF protease further, we assessed whether dHCF is cleaved at the taspase 1 cleavage site in vivo. We transfected Drosophila SL2 cells with an epitope-tagged expression vector encoding full-length wild-type and CS1-like mutant recombinant dHCF (rdHCF) and assayed cleavage by immunoblotting with an antitag antibody. As shown in Fig. 2D, synthesis of wild-type rdHCF resulted in an approximately 105-kDa N-terminal fragment (rdHCFN) (lane 2) (18), whereas synthesis of the CS1-like mutant rdHCF led to an approximately 180-kDa product, a size equivalent to that of the full-length protein (rdHCFFL) (lane 3) (18). These results indicate that dHCF is cleaved at a single taspase 1 recognition site.
We also analyzed proteolytic maturation of endogenous dHCF by generating an antibody directed to the N terminus of the protein (see Materials and Methods). Immunoblot analysis of the same SL2 extracts shown in Fig. 2D revealed that endogenous dHCFN comigrates with the processed rdHCFN (compare lanes 1 and 2 in Fig. 2D and E). With this antibody, we could not detect any endogenous full-length dHCF protein, but we could detect the ectopic CS1-like mutant rdHCFFL (compare lane 3 with lane 1). The lack of endogenous full-length dHCF indicates, as previously shown using a dHCFC antibody (9), that dHCF is efficiently processed in Drosophila cells.
To determine whether Drosophila taspase 1 is the sole protease responsible for dHCF processing in vivo, we depleted Drosophila taspase 1 from Drosophila cells by RNAi. As shown in Fig. 2F, independent treatment of SL2 cells with two different taspase 1 dsRNAs (RNAi 1 and RNAi 2) led in each case to the detection of dHCFFL (lanes 3 and 4) compared to untreated (lane 1) or mock RNAi-treated (luciferase, lane 2) cells. The levels of dHCFFL were limited, however, possibly owing to high levels of stable dHCFN remaining from before the RNAi treatment. In contrast, we observed prominent inhibition of rdHCF cleavage when the RNAi-induced Drosophila taspase 1 depletion was associated with concomitant synthesis of dHCF by simultaneous transfection of the epitope-tagged dHCF expression vector. Thus, as shown in Fig. 2G, in taspase 1 RNAi-treated cells, rdHCF processing is significantly reduced (lanes 3 and 4) compared to that in untreated (lane 1) or mock RNAi-treated (lane 2) cells. In summary, our in vitro and in vivo results indicate that Drosophila taspase 1 is responsible for dHCF processing at a single site and indeed may be the sole protease responsible for dHCF maturation. Thus, in flies, where dHCF lacks HCF-1PRO repeats, HCF protein maturation is apparently not autocatalytic as observed for an HCF-1PRO repeat region of human HCF-1 synthesized in E. coli (28).
dHCF is not an effective substrate for human taspase 1.We were surprised to find that dHCF is a substrate of Drosophila taspase 1, because Izeta et al. (12) have shown that dHCF is not processed in hamster cells and yet such cells would be expected to possess taspase 1. To explore this apparent discrepancy, we compared directly the abilities of human and Drosophila cell extracts to cleave the dHCF precursor. Indeed, as the results of Izeta et al. (12) would suggest, the dHCF precursor, albeit cleaved by a Drosophila SL2 extract, was not effectively cleaved by a human HeLa cell extract, as shown in Fig. 3A (compare lanes 2 and 3). This dissimilarity is likely the result of intrinsic differences in the human taspase 1 and Drosophila taspase 1 enzymes, because the same species-specific activity was also observed with purified recombinant human taspase 1 and Drosophila taspase 1 (compare lanes 5 and 6) (see Materials and Methods for enzymatic activity normalization). These results explain why Izeta et al. (12) did not observe dHCF processing by using mammalian cells and emphasize the importance of using a homologous system to assay HCF protein processing (18).
The human and Drosophila taspase 1 enzymes have coevolved with their specific MLL/Trx substrates.The dHCF Drosophila taspase 1 and human taspase 1 cleavage results suggest that taspase 1 activity has changed during evolution. To study this evolution further, we compared the human taspase 1 and Drosophila taspase 1 enzymes on homologous substrates: precursors from the human MLL and Drosophila Trx proteins (Fig. 3B). As shown in Fig. 3C, a human HeLa cell and a Drosophila SL2 cell extract display enhanced activity on the cognate MLL and Trx precursor, respectively (compare lanes 2 and 3 with lanes 5 and 6) (in this assay only MLL-CS2 cleavage is indicated). Furthermore, purified recombinant human taspase 1 and Drosophila taspase 1 display the same cognate substrate preference (Fig. 3D, compare lanes 2 and 3 with lanes 5 and 6). These results suggest that the taspase 1 enzymes have coevolved with their substrates MLL and Trx.
Zhou et al. (38) have defined a human taspase 1 recognition heptapeptide sequence. The MLL CS2 and Trx CS2-like sites differ by a single amino acid (from QLDGVDD in MLL CS2 to QMDGVDD in Trx). To examine the determinants responsible for the cognate enzyme substrate preference of Drosophila taspase 1 on Trx, we converted the Trx CS2-like taspase 1 site to the MLL CS2 sequence via a single amino acid change (Fig. 3B). As shown in Fig. 3E, the humanized Trx precursor [Trx(MLLCS2)] is still a better substrate for Drosophila taspase 1 than human taspase 1, indicating that the species-specific taspase 1 selectivity observed here results from sequence differences that lie outside the so-far-identified heptapeptide recognition sequence.
Human HCF-1 is not a taspase 1 substrate.HCF-1 is cleaved at any one of the six HCF-1PRO repeats (15, 31), and when the region containing all six sites is deleted (14, 32) or the six sites are all individually inactivated by amino acid substitution (29), the protein is no longer cleaved. To study HCF-1 proteolytic cleavage, we first asked whether the HCF-1PRO repeat is a taspase 1 substrate. Therefore, we prepared an in vitro HCF-1 protease substrate containing three of the six HCF-1PRO repeats (HCF-1PRO repeats 1, 2, and 3, called HCF-1rep123) (Fig. 4A) and incubated it with recombinant human taspase 1. Figure 4B shows that human taspase 1 had no effect on this HCF-1 precursor (compare lanes 1 and 2) at a concentration that effectively cleaved the MLL precursor (compare lanes 5 and 6), suggesting that taspase 1 is not an HCF-1PRO repeat protease.
To further exclude the requirement of taspase 1 in HCF-1 maturation, we asked whether HCF-1 processing is affected by the absence of taspase 1. Takeda et al. (26) have shown using MEFs that MLL processing is disrupted in taspase 1−/− knockout mice. In contrast, as shown in Fig. 4C, HCF-1 is processed normally in both wild-type and taspase 1−/− MEFs (compare lanes 2 and 3), with the pattern of cleavage products being identical to the one observed in a HeLa cell extract (lane 1). Together, these results indicate that taspase 1 is not the protease responsible for HCF-1PRO repeat processing. Indeed, taspase 1 is unlikely to be involved in HCF-1 processing generally, as there is no identifiable taspase 1 cleavage site in the entire HCF-1 amino acid sequence.
A human cell activity cleaves HCF-1PRO repeats.To identify an activity responsible for HCF-1PRO repeat processing, we used a wild-type HCF-1rep123 substrate or substrates with HCF-1PRO repeats 1 and 2 mutated either individually (HCF-1repX23 and HCF-1rep1X3) or together (HCF-1repXX3) as illustrated in Fig. 5A. As shown in Fig. 5B, in the absence of cell extract, none of the precursors was effectively cleaved (lanes 1, 3, 5, and 7). In the presence of HeLa cell extract, however, each precursor was cleaved at the wild-type but not mutated HCF-1PRO repeats (compare lanes 2, 4, 6, and 8), displaying a cleavage pattern consistent with HCF-1PRO repeat specific cleavage. These results suggest that HeLa cell extracts possess an activity that can specifically cleave an HCF-1PRO repeat precursor.
To establish the nature of this HCF-1PRO repeat enzymatic activity, we performed a time course experiment using the HCF-1rep123 precursor. As shown in Fig. 5C, under the same assay conditions used for Fig. 5B, the HCF-1PRO repeat cleavage product increased with incubation time for approximately 8 h. Accumulation of the product resulting from cleavage at the HCF-1PRO repeat 1 was quantified as shown in Fig. 5D. These results suggest that in cell extracts the HCF-1PRO repeat proteolytic cleavage is relatively slow but stable for 8 h.
The HeLa cell HCF-1PRO repeat proteolytic activity is heat sensitive but protease inhibitor resistant.To characterize the HCF-1PRO repeat protease further, we assayed the sensitivity of the HeLa cell extract activity to heat and protease inhibitor treatment as shown in Fig. 5E. The HeLa cell activity was sensitive to heat treatment (compare lanes 1 to 3) but resistant to Pefabloc, a serine protease inhibitor (compare lanes 3 and 4) (an assay with thrombin and a substrate with a thrombin cleavage site showed that the serine protease inhibitor was active [data not shown]), and Complete, a serine and cysteine protease inhibitor cocktail (compare lanes 3 and 5). Thus, the HCF-1PRO repeat protease displays both sensitivity (heat) and resistance (serine and cysteine protease inhibitors) to various treatments. We note that the resistance of the HCF-1PRO repeat protease to a serine protease inhibitor differentiates it from the autocatalytic activity described by Vogel and Kristie, which was serine protease inhibitor sensitive (28).
HCF protein maturation mechanism has changed during evolution.The identification of taspase 1 as the protease responsible for dHCF maturation and the observation that human HCF-1 is cleaved by a different activity at the HCF-1PRO repeats indicate that, from fly to human, HCF proteins have conserved proteolytic maturation but evolved different mechanisms. These differences between flies and humans appear to be generally specific to insects or vertebrates, as illustrated in Fig. 6. Thus, the Apis mellifera (honeybee) HCF protein possesses a consensus taspase 1 cleavage site at the same relative location as in the dHCF protein (Fig. 6A), suggesting that taspase 1 cleavage of HCF proteins may be generally conserved in insects. In parallel, a comparison of human, frog, and fish HCF-1 proteins shows that the position (Fig. 6B) and sequence (Fig. 6C) of the HCF-1PRO repeats have been very highly conserved in these three distantly related vertebrate species. Thus, between insects and vertebrates, there appears to have been an evolutionary switch in HCF protein processing mechanism that has been highly conserved within each group.
Interestingly, in some fish (e.g., F. rubripes) and frog (e.g., X. tropicalis) species, in addition to the perfectly conserved HCF-1PRO repeats there is a partially conserved taspase 1 cleavage site (six-of-seven match to a heptapeptide consensus taspase 1 cleavage site) (38) located just C terminal of the last HCF-1PRO repeat (Fig. 6B). This partially conserved taspase 1 cleavage site could represent an active site, indicating that in some species HCF-1 could be processed by both taspase 1 and an HCF-1PRO repeat protease. Alternatively, it could be inactive and instead represent the vestige of an active taspase 1 cleavage site in an ancestral HCF-1 protein. To distinguish between these two possibilities, we asked whether an xHCF-1 cleavage precursor containing the partially conserved taspase 1 cleavage site and the adjacent two HCF-1PRO repeats (xHCF-1rep89 [Fig. 6D]) could be a substrate for either taspase 1 and/or the HCF-1PRO repeat protease. As shown in Fig. 6D, the xHCF-1rep89 substrate is not cleaved by either Drosophila taspase 1 or human taspase 1 at concentrations that can effectively cleave the dHCF precursor (compare lanes 1 to 3 and 4 to 6). Instead, as for human HCF-1 (lanes 8 and 9), the xHCF-1rep89 precursor is cleaved by the HeLa cell extract at positions corresponding to the HCF-1PRO repeats (compare lanes 4 and 7). These data suggest that, like mammalian HCF-1 proteins, xHCF-1 is not cleaved by taspase 1 but is instead processed at the HCF-1PRO repeats by the same activity responsible for human HCF-1 maturation. The ability of the human cell extract to cleave the Xenopus HCF-1PRO repeats provides functional evidence of the high degree of conservation of the HCF-1PRO repeat proteolytic process in vertebrates.
In this study, we have compared the proteolytic maturation processes of two pairs of evolutionarily conserved proteins: the trithorax group proteins human MLL and Drosophila Trx and the HCF proteins human HCF-1 and dHCF. We have shown that, like MLL, the Drosophila Trx is a substrate for the taspase 1 protease. Interestingly, the enzyme-substrate specificity, i.e., human taspase 1 with MLL and Drosophila taspase 1 with Trx, has coevolved. In contrast, with respect to HCF proteins, very different proteolytic pathways have evolved. Thus, the dHCF protein is cleaved by Drosophila taspase 1 but its human (and Xenopus) HCF-1 counterparts are cleaved by what is likely an unusual proteolytic activity.
Evolution of taspase 1 enzyme-substrate specificity.Hsieh et al. (10) noted that sequences encoding the taspase 1 enzymes have been conserved in vertebrates and insects but not in the worm Caenorhabditis elegans and that, correspondingly, the MLL/Trx developmental regulators in vertebrates and insects but not in worms have taspase 1 recognition sites. Consistent with this observation, Trx is indeed a taspase 1 substrate at its CS2-like taspase 1 cleavage site. Examination of the activities of the human and Drosophila taspase 1 enzymes on the MLL and Trx proteins also revealed a more refined coevolution of enzyme and substrate, as human taspase 1 is more active on its cognate MLL substrate and Drosophila taspase 1 is more active on its cognate Trx substrate. This observation suggests that the presence of the taspase 1 enzyme and MLL/Trx protein maturation have been conserved because the cleavage is critically important for proper development (16, 19, 26). Nevertheless, there is considerable flexibility in the interaction between protease and substrate, and for such enzyme-substrate coevolutionary flexibility to be possible, it is likely that taspase 1 does not possess a large number of essential targets.
We note with interest that vertebrates and insects display common longitudinal body segmentation, which is lacking in C. elegans, and that proper MLL and Trx maturation is functionally linked to proper homeodomain gene expression, which is intimately involved in segmentation determination (10, 16, 19, 26). Consistent with the suggestion that MLL/Trx cleavage is required to control the expression of genes involved in body segmentation, in the sea urchin Strongylocentrotus purpuratus, which, like C. elegans, is not longitudinally segmented, the Trx homolog lacks an evident taspase 1 recognition site and there is no evident taspase 1-encoding gene (data not shown).
Although HCF proteins are not known to be involved in regulating segmentation, invertebrate HCF proteins display species-specific patterns of proteolytic maturation that follow the Trx proteins. Thus, in Drosophila, dHCF and Trx are both cleaved by Drosophila taspase 1, and in the honeybee the HCF and Trx homologs possess taspase 1 recognition sites. In contrast, as with the Trx homologs, the sea urchin and worm HCF proteins do not possess evident taspase 1 recognition sites (data not shown); indeed, worm HCF does not undergo proteolytic maturation (34). We suggest that in invertebrates, the HCF and Trx proteins coevolved taspase 1-induced maturation. In contrast, in vertebrates, HCF-1 proteins took on a very different proteolytic maturation mechanism. Nevertheless, the parallels between MLL/Trx and HCF protein maturation suggest that in addition to regulation of the cell cycle, HCF-1 may also play a role in regulation of genes involved in segment determination.
HCF-1 is likely cleaved by an unusual protease.We have described an enzymatic activity in HeLa cell extracts that specifically cleaves the HCF-1PRO repeat. We do not know at this time the precise relationship between the HeLa cell activity described here and the HCF-1 autocatalytic activity described previously (28), but we note some differences between the two activities. The autocatalytic activity, which depends on sequences within the HCF-1C region, results in cleavage at not only HCF-1PRO repeats but also an additional site, called 102, which lacks evident HCF-1PRO repeat sequence similarity and is located just C terminal of the sixth HCF-1PRO repeat (15, 28). This autocatalytic site is, however, not used in vivo when the HCF-1PRO repeats are inactivated by mutation or deletion (14, 29, 32), and we have not observed 102 site cleavage using a precursor substrate spanning this site in the HeLa cell extract assay (F. Capotosti and W. Herr, unpublished results). Thus, in both HeLa cells and HeLa cell extracts, HCF-1 proteolytic maturation appears to be HCF-1PRO repeat specific. We also note that, unlike the autocatalytic 102 site activity, which is sensitive to the serine protease inhibitor Pefabloc (28), the HeLa cell HCF-1PRO repeat activity is resistant to this protease inhibitor (Fig. 5E). Clarifying the relationship between the HeLa cell activity described here and the HCF-1 autocatalytic activity will require further investigation.
Whatever the relationship, however, the properties of the HCF-1PRO repeat protease are likely to be unusual. As shown in Fig. 6C, there are 16 amino acid residues over 19 positions that are identical among the HCF-1PRO repeat consensus sequence of the fish, frog, and human HCF-1 proteins, an impressive level of sequence conservation for a proteolytic recognition sequence. Furthermore, as described previously (32), alanine substitutions at 12 positions over 18 of the human HCF-1PRO repeat 2 affect HCF-1PRO repeat cleavage in HeLa cells (Fig. 6C), a surprisingly large sequence requirement for proteolytic cleavage. Given this extensive sequence conservation and sequence requirements, we suggest that the functional HCF-1PRO repeat element reflects the assemblage of different recognition sequences for (i) the protease itself and (ii) accessory factors that could regulate proteolysis. Such accessory factors could directly recruit or stimulate the protease or could reflect enzymes that modify the HCF-1PRO repeat (e.g., by phosphorylation or glycosylation) to regulate cleavage. Whatever the reason, the large size of the HCF-1PRO repeat results in considerable specificity, as we have been unable to find a match to the HCF-1PRO repeat in any other protein sequence found in public protein databases, suggesting that in toto the mechanisms of HCF-1PRO repeat cleavage are unique to HCF-1 maturation.
Alternate mechanisms of HCF protein cleavage result in HCFN and HCFC subunits of similar structure.The evolutionary comparisons shown in Fig. 6 indicate that although the mechanisms of HCF protein maturation have switched between insects and vertebrates, the end results of proteolytic processing are similar. To illustrate this point, Fig. 6A and B show charge profiles for the two insect (fly and honeybee) and three vertebrate (fish, frog, and human) HCF proteins. These profiles show that the insect and vertebrate HCF proteins all contain basic and acidic regions at corresponding positions. This conservation of amino acid composition is consistent with the important cellular functions these regions have been shown to possess in human HCF-1: G1 phase progression in the case of the basic region (30) and M phase progression (13), transcriptional activation (17), and chromatin association (13) in the case of the acidic region. We note with interest that whether the HCF protein is cleaved by taspase 1, as appears to be the case with the insect HCF proteins, or at the HCF-1PRO repeats, as appears to be the case in the vertebrate proteins, the cleavage site(s) is always positioned between the basic and acidic regions. Thus, although the mechanism for HCF protein proteolysis has changed during evolution, the resulting HCFN and HCFC subunits are very similar in structure.
How and why might a transition from taspase 1- to HCF-1PRO repeat-dependent HCF-1 proteolysis have evolved?The unexpected switch in proteolytic processing mechanism between insect and vertebrate HCF proteins leads to the questions of how and why. Concerning how the HCF-1PRO repeats may have arisen, we note with interest that the six HCF-1PRO repeats in fish and human HCF-1 are all encoded by a single large exon of 1,477 bp in human. (The nine repeats in X. tropicalis are encoded by two exons [five in the first and four in the second], which may have resulted from a duplication of the six-repeat-containing fish/human exon [Fig. 6C].) We imagine that an HCF-1 progenitor acquired the six HCF-1PRO repeats as a single genetic element by recombination, perhaps transposition, prior to vertebrate evolutionary divergence.
Concerning why a switch may have occurred, one possible explanation is that the region containing the HCF-1PRO repeats has been evolutionarily selected as an additional platform for protein-protein interactions, as Vogel and Kristie (29) have shown that the transcriptional coactivator/corepressor FHL2 interacts with nonprocessed HCF-1, stimulating transcription of an HCF-1 target gene. This interaction between HCF-1PRO repeats and cofactors could allow the modulation of HCF-1 processing and activity. Another possible explanation is that, with an HCF-1PRO repeat protease, HCF-1 protein maturation has become independent from taspase 1 and thus from MLL/Trx maturation. This could result in a more flexible regulation of these two important cell cycle regulators. In any case, however, the importance of HCF-1PRO repeat processing during vertebrate evolution is underscored by the remarkable similarity among the repeats themselves in one species (e.g., human) and between vertebrates as divergent as fish and human (Fig. 6C).
In conclusion, in human and fly, the MLL/Trx and HCF proteins have conserved the process of proteolytic maturation, but the MLL/Trx process coevolved with the cognate taspase 1 proteases, whereas HCF-1 and dHCF have apparently evolved very different proteolytic pathways. Whatever the reason for the evolutionary change to HCF-1PRO repeat-dependent processing in vertebrates, it is evident that the HCF-1PRO repeats are unusual cleavage sites, and it is likely that the protease responsible for their cleavage has unusual properties.
We thank Angus Wilson for the dHCF expression vector; Deborah Aufiero for help with characterizing the anti-dHCF antisera; Nouria Hernandez, Christina Hertel, Sara Rodriguez Jato, and Shweta Tyagi for critical readings of the manuscript; and Henrik Kaessmann for advice.
F.C. is the recipient of a Roche Research Foundation Fellowship. These studies were supported by the Swiss National Science Foundation and the University of Lausanne.
- Received 2 May 2007.
- Returned for modification 15 June 2007.
- Accepted 27 July 2007.
- Copyright © 2007 American Society for Microbiology


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All praise is due to Allah SWT who is our Creator and Sustainer. It is Allah alone who deserves all praise. Almighty Allah is unique in all His attributes and His power evades everywhere and all time. May peace and blessings be upon our final and beloved Prophet Muhammad salAllahu alaihi sallam.The following Qur'anic ayah shows the high place that education has in Islam.
'Allah will exalt in degree those of you who believe, and those who have been granted knowledge and Allah is Well– Acquainted with what you do.'
Surah Al Mujadilah: 11
Whilst the benefits of being in the path of knowledge are truly immense;
'He who issues forth in search of knowledge is busy in the cause of Allah until he returns from his quest.'
"If you educate a man, you educate an individual. If you educate a woman, you educate a nation."
African proverb
It is only through gaining knowledge can we advance as a society. At Abu Bakr Girls School we aim to provide a strong Islamic environment in which our pupils achieve their full potential and go on to become outstanding ambassadors for Islam. We are committed to creating a culture of learning in which pupils will strive and flourish Insha'Allah.
Our school is based on a foundation of the teachings Islam and we endeavour to provide a journey that fosters true Iman and love for Islam in the hearts and minds of our pupils, staff, and parents.

subset: commoncrawl


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White Striped Faux Fur Hem Snowman CostumeSKU: 024133
DescriptionLook hot and festive this holiday season in this sparkly snowman costume featuring stripe detailing, white faux fur hem, a striped Santa hat and leg warmers.
Fabric: Polyester
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2. Items received unused, undamaged and in original package.


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Legacy systems and old data: these artifacts pose serious challenges for the modern enterprise. How can companies reconcile the past with the present, and prepare for the future? Check out this episode of InsideAnalysis to find out! Host @eric_kavanagh will interview Dr. Geoffrey Malafsky, new Data Scientist in Residence at InsideAnalysis, and Justin Reock, Evangelist for Rogue Wave Software.
About Eric Kavanagh
Eric has nearly 30 years of experience as a career journalist with a keen focus on enterprise technologies. He designs and moderates a variety of New Media programs, including The Briefing Room, DM Radio and Espresso Series, as well as GARP's Leadership and Research Webcasts. His mission is to help people leverage the power of software, methodologies and politics in order to get things done.
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- Description
DetailsDescription:6/8/10/12mm Diameter Rod Length 400mm Steel Cylinder Liner Rail Linear Shaft Optical AxisSpecification:Material: High Carbon Steel (C45#) + Bearing Steel (GCr15) + Stainless Steel (SUS304)Rated Static Load: 50Rated Dynamic Load: 30Slider Fixed Mode: LockStatic Moment: 50 (kn·m)Fixed Bolt Size: M4 (mm)Diameter: 6/8/10/12mm (Please choose the size you want)Length: 400mm / 15.75" (approx.)Quantity: 1pc
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Homewood Suites by Hilton New Hartford Utica Opens
All Suite Hotel Welcomes Guests Visiting Central New York State
CLINTON, N.Y. and MCLEAN, Va. - Homewood Suites by Hilton, part of Hilton's (NYSE: HLT) All Suites portfolio, announced today its newest property, Homewood Suites by Hilton New Hartford Utica. Designed for guests who want to travel on their own terms, Homewood Suites by Hilton New Hartford Utica offers guests all the comforts of home, whether they are traveling for work or enjoying a well-earned getaway. With 85 new suites, the hotel offers a convenient place to stay for those visiting major sites in the region, such as National Baseball Hall of Fame and Museum, the Finger Lakes area and Hamilton College.
"Our team members want our hotel to feel like a 'home away from home' for guests, offering a comfortable atmosphere where travelers can maintain their normal routine while also experiencing our friendly service," said Marie Borrill, general manager. "From starting the day with a bountiful complimentary breakfast to winding down during our evening social Monday-Thursday nights, our extended stay hotel creates opportunities for guests to interact."
Developed and owned by New Hartford ExStay, LLC, Homewood Suites by Hilton New Hartford Utica offers all suite accommodations featuring fully equipped kitchens and separate living and sleeping areas. Guests are provided all the essentials needed for a smart, reliable and convenient stay including complimentary daily full-hot breakfast, evening social Monday-Thursday, Wi-Fi and a grocery shopping service**. Homewood Suites by Hilton New Hartford Utica also makes it easy for travelers to relax or stay active with an indoor heated pool, outdoor patio with fire pit, a state-of-the-art fitness center, sports court and a practice green for putting. There is also a nature trail adjacent to the hotel, where guests can rent bikes or order boxed picnic lunches at their leisure. In addition, the property also offers meeting space that can accommodate up to 70 people, fully equipped with presentation technology, which is ideal for seminars, training or catered social gatherings.
Located at 185 Woods Park Drive, Homewood Suites by Hilton New Hartford Utica offers guests convenient access to Interstate 90 as well as Hamilton College, Colgate University, National Baseball Hall of Fame and Museum, F.X. Matt Brewing Co., Munson-Williams-Proctor Arts Institute and several dining and shopping options.
Homewood Suites by Hilton New Hartford Utica participates in Hilton's award-winning customer loyalty program, Hilton Honors. Hilton Honors members who book directly through preferred Hilton channels have access to instant benefits, including a flexible payment slider that allows members to choose nearly any combination of Points and money to book a stay, an exclusive member discount, free standard Wi-Fi, as well as digital amenities that are available exclusively through the industry-leading Hilton Honors app, where Honors members can check-in, choose their room, and access their room using a Digital Key. For more information or to make a reservation, visit Homewood Suites by Hilton New Hartford Utica or call 315-801-7200.
Read more about Homewood Suites by Hilton at www.homewoodsuites.com and news.homewoodsuites.com.
**Guest pays for groceries. Other restrictions apply.
Marie Borrill
General Manager, Homewood Suites by Hilton New Hartford Utica
Kristen Wells
Director, Brand Public Relations, Hilton
About Homewood Suites by Hilton
Homewood Suites by Hilton, Hilton's upscale, all-suite, extended-stay hotel brand with more than 500 locations in the United States, Mexico, Canada, and the Caribbean, with 115 properties in the pipeline, is an award-winning leader. The first choice for guests seeking comfortable accommodations when traveling for extended or quick overnight stays, Homewood Suites offers inviting, generous suites featuring separate living and sleeping areas, and fully equipped kitchens with full-size refrigerators. Additional value-driven essentials include complimentary internet, a daily full hot breakfast, and complimentary evening socials every Monday - Thursday. Hilton Honors members who book directly through preferred Hilton channels have access to instant benefits, including a flexible payment slider that allows members to choose nearly any combination of Points and money to book a stay, an exclusive member discount that can't be found anywhere else, free standard WiFi, and digital amenities like digital check-in with room selection and Digital Key (select locations), available exclusively through the industry-leading Hilton Honors mobile app. Homewood Suites is focused on guest satisfaction and stands behind each stay with its 100% Suite Assurance® guarantee. For more information on the extended-stay advantage, visit www.homewoodsuites.com or newsroom.hilton.com/homewoodsuites, and connect with Homewood Suites on Facebook, Twitter, LinkedIn, and Instagram.
About Hilton
Hilton (NYSE: HLT) is a leading global hospitality company with a portfolio of 18 world-class brands comprising more than 6,200 properties with more than 983,000 rooms, in 118 countries and territories. Dedicated to fulfilling its mission to be the world's most hospitable company, Hilton welcomed more than 3 billion guests in its 100-year history, earned a top spot on the 2019 World's Best Workplaces list, and was named the 2019 Global Industry Leader on the Dow Jones Sustainability Indices. In 2020, Hilton CleanStay was introduced, bringing an industry-defining standard of cleanliness and disinfection to hotels worldwide. Through the award-winning guest loyalty program Hilton Honors, the 108 million members who book directly with Hilton can earn Points for hotel stays and experiences money can't buy, plus enjoy instant benefits, including digital check-in with room selection, Digital Key, and Connected Room. Visit newsroom.hilton.com for more information, and connect with Hilton on Facebook, Twitter, LinkedIn, Instagram and YouTube.Download Article Photos


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April 19, 2014 02:28:52
Posted By Mike Luoma
Like listening to science fiction adventure? Join Writer, Reader and Host Mike Luoma for the 300th episode of Glow-in-the-Dark Radio! Mike reads from his books every week – right now it's The Adventures of Alibi Jones! For this special 300th episode, you'll hear chapters twenty-one, twenty-two and twenty-three of the novel Alibi Jones and The Sunrise of Hur! Mike runs down the story so far beforehand, so you can enjoy the chapters even if it's your first time listening in.
Get access to all 300 episodes of Glow-in-the-Dark Radio on Google Drive: https://drive.google.com/folderview?id=0B0uV2VUMyDHwRXczSVBJSUtreGs&usp=drive_web
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We're part of the Comic Related Podcasting Network – Find Us Each Week at http://comicrelated.com


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India's Gaming Business – a deeper Look
India had a conservative outlook toward the gaming industry until the 2000s when the industry saw a major makeover in India. With the ever growing youth population, which now stands at 350 million, and the increasing access to PC and gaming parlors, the future of the gaming industry is indeed bright in the country.
Earlier, the games and consoles created by Nintendo and Sega were not accessible to the middle-income group because of their high prices. However, with the advent of high-speed internet, it has become easier for gamers from the economical sections of the society to access them. Games like the Call of Duty, Counter Strike, FIFA, and DOTA are known for their high quality, action and multiplayer themes. Although designed in the western market, they are highly popular in the Indian market too because of the reformed gaming platforms that now included social media platforms like the Facebook. In fact, Facebook's Farmville and Mafia Wars got millions of Indians hooked.
Status of global gaming industry and India's contribution
One can conceive how fast the gaming industry is expanding, owing to the million dollar contribution it has in the global economy. According to an evaluation in 2017, the valuation of the industry was $108.9 billion with a year-on-year growth of 7.8%. Breaking down the percentage of growth rate, it was found that the contribution of the Asia Pacific region was 47%; India ranked 17th in the list of contributors in the APAC region, generating $818 million with a CAGR of 11.14 percent. Needless to say, it is one of the largest markets for the global gaming industry.
In fact, there was a drastic increase in the numbers in the next two years. In 2019, the industry has been estimated to reach USD 890 million and is estimated to have an annual growth rate of 14.3%. The mobile gaming industry is estimated to lead at 71% share.
Why is India a future hub of the gaming industry?
· World's largest youth population
· Presence of some of the biggest development centers like Nvidia, Microsoft, Zynga, Electronic Arts, Playdom, Sony, Digital Chocolate, etc
· Huge skills base across IT, testing and arts
· Huge base of creative talent
· World's second largest Internet population
PC gaming in India
PC gaming is fast becoming a favorable choice among youngsters who have easy access to quality of game play, affordability and availability of top-end hardware and software systems and improved Internet bandwidth.
India has more than 60% of the population below 35 years of age, which makes it one of the largest potential market for gaming. Certainly, the gaming industry is here to stay!
Mobile gaming
With the advent of smartphones, gaming has become an easy means of entertainment even while one is on the move. Revenue in the Mobile Games segment is expected to show an annual CAGR of 9.8%, which resulted in a market volume of USD 943 million in 2022. In fact, the user penetration in the mobile gaming industry is at 19.9% in 2018 and is expected to hit 26.0% in 2022. Owing to the increasing demand of the mobile gaming industry, 95% of the games are mobile-centric.
Meet the world's top dealmakers of the gaming industry at BizzTech's next online matchmaking event MOBILE GAMES. PC GAMES. VR/AR. CONNECTION | May 15/16to do business in pre-scheduled video calls.


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CaraCompact is a new model of semi-integrated motorhome from Weinsberg. As for now, this camping car, for which the base is new Fiat Ducato, will come only in one variant.
CaraCompact may look like a compact vehicle, and even a little "muscular". Its silhouette is very proportionate thanks to a relatively small width - 220 cm. The width of the interior amounts 205 cm. The car is 674 cm long, and its height amounts 272 cm (inside - 200 cm).
Permissible gross weight is standard, and amounts 3.5 tons. The maximum load capacity is 740 kg.
With unfolding worktop
In the variant, which for now will be the only one available, there're two single beds placed in the rear of the vehicle, as well as a sizable garage. What's interesting, access to the storage compartments placed beneath high suspended beds (measuring 196 x 80 cm) is facilitated by the presence of the blinds moved aside.
The kitchen, with a gas stove and sink, is small, but it can be enlarged with an additional folding part. Under the worktop there're large, soft closing drawers.
Also, a table was placed between the driver and passenger seats, while double sofa has an extra element enabling to enlarge its surface.
In the car, there's of course a bathroom with shower - not too big, but fits in everything what's needed.
Lots of shelves
The interior looks modern and aesthetically due to both small things such as solid handles and practical solutions in the form of a large number of open shelves. The whole interior looks even better thank to the LED lighting.
The equipment of CaraCompact includes, among others, a 149-liter fridge, 110 liter fresh water container and 95 liter waste water container, but also a gas heating system Truma Combi 4 with 10-liter boiler. Some equipment packages are also available with LED strips placed above the cabinets and under the worktop.
Valves available outside
The vehicle has a fairly conventional construction. The walls are "sandwiches" that use polystyrene and aluminum. Glass fibers were used in the construction of the roof, which apart from that was made of plastic GfK and is also insulated.
From the outside CaraCompact isn't too impressive – it's aesthetic, but certainly not extravagant. A very convenient access to valves of onboard devices, all of which are appropriately marked, should be appreciated.
The basic version of the car costs 43,990 euros in Germany, but the manufacturer gives you the opportunity to enrich the vehicle with additional packages. They include, among others, a heated and insulated waste water container, alloy wheels, LED daytime lights and reversing camera.
As for now, CaraCompact is only available in one version, but the manufacturer is already working on new variants and new interior layouts.
During the Caravan Salon 2014 in Dusseldorf, the WEINSBERG brand will present the showcar CaraCompact "Glut" with its optically striking design.


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Chagall Guevara - Chagall Guevara
1991, Sparrow
1. Murder in the Big House
2. Escher's World
3. Play God
4. Monkey Grinder
5. Can't You Feel the Chains?
6. Violent Blue
7. Love is a Dead Language
8. Take Me Back to Love Canal
9. The Wrong George
10. Candy Guru
11. I Need Somebody
12. The Rub of Love
13. If It All Comes True
I don't know how Chagall Guevara wasn't the next big thing back in '91. This is some pretty proficient alt-rock from the likes of Steve Taylor and Dave Perkins (Passafist). "Murder in the Big House" has a definite INXS vibe, which is cool, and the whole album is just filled with great catchy songs. "Love is a Dead Language" is one of my favorites. Steve Taylor pens some pretty thought provoking lyrics this time out with nary an overcooked idea in sight. This is way out of print so it's pretty hard to find these days as far as I know.
Useless Fact: The band's name is taken from "freedom fighter" Che Guevara and an artist... Chagall presumably. I think it's a subtle cry for artistic freedom.


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If anything is true in IT, it's constant change. Every few years—a new evolution happens. While there are always incremental improvements in technologies such as chip designs, network speeds, and storage technologies, sometimes these small changes synergistically align on multiple fronts into something that's new and exciting.
Intent-based networking is such a combination. It's a new way to conceptualize and interact with the network that may fundamentally alter the foundation of networking in your organization. But what is "intent-based networking"? And how does it differ from the day-in, day-out networking that's been running your enterprise for years?
According to the analyst firm ESG, "The concept of intent-based networking is that the network team could simply describe, in plain language, what they wanted to accomplish (the intent) and the network would be able to translate the intent into the numerous policies that would establish the appropriate configuration and settings changes across a complex and heterogeneous environment leveraging automation."¹
If we quickly look at the underlying meaning—there's a whole lot of intelligence and automation behind the scenes to describing something in plain language and having the network automatically build policies. Normally this activity would require a significant manual effort by highly skilled network engineers to modify each device impacted by the desired change.
That's just the "hands on" of the network too—the manual work done by network engineers to modify or configure the devices involved. The benefit of increasing the amount of automation is the reduction of manual processes involved in any network change or implementation process. However, there's also software-derived "intelligence" driving all those changes. Read into that "machine learning" or "artificial intelligence" operating under the covers. The idea is that when performance or any other criteria is not met, the intent-based network system can either notify operations and suggest corrective action. And perhaps even automatically reconfigure the network to ensure compliance with performance goals.
While this may all sound like science-fiction build for a networking future decades away, many of the building blocks for intent-based networking are being deployed today. Organizations leveraging software-defined networking, virtualization technologies, and network analytics platforms should already be able to see how an intent-based architecture could orchestrate an information-rich and highly flexible network environment for them today.
The key to that network intelligence promised in intent-based networking is two-fold: the marriage of advanced hardware and software. Let's concentrate on the software part of the equation since software-defined is one of the most promising trends in IT today. Software is being used to define just about everything in IT today—and networks are no exception. Software can be used to help your wide area networks to network access. Using a software-defined approach, you can provide access to any application--without compromising on security--while gaining awareness of what is hitting your network. In fact, Cisco's Software-Defined Access (SD-Access) can offer you network insights, easier management, and a highly secure network—all delivered through software. SD-Access is part of building Cisco's Digital Network Architecture (DNA) to deliver that much anticipated intent-based networking.
Just like your network, Cisco's Software-Defined Access (SD-Access) is a collection of many different parts spanning identity, network management, and network security. All are critical to true software-defined access, but much of IT's time is spent on network management tasks such as configuring and provisioning and troubleshooting—and Cisco's DNA Center™ can definitely help out your IT staff.
Cisco DNA Center is the foundational controller and analytics platform at the heart of Cisco's intent-based network. It supports the expression of intent for multiple use cases, including base automation capabilities, fabric provisioning, and policy-based segmentation in the enterprise network. However, the latest version adds further context to this journey through the introduction of Analytics and Assurance. You get end-to-end visibility into the network with full context through data and insights.
Why not explore the possibilities that intent-based networking could bring to your business or organization? Our trusted advisors can help you design and implement a software-defined network access (SD-Access) solution that will allow you to configure and provision thousands of network devices across your enterprise in minutes, as well as use advanced assurance and analytics to proactively monitor, troubleshoot, and optimize your network.
The future of intelligent, intent-based networking is fast approaching. Will you be ready for it? Clearpath can help your organization come up with a networking strategy and plan so you'll be ready for a fully intelligent, intent-based network in no time!
1 Enterprise Strategy Group, "The Journey to Intent-based NetworkIng" January 2018


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Stop Predicting the Future — You Can't
A former boss asked me last week what I thought the next big thing in online advertising was going to be. I stopped to think for a few seconds, and then told him that it will be offers delivered to mobile devices, triggered by an ad network, serving creative based on the GPS location of the user's cell phone or PDA.
Although eMarketer recently predicted that the total market for mobile text (SMS) ads will reach $1.5 billion this year, I'm not talking about an offer being delivered as 150 or so characters of a text message. Instead, the promise of something like the iPhone's Safari Web browser on more mobile devices should have marketers really excited.
Then I started to think about how many Internet prophets I've run into over the past few months, all trying to either make a living or get on the speaker circuit predicting the future of the Internet. It's really starting to bug me.
I'll lump these people into the same heap as consultants and analysts. The longer they spend on the outside, the further they are from telling you anything worthwhile. I'm pretty sure that I could drop everything today and pimp myself out to small- to mid-size publishers and give them input on how to make their e-media efforts stronger. In less than a year, I would be out of things to tell them and out of people to tell it to. You need to live and breathe this stuff every day, and even then you sometimes feel you know very little.
Now back to the futurists. Yes, the next logical progression from Web 2.0 is Web 3.0, my 8-year-old daughter could tell me that one. Sure, traditional print advertising is taking a huge hit from online advertising, while paid search is highly measurable and social media sites like Facebook and MySpace may someday be the only way we communicate. Enough already!
Is it me, or are there more and more people analyzing, predicting and consulting rather than doing? All I know is that over the past decade, what I do for a living has changed a lot. If any of you folks can tell me what the next 10 (or even five) years will look like, I'm hoping you have something creative to predict. Otherwise, you may have better luck convincing me my beloved Philadelphia Eagles will win the Super Bowl this season.


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News | January 27, 2020
Irvine School Wins Regional Science Bowl at JPL Third Year in a Row
High school teams had mere seconds to answer complex science and math questions in in the regional round of the National Science Bowl.
Southern California high school students had mere seconds to answer complex science questions that would rattle even the most stalwart rocket scientist in the regional round of the U.S. Department of Energy's National Science Bowl on Jan. 25, 2020.
The team from University High School in Irvine, California, won first place for the third year in a row at the tournament, which has been hosted by NASA's Jet Propulsion Laboratory in Pasadena, California, for almost three decades. They'll advance to the national competition, held in Washington from April 30 to May 4.
Twenty-three teams competed in this year's competition, which has become so popular that schools are chosen via a lottery system.
John Callas, project manager for the Spirit and Opportunity Mars rovers and manager of the NASA-NSF Exoplanet Observational Research program, has volunteered as a Science Bowl moderator for 26 years. "I'm still impressed with how quickly these young minds can answer a very complicated question. A question that I, today, would have to take out a piece of paper and spend maybe five minutes on doing the calculation, they do within seconds in their heads," he said. Needless to say, calculators aren't allowed.
One example of a past tournament's "easy" questions reads: "A famous principle of fluid mechanics, traditionally attributed as the explanation behind lift on an airplane wing, is named after what scientist?"
The team from Santa Monica High School confers during the final rounds of the L.A. regional Science Bowl, before winning second place. Teams were given mere seconds to answer complex science and math questions. Image Credit: NASA/JPL-Caltech
Larger view
Despite the serious brainwork required, the atmosphere was fun and energetic, with even a milk-and-cookie break in the afternoon.
"I love learning science with other people. I love competing, and I think Science Bowl is one of the best competitions out there," said senior Nyle Wong, captain of University High School's winning team. "We feel really good - we worked really hard for this."
In between the round-robin and double-elimination rounds, Callas explained astrophysics and Mars exploration and students went to see the Mars 2020 rover in the Spacecraft Assembly Facility. Some former Science Bowl participants have gone on to work at JPL or Caltech, which manages JPL for NASA.
"These kids are so smart, and it just reminds you that this is the next generation. And if we can help them get to JPL, a lot of them will come work here, which is great. We just try to give back to the community as much as we can," said Nancy Kapell, a JPL human resources specialist who has volunteered for the Science Bowl for 17 years.
While students may be preparing for careers in STEM, some of them do it purely for the love of science.
Sophomore Alissa Kopylova - one of University High's younger team members -dedicated her summer and free time to studying in order to make the team. "I feel like I got interested in biology when I first came to school, and I felt like it was a great way to expand my ability in this area. And also, the subject interests me to a point where I was willing to do something fun," she said.
Silver medals for second place were given to Santa Monica High School and bronze medals for third place went to Troy High School in Fullerton. Culver City High School won for best sportsmanship.
News Media ContactArielle Samuelson
Jet Propulsion Laboratory, Pasadena, Calif.


text: While you may not like to argue with your significant other, it probably happens sometimes. You may know couples who argue often and those who don't. But there are several reasons why arguing may be good for your relationship. In fact, an online study, "Able Arguers," among 976 individuals in 2012 found that couples who engage in healthy conflict are 10 times more likely to have a happy relationshipversus those who ignore difficult conversations. The study was conducted by David Maxfield and Joseph Grenny. Grenny co-authored The New York Times bestseller Crucial Conversations: Tools for Talking When Stakes are High, and Maxfield co-authored follow-up books in the series, all of which were also NYT bestsellers.


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3rd Transit Oriented Development World Summit 2019Posted in Civil Engineering Events | Email This Post |
The Leading Transit Oriented Development World Summit!
While the development of cities has been historically linked to investing in urban regions, the concept of Transit Oriented Development has emerged as a response to traffic congestion and the prevention of urban sprawl. Transit Oriented Development has proven to help integrate modes of transport to save time, dependence on cars and improves physical health. It has also been seen a major solution to the serious growing problems of climate change and global energy security by creating dense, walkable communities that greatly reduce the need for driving and energy consumption, whilst driving urban and economic development.
Equip Global's3rd Transit Oriented Development World Summit 2019taking place in Singapore from 17-20 June 2019returns in June and will gather various Ministries of Transportation, National Development, and Infrastructure, Housing Authorities, Urban Development Authorities, Rail & Transport Operators, Investment firms and City Planners across Asia and globally to discuss their experience and knowledge on successful masterplans for today and tomorrow's Transit Oriented Development Projects around the globe.
For more information, please visit our website or email us at email@example.com now!
Start: 17 June 2019
End: 20 June 2019
Organizer: Equip Global Pte Ltd


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Congratulations to PREPD ambassador Tim Reed on an epic performance taking out 2nd place in IRONMAN in Port Macquarie earlier this month. A PB performance in a time of 8:09:50 – almost 7 minutes faster than when he won in 2016!
After heading into the run almost 10 mins behind Cam Wurf after a blisteringly fast bike leg, Reedy chipped away before reigning in an extra five minutes over the last 10k's. Not quite enough to take out the win but Tim described it as "a clear standout race for me over this distance" and finished with a staggering 2:44 hour marathon!
Just a week later, Reedy backed up with a podium finish in hot and humid conditions in IRONMAN 70.3 Vietnam against a world class field including IRONMAN World Champion Patrick Lange.


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The Toronto Outdoor Art Fair is Canadas largest and longest-running juried outdoor contemporary art fair. This year, the free public event showcases works by over 360 contemporary visual artists and makers, ranging in medium from painting, photography, sculpture and mixed media to jewellery and ceramics.
This year, TOAF brings an array of programming including Art Talks where visitors can catch the exuberant Tiffany Pratt, one of Torontos most recognizable design figures, as she shares her love of art or join a walking Art Tour to meet the talented artists of the fair. The third annual Henderson Brewing Co.s Beer Garden takes over the stage at Nathan Phillips Square while Karen Kaeja of Kaeja dDance premieres Calibrate, a special contemporary dance piece.
If thats not enough, there are art tours, nested exhibitions, adventures in photo darkrooms as well as exciting Power Kids workshops and a special collecting program for our young art lovers to keep attendees inspired and stimulated.
The 57th annual edition runs rain or shine in the square, Jul 6-8, Fri-Sat 10 am-7 pm, Sun 10 am-5 pm. Free.
Location - Nathan Phillips Square


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Guests: Paul Hurley, former editorial page editor of the Visalia Times Delta; Lisa Bryant, a political science professor at California State University Fresno; James Burger, government reporter for the Bakersfield Californian; and John Ellis, political reporter for the Fresno Bee / John Meyers, reporter of the LA Times and Dan Walters, Reporter of the Sacramento Bee. Host: Mark Keppler, Executive Director of The Maddy Institute.
Podcast: Play in new window | Download
Subscribe: Apple Podcasts | Android | RSS


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Previously On….
- 3rd Annual Broadcast/Real Weird Sisters Crossover!
- Christie Runs a Marathon
- The Broadcast Ep. 4.17 "Little Mermaid Madlibs"
- Halloween 2019!
- A Geocache Adventure with Dad & Zach!
- Leslie Odom Jr. in Raleigh!
- NCGeocachers' Fall Fling 2019
- The Joker Isn't a Joker After All (Movie Review)
- The Lizzo Concert!
- Geek. Dad. Life. Ep. 1 is Out!
Tag Archives: Podcast
Vote for MetroBuzz for a Podcast Award!
MetroBuzz has been nominated for a podcast award in the category "Best Comedy." We are going up against some very fierce competition, but you can vote once daily from Nov. 1 through Nov. 15, so if you have a minute … Continue reading
Posted in John Norton, Metrobuzz Tagged Metrobuzz, Podcast, Podcast Awards 2 Comments
An Oldie Jolleen Story
I know that I have told to this story before but it's one of my favorite memories and since it's Podcast Award nomination times once again I figured that I would fondly revisit the past. On October 19, 2008, Jay … Continue reading
Posted in Jay Glatfelter, John Norton, Memories, Metrobuzz, Movies, Podcasting Adventures Tagged Metrobuzz, Movies, Podcast, Podcast Awards, Tequila, The Lost Podcast Leave a comment


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Before you listen to Kylie's new single Whistle there are three things you should know:
1) This is not a single from a new Kylie album - it's from the soundtrack to the movie Jack & Diane (that also features Kylie)
2) It's not a cover of a Flo-Rida song
3) Don't expect Love At First Sight or All The Lovers here - think more Where The Wild Roses Grow but way quirkier thanks to teaming up with the Icelandic experimental band Múm
Ok so we've prepared you now...
This is different - survey most on the street and they'd never guess this was a Kylie track. But we like it. Listening to it, it's more Múm than it is Kylie but she's the overriding vocal in the song. It's sweet, delicate and melodic with an interesting use of vocals and computer generated sounds. Each and every listen you like it more and more. Out now on iTunes to buy, we could go on about it but we think it's best you listen to Whistle yourself..!


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The Lena Blouson Blouse
The Lena Blouson Blouse
Free US shipping and returns
Ships Within 2 Business Days
The Lena Blouson Blouse is the perfect lightweight top for transitioning into warmer weather. With a feminine V neckline, front pleat details and a modern stand collar, this easy fit, long sleeve top is a vacation must-have. Subtle seam detailing enhances slightly puffed shoulders for that added touch you love from Brochu Walker. We adore this shirt with everything…untucked or front-tucked for an effortless, chic look.
Model is 5'10" and is wearing a size Small.
Shown here with: Coombe Pant
- Slight crossover V neck with front pleat
- Easy fit
- Stand collar
- Slight puffed shoulders with seam detail
- Long sleeves with button slit cuffs
- Yoked back with center back pleat
- Straight hem
- 100% Polyester with 100% Silk Trim
- Imported


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Here we are, back at the office. One more day of glorious hooping by the top remaining teams in Illinois high school basketball.
The fans are waking up and trickling in for the third-place 3A game. The town of Lincoln will be here in a while to split rails and ears in support of their team in the title game, against Chicago Morgan Park.
And another day of Bitting around.
Can't wait.


text: I am querying data from another Oracle database, doing some work on the data, geocoding it, then appending it to a feature class in our enterprise geodatabase. I use ArcSDESQLExecute(), then NumPyArrayToTable() to pass into GeocodeAddresses_geocoding(). However, during the NumPyArrayToTable() process, null values always come through as a text string 'None'. I tried changing None values to numpy.NaN but those just come through as a string 'nan'. My datatypes for my numpy array fields are all string. How do you deal with null values with NumPyArrayToTable()?


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Marketers must move past pink, embrace female empowerment
Whether it is razors, headphones, toys or even firearms, the "pink it and shrink it" approach has been implemented in marketing strategies for a wide range of products — not just sports. Dell did it with its pastel-painted Della website and marketing campaign targeted at women. The second-largest manufacturer of PCs attempted to introduce technology as a fashion statement for women by pitching a new lightweight laptop notebook computer stocked with a calorie counter and tips for finding recipes. The audience backlash was immediate, forcing Dell to drop Della within two weeks of its launch in 2009.
Increased national dialogue about gender equality has challenged the long-held notion that such marketing is effective in attracting female consumers. Instead, pro-women ads are being created by some of the world's top brands in an effort to win women's spending through inspiring, motivational, and emotional advertising. These female empowerment ads are not only developed by women's product companies like Always, but are also implemented by sports marketers, historically a boys club.
With women now controlling 70 percent to 80 percent of consumer purchases, according to a 2016 infographic created by UNC Kenan-Flagler Business School's online MBA program MBA@UNC, companies catering to the biggest sports leagues' audiences can no longer afford to ignore the stats. They must instead focus on developing ads that speak to their audience.
Today, pink is a trigger color. It's a lack of creativity, a lack of understanding of women as a market. It's a trigger that makes a company appear as if it believes all women are the same. It's offensive, ineffective, and no longer tolerated.
Many brands make their livelihood selling on the stereotypes of women, and that's not going away, but with the purchasing power of women growing, sports marketers must work harder to understand female sports audiences from a psychographics standpoint or risk their campaigns falling flat. They must engage with their female audiences in a meaningful way that goes beyond coloring something pink.
|A Gatorade ad with flashbacks of Serena Willams (right, with sister Venus) reaches out to female consumers.
Gatorade, for example, features women in advertising campaigns to connect with female consumers. Their most recent series shows flashbacks from Serena Williams' career. The ad builds on Gatorade's "Win from Within" tagline, asking a young Williams who she wants to be like when she grows up. She says she wants people to be like her.
Touch by Alyssa Milano changed the retail game as the only apparel company with licensing agreements for women's clothing with major American sports leagues — MLB, the NFL, NHL, NBA and, more recently, NASCAR and MLS. Tired of having options only in pink for sports apparel, Milano developed a fashion line with the motto: "Where the game meets the after party." The idea was to make Touch fashionable enough for women to wear both in and outside of the sports arena. Milano is now credited with being a trailblazer in the fan-gear industry. As soon as she proved that women wear jerseys away from games and around other women, large sports apparel companies like Nike and Reebok followed with more fashionable lines of women's team wear.
Speaking of Nike, the largest retailer of women's athletic wear launched a campaign called #betterforit aimed at women in 2015. Nike says #betterforit is its largest initiative yet in supporting and motivating women's fitness quests.
In a 2014 survey conducted by SheKnows, 52 percent of the more than 600 women surveyed indicated that they purchased a product based on the marketer's portrayal of women.
The more marketers develop powerful, pro-female campaigns and ads that inform, empower, and avoid borderline offensive advertising practices, the better.
Molly Greenberg (firstname.lastname@example.com) is the community content manager for MBA@UNC, UNC Business School's online MBA for executives. She was named one of Washington's 100 top tech leaders by the Washingtonian in 2015 for her in-depth media coverage of the business of higher education, education policy, education technology, and startups across the D.C., Maryland and Virginia region.


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If you follow along on Instagram, then you will know that two of my girlfriends and I took a little vacation to Isla Holbox at the beginning of the month. Holbox (pronounced "Ole-bosh") is a tiny island about 40 miles northwest of Cancun. It's absolutely beautiful and totally off the grid. The beaches reminded me of Tulum (before it got ruined) and the town reminded me of Sayulita (but cleaner). It made for the perfect remote getaway.
I got so many questions about our trip (especially our hotel + how we got there, etc.) so am going to do my best to answer them all while also sharing some pics from our trip!
Isla Holbox Photo Diary
Where we stayed:
We stayed at Punta Caliza. We LOVED It. The hotel was actually the reason we booked the trip; we'd been stalking it on Instagram for ages! It's a tiny, family-owned boutique hotel (just twelve rooms!) and so beautifully designed. The pool is amazing (as is the bar + restaurant… they had the best ceviche + mezcal margaritas and a really yummy (free!) breakfast in the morning). This place is going to blow up – if it hasn't already. They just were featured in Conde Nast Traveler's Hot Issue as one of the best new hotels in the world so it's getting popular fast and they're small!
Every room has its own entrance to the pool and access to the beach which is wonderful. The beach is about a 5-10 minute walk and Punta Caliza has its own little area with chairs/umbrellas, etc. The water is beautiful and crystal blue and reminds me of how Tulum used to be before it got ruined.
I will note that they do have a no children under 12 policy (sorry). This was a draw for us as we wanted a quiet girls weekend where we could drink margs and read in the sunshine without any little ones running around. I have heard great things about Casa Los Tortugas which sounds more family friendly. I will also say that it seems they raised their rates quite a bit. Our room was about $150 a night (slightly more with taxes). A friend of mine reached out and was quoted $800 USD/night (yikes) so I don't know if it's just a very busy time of year or they've increased their prices. (We didn't receive a media discount or anything; Becca was the one who booked it so they didn't even know about my blog.)
Getting There:
Okay, it's a hike! I would recommend booking transportation through your hotel, which is what we did.
You fly to Cancun (very easy from New York) and then drive (we had a driver) two hours and then take a 30 minute ferry. Our hotel picked us up in a golf cart. For the three of us (total) this cost $185 each way, which includes ferry tickets. We took an 8am flight and weren't settled into our hotel til about 4pm later that day.
What to Do:
Honestly, we didn't do a lot! The town is REALLY cute (it reminded me a lot of Sayulita). Every day we wandered around the town, got juice, shopped a little bit (there are a LOT of cute shops!), and then went to the pool/beach. But we were only there for three days and we were all pretty exhausted after our live show / work stuff so we were happy to just lay low and read in the sun. We were all on a very strict book a day diet!
We heard good things about the bioluminescence tours but didn't do that – maybe next time! You should also rent bikes and explore the island.
I will be honest (and someone is going to rip me apart for saying this but I would want to know this – also know that I'm tough crowd and am always honest) – the food was kinda underwhelming! We kept thinking we were missing something but talked to a few other groups of tourists we met and they felt the same way so we felt a little better. The ceviche at Punta Caliza was amazing – we had it every day as lunch or at least a snack. But what we really wanted was authentic Mexican and had a hard time finding that.
- Mandarina at Casa Tortuga – we went here twice for dinner! The first night we ate at their main restaurant (a mix of seafood and Italian) and on our last night ate on the roof and had sushi.
- Barbra Negra – really good fish tacos.
- Luuma was probably my favorite meal we had and the nicest restaurant we went to. We got the veggie board and a ton of tapas.
There are tons of spots to grab a drink if you walk along the beach.
We also had really great coffees at Clandestino.
What to Know Before You Go:
- There is one ATM on the whole island (or so I'm told, we didn't see it!) and it doesn't always work so take out pesos before you go! I took about $180 in USD for the whole time we were there (three full days/four nights) and it was more than enough although we did eat a lot of ceviche + drink a lot of margs at our hotel. The food is very inexpensive.
- If you love dogs, you are in for a treat. There are SO many cute pups, everywhere! I was dying and wanted to take them all home!
- Most restaurants do not take reservations so get there a little early and plan to grab a drink nearby (there are tons of great little bars around every corner.)
- The Wifi and cell service are NOT great. This is a draw for a lot of people but I wish I knew ahead of time as I would have prepared as if I were going to Cuba – you will be disconnected.
- There are no cars on the island! Everyone gets around on golf carts which is very charming.
- It's so casual. Like, you don't even really need shoes (seriously, all the roads are sand – at dinner a lot of people were shoe-less!)
Would I go back?
I definitely would. I will just say that I am a little torn on if I actually will. It is such a hike to get there, and once you are there it's amazing. But I got restless and was a little bored by the food options. I probably wouldn't want to go for more than three or four days. And I would also make sure I didn't have to work at all while I was there!
So it's a pain to get to for just a few days but if you are the type of person who wants a week long sleepy vacay (which makes the long travel days more worthwhile), this is your spot!!
If you like this"Isla Holbox Photo Diary", also, check out, Cuba Travel Guide and Tulum Travel Guide.


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Do not let an accident ruin your life. Contact my Ordaz Law, APC personal injury office to speak with a premier Chula Vista personal injury lawyer – (619) 550-3617.
We offer free home consultations to your Chula Vista home.
Continue to read and learn about 5-Things To Do after a car accident.
About Ordaz Law, APC – A San Diego Personal Injury Attorney, and his Distinguished Case Results.
We have recovered millions on behalf of accident injury victims.
Five Things You Should Do after You have been Involved in Car Accident
It is important that you remain calm if you are ever in a car accident. It is also important to do the following five things:
- Call the Police;
- Receive Immediate Medical Attention;
- Photograph the Damage to your Vehicle;
- Follow-up with your Medical Care; and
- Call an Experienced and Well Respected Chula Vista Personal Injury Lawyer.
Chula Vista | Free Home Consultation
We offer a free home consultation to your home in Chula Vista.
Chula Vista Personal Injury Lawyer | What to Expect from Your Attorney
- Expect Individual Attention from Your Personal Injury Attorney.
- Expect Aggressive Representation from Your Personal Injury Attorney.
- Expect A Strong Expertise and a Good Reputation from Your Personal Injury Attorney.
Blog: Chula Vista Personal Injury
Ordaz Law, APC | Chula Vista Personal Injury Lawyer
Juan J. Ordaz Jr. provides candid, hardworking and personal legal representation to individuals seeking a Chula Vista personal injury lawyer. We help victims suffering from a variety of injuries, who have sustained their injuries through different types of injury producing events including the following:
- Personal Injury;
- Car Accidents / Automobile Accidents;
- Trucking Accidents;
- Motorcycle Accidents;
- Bicycle Accidents;
- Pedestrian Accidents
- Premises Liability;
- Products Liability;
- Aviation Accidents;
- Government Claims; and
- Indian Tort Law.
We believe that it is a necessity to represent people who have sustained traumatic and debilitating injuries, and suffered ultimate losses.
Call (619) 550-3617 today so that we may schedule your free and discreet consultation with a premier Chula Vista personal injury lawyer.
We are for Justice no Matter Who it's for or Against.
Learn More:
What is the Statute of Limitations for a Personal Injury Action?
What Should You Expect from Your Personal Injury Attorney?
What Should You Expect from Your Injury Lawsuit?
Will My Personal Injury Case Go To Trial or Be Settled?
Chula Vista | Personal Injury Lawyer
Blog: Personal Injury
Blog: Chula Vista Personal Injury
Chula Vista Car Accident | Personal Injury


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Oh hey there hey!
I'm Kathryn, and you've found Worn Slap Out, a Southern food blog dedicated to quick, healthy meals and indulgent desserts that are designed to be simple, kid-friendly (or easily adapted) and can be made while getting poked in the eye or tripping on small legos.
I'm fixin' to tell you a little bit more about me and WSO.
First, I love food.
This probably goes without saying since this is a food blog, but I've always had a passion for food. And good food. Maybe it was my dad imparting some wisdom when I was very, very young — "Kathryn, never get separated from your lunch." Maybe it was me seeing other pre-professional ballet dancers eating carrots and celery when I chowed down on a hearty sandwich and carrot cake thinking, "WTF?"
Regardless, I've always had a healthy obsession with food. I love creating my own recipes, experimenting with existing ones, and making food look pretty (because, let's face it — so much of food is in the appearance and presentation). I'm 100% self-taught, though I've had the great fortune of taking cooking classes with some of the best chefs in Washington, DC.
I believe food is so much more than the actual ingredients used to cook or bake it. It activates all your senses. Those smells, the way the food tastes on your tongue, the people you're sharing the food with, the topics of conversation you're having while eating — all sear special memories into your brain. These memories are often much sweeter than the food that was consumed.
I'm worn slap out but full of joy.
Hence, the name of this blog. Being "worn slap out" is Southern for totally exhausted. I've always enjoyed staying active, but this lifestyle took a whole new meaning with kids. The greatest joys of my life are my husband of 8 years, son (4) and daughter (2). I keep busy chasing kids and working full-time as a staff development teacher.
What you'll find on WSO
We don't subscribe to a particular diet. In fact, I don't believe in the whole diet concept. I think eating healthy is a habit, and we make choices eery day about what we put in our body. I try to fill our bodies with clean, whole foods. I shop at Whole Foods way too much and buy organic when possible. Baking is my favorite though, and when I bake…I BAKE. You'll find very few "healthy" desserts here. I'm a firm believer in moderation.
On the website, you can find recipes that are generally:
- healthy (hello, stuffed peppers!)
- quick (30 minutes or less) OR can be made in advance and last a couple of nights
- not full of fancy ingredients you can't buy at your neighborhood grocery store
Don't forget about the dessert.
Is a meal really complete without dessert anyway?
Some of my favorite desserts are those with chocolate: Double Chocolate Bundt Cake with Chocolate Ganache, Classic Chocolate Mousse, or these delicious Chocolate Pots de Creme modeled after the ones from Le Diplomate.
Other fun facts about me:
- I'm a Georgia Bulldog through and through (Go Dawgs!) and you'll find us cheering the Dawgs on every Saturday in the fall (and hopefully beyond!).
- I'm always a little jealous when people have those special biscuits their grandmother used to make that bring them right back to being in her house or Aunt Sue's collard greens.
- I can't turn down a good chocolate chip cookie.
Through the exhaustion, there's truly nobody else I'd rather be in this world than me.
Thanks for reading!
All photo credits of our family go to Aaron Taylor Photography.
Amazon Disclosure
Worn Slap Out is a participant in the Amazon Services LLC Associates Program, an affiliate advertising program designed to provide a means for sites to earn advertising fees by advertising and linking to amazon.com. This comes at no cost to you.


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Giordano Bellincampi | Conductor
Ingrid Fliter | Piano
Leonie Holmes | New work
Schumann | Piano Concerto
Tchaikovsky | Symphony No.4
Tchaikovsky's Fourth Symphony opens with a stark fanfare which, Tchaikovsky said, represented 'Fate, that ominous power', adding 'There is no alternative but to submit to Fate'. His friend and patron, Mme von Meck, heard 'profound, terrifying despair' in this music. True, but it has also passion, drama and ultimately catharsis.
The superb pianist Ingrid Fliter returns with Schumann's ineffably poetic concerto, and to open the concert, a new work from the multifaceted Auckland composer, and former APO composer-in-residence, Leonie Holmes.
Start time
November 12, 2020 8:00PM
Auckland Town Hall


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Some observers are comparing the U.S. today to the Gilded Age at the end of the 19th century, a time during which a relatively small number of financiers and entrepreneurs — the so-called Robber Barons — amassed enormous fortunes and the gap between the haves and have-nots became a yawning chasm. The Progressive Movement arose slowly during the last decade of that century and then gained national momentum during the years between Theodore Roosevelt's ascendancy to the Presidency and the election of Warren G. Harding as President in 1920. A central aim of the Progressive Movement was to address the excesses of the Gilded Age and to bring a greater sense of equity to national life. We are living through a similar period of great income and wealth inequality. The lecture will identify how the Progressives grappled with the issue in their time — for example through establishment women's suffrage, the graduated income tax, and the direct election of the Senate — and what we can learn from their experience and apply to today's problems.
Steven Sinding was the director general of the International Planned Parenthood Federation in London. He had a 20-year career at the U.S. Agency for International Development and served as senior population advisor to the World Bank and later as director of the Population Sciences program at the Rockefeller Foundation. He was clinical professor of public health at Columbia University. Dr. Sinding serves on a number of boards and works as an international consultant.


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Located 15 kilometers east of Antalya, one of the largest cities in Turkey, in the region of Pamphylia, is a beautiful resort town called Side, which is regarded as one of the renowned classical sites in the Mediterranean Turkey. If you want to experience the perfect harmony of history and nature, along with world class tourist attractions, then visiting Side should definitely be included in your itinerary.
Roman ruins
- Details
- City: Side
Roman ruins of Side that are in fairly good condition include among others a quite large amphitheatre, a temple (to Apollo), and a gate. There are plenty of ruins are remaining from the Roman Empire. ...


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Embedded A.I. Solutions company featuring the NeuroMem Smart® technology of General Vision Inc. Enabling custom design of edge computing modules with application development.
UAB Neurotechnologijos offers an NТ Industrial Automation system for solving problems of recognition of signals and images. The NT Industrial Automation hardware is composed of Adaptive Controllers each featuring NeuroMem networks for the learning and classification of patterns. The software part consists in specialised software for the retrieval of data from any MES system.


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Tagged With: Lithuania
What else to do in Lithuania? Go to Latvia!
Since the country of Lithuania is fairly small compared to a lot of other bigger countries, Lithuania is in perfect location to drive hard and fast into Latvia. If you got a rental car, then you might as well as drive like you stole it! Just kidding. Tip: There are a lot of speed cameras … Continue reading
What to buy in Lithuania – Wine, Amber, and Basketball Jerseys
So what to buy in Lithuania? Plenty of stuff, actually. Note that these people don't really bargain so everything pretty much has a price tag to it. So don't try to haggle or negotiate a lower price. That might work if you are buying over 1000 euros worth of stuff and it's at an outside … Continue reading
Underrated Lithuania – Hill of Crosses, Trakai Castle, KGB Museum
I know I have not been diligent in writing which resulted in this website being off for quite some time. But this was due to the lack of traveling and enticing adventuring to write about on my part. However, the travel gods have struck lightning into me once again and I have started to rack … Continue reading


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Home for the Holidays
How can you help the less fortunate in our community move out of the cycle of homelessness? Director of Development reflects on the reason for the season.
Director of Development Marty Sanders reminds us to be thankful and help those in need this holiday
Serving Veterans at the Center of Hope
Director of Development Marty Sanders writes about how we serve our homeless veterans by providing temporary shelter at the Salvation Army Center of Hope