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An Afternoon at the Met- Finding Peace in the Middle of NYC
I was in NYC all last week working on Picha, the new startup.
The week was full of all kinds of techie goodness: hanging out at the mothership (aka Babble HQ), attending Latino2 at Google NY HQ (here are the pictures) and attending my first NY Tech Meetup.
By Friday my head was spinning, I was eating Reese's cups and Snickers bars constantly (always a sign I need a nap) and I was so homesick for home-sweet-chaotic-home that I wanted to start walking to Nashville.
What does a gal do when she is heartsick, overtired and has a free afternoon in Manhattan?
She (ahem, actually it was Megan and I) goes to the Met, of course! Instant inspiration, a sense of wonder at the beauty of many of the pieces and an overwhelming feeling of awe that everything else went on hold for a few hours. (Even my feet magically quit hurting for a few hours.)
Here is a look through my lens. I hope you enjoy…
Read more from me at Alli Worthington .
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Don't miss the latest from Babble Voices Like Us on Facebook!
More of me on This is Alli.

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Proper repair of DNA double strand breaks (DSBs) is vital for the preservation of genomic integrity. There are two main pathways that repair DSBs, Homologous recombination (HR) and Non-homologous end-joining (NHEJ). HR is restricted to the S and G2 phases of the cell cycle due to the requirement for the sister chromatid as a template, while NHEJ is active throughout the cell cycle and does not rely on a template. The balance between both pathways is essential for genome stability and numerous assays have been developed to measure the efficiency of the two pathways. Several proteins are known to affect the balance between HR and NHEJ and the complexity of the break also plays a role. In this review we describe several repair assays to determine the efficiencies of both pathways. We discuss how disturbance of the balance between HR and NHEJ can lead to disease, but also how it can be exploited for cancer treatment.
DSB repair; HR; NHEJ; DNA repair assays; PARP inhibitorsIntroduction
Genomic integrity and faithful replication are essential to prevent mutations and chromosomal rearrangements, which may otherwise lead to diseases and in some cases even death. DNA damage is generated by several different genotoxic agents such as reactive oxygen species, UV light from the sun and mutagenic chemicals [1]. These agents cause many types of DNA damage, ranging from base damage to double strand breaks (DSBs). To protect the genome from the deleterious effects of these lesions, several mechanisms have evolved that detect and repair DNA damage. Together with mechanisms that regulate cell cycle progression and cell death pathways this is known as the DNA damage response (DDR).
In this review we concentrate on DSBs, which are among the most cytotoxic types of DNA damage. The therapeutic effect of several commonly used cancer treatment modalities, such as ionizing radiation and the chemotherapeutic doxorubicin, are based on the cell-killing effect of DSBs. However, DSBs are also the initiating lesion of disease-causing chromosomal translocations in cancer. Therefore, it is important to understand the intricate regulation of the DDR upon DSB formation. We mainly concentrate on the two main DSB repair pathways, Non-homologous end joining (NHEJ) and Homologous recombination (HR), with a special emphasis on the balance between both repair mechanisms in health and disease.
NHEJ is a relatively simple DSB repair pathway (Figure 1). Both ends of the break are first bound by the Ku70/Ku80 heterodimer, which then recruits the catalytic subunit of the DNA dependent protein kinase (DNA-PKcs) [2]. If necessary, the ends can be trimmed by nucleases (such as Artemis) or filled in by DNA polymerases (such as Polμ or Polλ) to create compatible ends [3]. Finally, the ligation complex, consisting of DNA ligase IV, X-ray cross-complementation group 4 (XRCC4) and Xrcc4 like factor (XLF)/Cernunnos ligates the ends [4,5]. NHEJ can take place throughout the cell cycle. For an extensive review on NHEJ see [3].
Figure 1. HR and NHEJ. NHEJ) NHEJ starts with recognition of the DNA ends by the Ku70/80 heterodimer, which recruits DNA-PKcs. If the ends are incompatible, nucleases such as Artemis can trim the ends. The XRCC4-DNA Ligase IV-XLF ligation complex seals the break. HR) The MRN-CtIP-complex starts resection on the breaks to generate single stranded DNA (ssDNA). After resection the break can no longer be repaired by NHEJ. The ssDNA is first coated by RPA, which is subsequently replaced by Rad51 with the help of BRCA2. These Rad51 nucleoprotein filaments mediate strand invasion on the homologous template. Extension of the D-loop and capture of the second end lead to repair.
HR uses a sequence similar or identical to the broken DNA as a template for accurate repair. The sister chromatid is used as an identical template in the S and G2 phases of the cell cycle, when the DNA has been replicated. HR is restricted to these cell cycle phases in higher eukaryotes to prevent recombination between (repetitive) non-identical sequences. Spurious HR can lead to loss of heterozygosity (when HR takes place between paternal and maternal chromosomes) or insertions/deletion (when repeats are not aligned properly).
The HR pathway starts with resection of the broken DNA ends (Figure 1) by the MRN-complex [6,7], together with CtBP-interacting protein (CtIP) [8,9] and other exonucleases, generating 3'-single stranded DNA (ssDNA) [9,10]. The ssDNA tail is coated by Replication protein A (RPA) to remove secondary structure [11]. Subsequently, BRCA2 mediates the replacement of RPA by RAD51, to form a nucleoprotein filament that searches for the homologous sequence on the sister chromatid. After strand invasion, catalyzed by RAD51 and many other proteins, the DNA end is extended using the intact sequence as a template. After restoration of any lost sequence information, the second end of the broken DNA is captured and the junctions are resolved to give a precisely repaired DSB [12]. This resolution step can be accomplished via formation of two Holiday junctions, which are subsequently resolved to give crossover or non-crossover products (the double Holiday junction model). An alternative HR model, the synthesis dependent strand annealing (SDSA) model, does not involve Holiday junctions and results in non-crossover products only [13].
Microscopically, DSBs can be visualized as local spots of repair protein accumulation (also called foci) in the nucleus. For example, histone H2AX is phosphorylated locally around the DSB and 53BP1, RPA and RAD51 accumulate in foci after ionizing radiation. Changes in the number of foci per nucleus in time can be quantified to analyze the dynamics of DNA repair [14]. Not all repair proteins accumulate in sufficient numbers to form foci. For example Ku70/80 does not form foci, although it is recruited to DNA damage [15,16].
Alternative DSB repair pathways
In addition to classical HR, several subpathways result in slightly different products. For example the single strand annealing (SSA) pathway uses directly repeated stretches of homology to repair DSBs. After resection of the break (as described above for HR) complementary stretches in the ssDNA anneal and the intervening sequence and one of the repeats is deleted [17]. Since HR and SSA use the same substrate, these pathways compete when repeats are present on both sides of the break and SSA should be suppressed to prevent its mutagenic effect.
Alternative end-joining pathways can also join DSBs in an error-prone manner, especially when classical NHEJ is impaired by deletion of essential components. The genetics of this pathway are not well defined and there may even be several alternative end-joining pathways. A dependence on DNA ligase III, Xrcc1 and PARP1 has been found in genetic assays [18,19]. However, in another assay the repair of I-SceI induced DSBs in XRCC4-deficient pro-B cell lines did not require Xrcc1 [20]. Alternative pathways show increased DSB joining using microhomologies (stretches of 1–6 bp of direct repeat at the junction), possibly to stabilize the synapsed ends [3].
Although these alternative DSB repair pathways can work in specific experimental settings, they probably do not play a major role in repair of most DSBs in wild type cells. Therefore this review will focus on the balance between the classical forms of HR and NHEJ.
Repair assays
To study the balance between HR and NHEJ, one would ideally measure both types of repair at the same time using a defined chromosomal site. Unfortunately such an assay is not yet available. There are, however, many assays to measure HR and NHEJ separately. A good understanding of these assays is indispensable for correct interpretation of the results obtained using these different approaches. We therefore review the major assay systems and discuss their merits and drawbacks.
Assays to measure NHEJ
NHEJ can be measured in many different ways. The simplest version is transfection of linearized DNA into wild type and mutant cells. Joining of the ends can be monitored by cloning out individual plasmids or PCR amplification followed by sequencing or digestion of the junction [21] (Figure 2a). Recircularization can also be monitored by following the restoration of expression of a reporter gene, such as an antibiotic resistance gene or a fluorescent marker (Figure 2b). A major disadvantage of these assays is that the linear DNA is extrachromosomal and NHEJ cannot be measured in the normal context of chromatin. However, it is a simple assay that can monitor decreased activity of the core NHEJ machinery as a shift towards microhomology use at the newly formed junctions.
Figure 2. NHEJ repair assays. a) Linear plasmid DNA with 6 bp repeats at the ends is joined after transfection. The joints are amplified by PCR and digested using BstXI to distinguish between direct repair and microhomology mediated repair [21]. b) Repair of linearized plasmid DNA results in restoration of GFP expression. c) Cleavage by I-SceI and subsequent repair lead to loss of the middle splice donor and acceptor sites (SD and SA) and the adenoviral exon (AD), resulting in the expression of active GFP [22]. d) H2Kd fused to CD8 is expressed from the intact substrate. Repair of the oppositely oriented I-SceI breaks results in loss of H2Kd-CD8 and allows expression of CD4 [23]. e) Similar to d), the intact substrate expresses GFP, while the repaired substrate allows expression of RFP and loses GFP expression [24]. f) Between the opposite I-SceI sites, a translation start site is located, preventing translation of the XHATM resistance gene. Repair of the I-SceI breaks and loss of the intervening ATG results in XHATM resistance. The sequence around the breaks can be sequenced to monitor loss of nucleotides [25]. g) V(D)J recombination assay. Cleavage by the Rag1/2 endonuclease at the recombination signal sequences induces inversion of the intervening sequence. Small arrows indicate location of PCR primers to amplify joints [21].
Another type of NHEJ assays uses two I-SceI restriction sites. These sites can be in the same or opposite orientation, generating compatible or incompatible ends, respectively. These constructs are generally integrated into the genome. The general theme of all these assays is restoration of expression of a marker gene, in some cases accompanied by inactivation of another gene. Mao et al. (Figure 2c) interrupted the GFP gene with an intron containing an adenoviral exon (AD) flanked by two I-SceI sites. Repair of the two I-SceI induced DSBs leads to loss of the intervening exon and expression of functional GFP [22]. Guirouilh-Barbat et al. developed a similar assay (Figure 2d) with compatible or incompatible I-SceI sites, but they used surface antigens as a read-out for repair [23]. Coleman and Greenberg also used a comparable assay (Figure 2e) with GFP between the I-SceI sites and RFP downstream, resulting in loss of GFP and expression of RFP after repair of the I-SceI induced DSBs [24]. In these assays with a double I-SceI site it is also possible to sequence the joints and to determine the loss of nucleotides around the breaks (Figure 2f) [25].
A disadvantage of these assays using I-SceI restriction sites is that the individual I-SceI break has compatible ends and can recreate an I-SceI site if it is repaired precisely by NHEJ. Therefore, several cycles of cleavage and repair can happen before the site is lost due to inaccurate repair and these assays cannot measure the NHEJ efficiency accurately. However, sequencing of the junctions can provide interesting information about imprecise end-joining events. To avoid the cut-and-paste cycle problem of the I-SceI sites, some assays use transposon excision to create a break. Repair of transposon-induced DSBs can reveal details of efficiency as well as precision of NHEJ [26,27]. In principle, transposons would also be useful to study HR, although their DSB formation efficiency is generally lower than endonucleases.
The immune system depends on end-joining for V(D)J and class switch recombination (CSR). Pan-Hammarstrom and colleagues studied CSR by PCR amplification and sequencing of the junctions in normal individuals and patients. They found that patients with mutations in NHEJ components, such as DNA ligase IV, showed an increased dependence on longer microhomology stretches at the junctions [28]. An advantage of this assay is that repair is measured on endogenous substrates. However, it is not clear whether these loci are representative for other types of DSBs.
V(D)J recombination also depends on NHEJ factors to repair the breaks induced by the Rag1 and Rag2 proteins. This type of repair can be assayed using a specific repair substrate containing Recombination Signal Sequences, the recognition sites for Rag1 and Rag2 (Figure 2g) [21]. The V(D)J recombination assay gives a clear phenotype for defects in proteins involved in DNA end-processing, such as the Artemis nuclease [29]. The major disadvantage of these types of assays is the special nature of the DSBs formed by the RAG proteins, which may shuttle the breaks towards NHEJ [30].
Assays to measure HR
The most commonly used assay to measure HR is the DR-GFP assay developed by Pierce and Jasin [31] (Figure 3). The reporter construct can be inserted by gene targeting or random integration. It contains two GFP sequences separated by a selection marker. The 5' GFP sequence is inactivated by an I-SceI site and internal stop codons, preventing GFP expression. The 3' truncated GFP serves as a template for repair after DSB induction by I-SceI. Repair of the break by gene conversion using the downstream GFP sequence leads to restoration of the GFP gene and the percentage of GFP expressing cells can be determined by FACS analysis.
Figure 3. HR Assay. The 5' GFP is inactivated by several in frame stop codons and contains an I-SceI site. A downstream truncated GFP, lacking the I-SceI sites and stops, serves as a template. Accurate repair via HR results in GFP expression.
This HR assay has been used successfully to characterize defects in various (repair) mutant genetic backgrounds. An important advantage of this HR assay is that it measures repair using a chromatinized reporter construct in the chromosome. However, the template for repair is downstream of the break, whereas the normal template for HR is the equivalent position on the sister chromatid. Furthermore, the I-SceI site can be subject to several cycles of cleavage and repair by precise NHEJ or restoration of the sequence using the sister chromatid as a repair template, which leaves a high degree of uncertainty about the relative levels of HR and NHEJ.
Expression of I-SceI is usually induced by transfection of an expression plasmid into an asynchronously growing cell population. This creates DSBs in the reporter substrate throughout the cell cycle, whereas HR only takes place in the S and G2 phases. To overcome this problem, Hartlerode et al. developed an I-SceI fusion protein that is drug-activatable. Enriching cells in a certain phase of the cell cycle then allows restricted activation of I-SceI [32].
HR can also be estimated by scoring sister chromatid exchanges (SCEs). In this assays a nucleotide analog is added in the first cell cycle to allow incorporation into the newly synthesized strand in S phase. After a second replication round, only one of the sister chromatids is labeled, which allows visualization of recombination between the sister chromatids in metaphase spreads by staining for the incorporated nucleotide analog. SCEs can be formed in S phase during the repair of collapsed replication forks as well as in G2 phase at two-ended DSBs [33].
A completely different method to assess the efficiency of DSB repair is monitoring the disappearance of γH2AX foci. These foci form within a few minutes after DSB formation and disappear slowly as repair takes place. By comparing the kinetics of several known HR and NHEJ mutants, the efficiency and likely repair pathway can be determined. For a review on the advantages and potential pitfalls of this assay, see [34].
As a more sophisticated approach, the formation and disappearance of 53BP1-YFP and Rad52-Cherry foci has been followed to estimate the use of HR and NHEJ in single cells throughout the cell cycle [35]. Karanam et al. found that there is a gradual increase in HR at the beginning of S phase. The number of Rad52 foci increases till mid S phase and then decreases towards the end of S phase. In G2, very few Rad52 foci were observed, showing that HR is not the predominant pathway in G2. This is consistent with data from Beucher et al., who demonstrated that NHEJ repairs approximately 85% of all IR-induced DSBs in G2, as measured by γH2AX foci kinetics [36].
Balancing HR and NHEJ
The presence of large numbers of highly repetitive sequences in the DNA of higher eukaryotes makes HR between sequences other than sister chromatids prone to misalignment of the homologous sequences. Therefore, HR generally dominates in organisms with a small genome (with low abundance of repetitive sequences), whereas mammals mainly rely on NHEJ for DSB repair [35,37]. However, even in highly complex genomes, HR is used as the preferred DSB repair mechanism to deal with DSBs that are formed during replication. This necessitates intricate control mechanisms to prevent access of the wrong repair pathway to the DSB.
Resection & cell cycle
HR can only safely be used to repair breaks in the S and G2 phases of the cell cycle. The first mechanism to regulate this depends on S/G2 specific cyclin dependent kinases (CDKs). DNA end resection requires phosphorylation of CtIP on a CDK consensus sequence [9,38,39]. Proteasome-mediated degradation of the CtIP protein in G1 [40] adds an additional layer of regulation at the resection step.
CDK1/CyclinB also phosphorylates the NBS1 component of the MRN complex on Serine 432 during the S, G2 and M phases, which is required for resection and efficient HR. However, IR sensitivity was not affected in the Ser432Ala NBS1 mutant, consistent with the notion that NHEJ is the major DSB repair pathway in mammals [41].
Although activation of HR proteins in a cell cycle dependent manner helps to restrict their activity, it is insufficient to ensure safe use of HR. While replication is ongoing in the S phase, parts of the genome have not yet been replicated and recombination of these parts should be avoided to prevent loss of heterozygosity and non-allelic recombination. Therefore, another layer of regulation is provided by the structural maintenance of chromosomes (SMC) proteins such as Cohesin, Condensin and SMC5/6: they are able to confine repair to the sister chromatid and prevent HR between other sequences [42,43].
Complexity of the break
Whether HR or NHEJ is used also depends on DSB complexity. This phenomenon has been studied in detail in the G2 phase of the cell cycle, when both HR and NHEJ contribute to DSB repair. Treatment of cells with the topoisomerase II inhibitor Etoposide results in breaks with a 4 bp 5'-overhang with covalently attached protein [44]. The large majority of these breaks are repaired rapidly by NHEJ. The remaining 10% of the Etoposide induced breaks is repaired with slow kinetics via HR [45]. High linear energy transfer (LET) carbon ions, on the other hand, induce highly complex clusters of DSBs and other types of DNA damage [46], because this type of radiation causes a high number of ionizations in a small volume. These breaks are frequently resected and their repair takes place via HR with slow kinetics [45]. From the breaks induced by low LET ionizing irradiation (IR), which causes less complex DSBs, only 20-30% is resected and their repair is much less dependent on HR [36,45].
The chromatin structure around the DSB affects repair as well. Breaks in heterochromatin are repaired more slowly than breaks in euchromatin [47] probably because euchromatin is more easily accessible for repair and requires less or no remodeling. Repair of breaks in heterochromatin requires ATM [47]. ATM phosphorylates transcriptional corepressor Krüppel-associated box (KRAB)-associated protein (KAP)-1 [48], which disrupts the interaction between Kap-1 and CHD3 [49]. CHD3 is an ATP-dependent nucleosome remodeling enzyme and its dispersion allows chromatin relaxation, facilitating DSB repair in heterochromatin. Furthermore, the ATP-dependent chromatin remodeler SMARCAD1 can also be recruited to sites of DNA damage where it facilitates resection [50].
Genetic factors influencing pathway choice
The core HR and NHEJ machineries have been conserved from yeast to mammals [51]. However, several genes have been added to optimize or regulate both pathways in higher eukaryotes. For example, NHEJ has acquired DNA-PKCS and HR added several RAD51 paralogs. Furthermore, several additional genes in higher eukaryotes regulate DSB repair pathway choice without direct participation in the catalytic steps of the repair reaction.
The p53 binding protein 1 (53BP1) is recruited to DSBs, where it has functions in cell cycle checkpoint maintenance and double strand break repair [52]. The fast phase in DSB repair is normal in the absence of 53BP1, but repair of breaks in heterochromatin is severely impaired, probably as a result of impaired Kap1 phosphorylation [49,53].
A deeper understanding of the 53BP1 function has been gained from studies in the immune system. During class switch recombination, highly repetitive DNA segments are recombined to generate the different classes of antibodies. DSBs generated during this recombination reaction can be repaired via NHEJ or alternative end joining. In the absence of 53BP1, resection increases and microhomology mediated alternative end-joining takes over from classical NHEJ [54]. In V(D)J recombination, Variable (V), Diversity (D) and Joining (J) segments are recombined to create a large variety of functional coding sequences for immunoglobulins and T-cell receptors. DSBs created by Rag1/2 are repaired via NHEJ. 53BP1 prevents extensive degradation and it promotes synapsis of DNA ends and stabilizes long-range interactions, not only between breaks created during V(D)J recombination [55], but also between deprotected telomeres [56].
BRCA1 and associated protein
In contrast to the NHEJ promoting effect of 53BP1, the tumor suppressor BRCA1 is required for efficient HR [57] and formation of Rad51 foci after DSB induction [58]. BRCA1 is an E3-ubiquitin ligase that forms a complex with the E2 enzyme BARD1 via its RING domain. This interaction is required for the ligase activity, as well as protein stability and nuclear localization [59]. Although several RING domain mutations have been found in patients, it is currently unknown how the HR defect is related to the E3-ligase function and BARD1 interaction. Drost et al. recently showed that the ring domain is necessary for tumor suppression, but not required for the development of resistance to chemotherapeutics. Tumors with a C61G mutation in the RING domain rapidly develop resistance to platinum drugs and the PARP inhibitor Olaparib, while retaining this mutation [60].
In addition to its function as a ubiquitin ligase, BRCA1 may also function as a scaffold protein that associates with many interaction partners, such as Abraxas, BACH1 and BRCA2/PALB2 [59]. For efficient resection of DNA ends, its interaction with CtIP and the MRN complex is probably important [39,61]. BRCA1 also interacts with RAP80 and the BRCA1/RAP80 complex is recruited to ubiquitylated chromatin around DSBs [62-64]. In contrast to the BRCA1 interactions described above, the RAP80-BRCA1 interaction decreased HR: depletion of Rap80 stimulated recruitment of CtIP and Mre11 and thereby resection [24]. The BRCA1 interactions with CtIP and RAP80 are mutually exclusive, indicating that competition for this BRCA1 binding site affects resection and thereby pathway choice. For replication-associated breaks, BRCA1 clearly tips the balance towards HR.
Genetic interactions of BRCA1 and 53BP1
Recently, some unexpected genetic interactions between BRCA1 and 53BP1 shed new light on their function in balancing DSB repair pathways. Deletion of BRCA1 causes embryonic lethality, but this can be rescued by deletion of 53BP1 [65]. Unexpectedly, deletion of 53BP1 also restored HR and Rad51 foci formation in BRCA1 deficient cells [66,67], implying that both factors influence HR in opposite directions and that inactivation of both genes largely restores the balance. Inactivation of 53BP1 in BRCA1−/− ES cells led to an increase in both nucleolytic DNA end processing and RPA phosphorylation [67]. ATM inhibition in BRCA1 −/− 53BP1−/− cells reduced RPA phosphorylation and Rad51 foci formation, indicating that ATM-dependent resection allows partial restoration of HR [67].
Figure 4 presents a plausible model to accommodate these findings. One-ended DSBs that are formed during replication require BRCA1 to stimulate resection. In BRCA1 deficient cells, 53BP1 prevents resection of DNA ends, leading to aberrant diversion of breaks to NHEJ. This creates dead-end products (if only one DNA end is present) or inappropriate joining to distant sequences causing chromosomal translocations (if a DNA end combines with an unrelated other DNA end). Two-ended DSBs, on the other hand, require 53BP1 to limit resection and allow efficient NHEJ. Overactive resection in 53BP1−/− cells may result in aberrant HR reactions (such as single-strand annealing) or alternative end-joining pathways, creating microhomology-mediated translocations and/or junctions with excessive deletions [68,69].
Figure 4. BRCA1 and 53BP1 in DSB repair. a) Repair of replication associated breaks requires HR. 53BP1 blocks resection of the one-ended break in BRCA1 deficient cells, preventing repair via HR. The breaks are either left unrepaired or repaired via NHEJ using other random DNA ends, which leads to chromosomal rearrangements and genomic instability. In the absence of 53BP1, resection of the DNA ends can take place, allowing faithful repair via HR. b) IR induced two ended DSBs are mainly repaired via NHEJ, however part of the breaks is repaired via HR or alternative end-joining (alt-EJ). Repair via HR or alt-EJ increases when classical NHEJ is impaired by a mutation in one of the core NHEJ genes or 53BP1.
Further insight into the role of BRCA1 and 53BP1 in repair pathway choice was recently obtained using super resolution microscopy of IR induced foci (IRIF). The core of the focus contained mainly 53BP1 molecules in the G1 phase of the cell cycle, probably representing repair via NHEJ. In S phase, however, the core of the IRIF was filled with BRCA1 and 53BP1 formed a ring around this core, suggesting that BRCA1 physically excludes 53BP1 from the break to allow repair via HR [70].
BRCA1 deficient cells are exquisitely sensitive to PARP inhibitors, which inhibit single strand break repair [71,72]. The rationale for this observation is that replication of DNA with single strand breaks results in formation of single DNA ends, which require HR for their repair (Figure 4). As described above, deletion of 53BP1 in BRCA1 deficient cells rescues embryonic lethality. However, loss of 53BP1 also leads to resistance to PARP inhibition [66,73]. In the BRCA1-deficient cells that have also lost 53BP1, the number of chromosome and chromatid breaks is decreased and checkpoint activation is diminished compared to cells that are only BRCA1 deficient [66], suggesting that the regained HR capacity in these cells is largely sufficient to restore genomic stability. A subset BRCA1 and BRCA2 mutant tumors shows loss of 53BP1, indicating that therapy resistance via loss of 53BP1 may be clinically relevant [66].
Ubiquitylation and sumoylation
Ubiquitin and the small ubiquitin-like modifier (SUMO) are small polypeptides that can be attached to proteins as a posttranslational modification. After activation of ubiquitin or SUMO by an E1 enzyme, they are transferred to an E2 conjugating enzyme. With the help of an ubiquitin (or SUMO) ligase (E3) the modification is attached to the substrate. Deubiquitylating enzymes (DUBs) can reverse the ubiquitin modification.
Many proteins involved in the DDR can be ubiquitylated or sumoylated [2,74-76]. For the sake of simplicity, we will focus on one part of the DDR signaling cascade as an example. Upon DSB formation, histone H2AX is phosphorylated by ATM or DNA-PK. MDC1 is recruited to this phosphorylated histone (γH2AX) and is in turn phosphorylated by ATM. This attracts the E3 ligase RNF8 which ubiquitylates H2A and H2AX. Subsequent action of the E3 ligase RNF168 leads to more extensive ubiquitylation of the chromatin around the break, creating a recruitment platform for many other repair proteins, including 53BP1 and BRCA1 [77]. These ubiquitylation events are also required for phospho-KAP-1 foci formation and thereby chromatin relaxation at sites of damage [53].
In addition to an effect on recruitment of repair proteins, ubiquitylation can also affect release of proteins from the lesion. The transient binding of Ku at DNA ends affects pathway choice. Ku binds in all phases of the cell cycle and must be removed to allow resection [78,79]. This removal can be facilitated via ubiquitlyation of Ku by the E3 ligase RNF8 and an unknown E2 conjugating enzyme, leading to proteasome-dependent Ku degradation [80]. Since Ubiquitylation is a very abundant modification on DDR proteins, it is likely that more modifications affecting pathway choice will be discovered in the future.
Concluding remarks
A unifying model for DSB repair pathway choice should take into account that NHEJ is relatively fast, while resection is a slow process that probably creates a point of no return. Therefore, it is to be expected that NHEJ initially tries to repair all DSBs and only if this repair pathway fails to repair the lesion, the chance that resection takes place increases over time, necessitating repair via HR. This is consistent with the observation that the binding of the Ku heterodimer to DNA ends is a very fast process, but the assembly of end-joining complexes is dynamic and may in the long run give way to proteins mediating resection if they are active [16]. This means that initiation of HR will mainly be restricted to the S and G2 phases of the cell cycle, when CtIP is active. Indeed, a subfraction of DSBs in G2 requires BRCA2 for their repair, but knock-down of both CtIP and BRCA2 alleviates this repair defect [45], suggesting that avoiding resection prevents HR and allows repair of these DSBs by NHEJ. Replication associated breaks, on the other hand, should be channeled to HR, which is the only pathway that can restart a replication fork from a single broken DNA end.
The study of the balance between HR and NHEJ is important for the prediction of treatment responses upon inhibition of these pathways in various genetic backgrounds. Combined treatments might backfire when the balance is tipped the wrong way. For example, the treatment of BRCA1 and BRCA2 deficient cells is most effective when NHEJ is functional, whereas impaired NHEJ prevents lethal genomic instability and cytotoxicity, which counteracts the effect of PARP inhibitors in HR deficient cells [73,81].
The balance between HR and NHEJ is heavily regulated, but the wiring and hierarchy of this regulatory network is still incompletely understood. Development of targeted therapies using DNA damage response defects requires a much more detailed knowledge of the precise network of the cellular responses to DNA damaging treatments. It is to be expected that new assay systems will be developed and that a flurry of novel combinations of chemical inhibitors and genetic defects will increase our understanding of these processes in the near future. This knowledge will then be an invaluable source for developing new targeted therapies for tumors with DNA damage response defects, which should yield more specific and effective therapeutic approaches to combat cancer. Novel tools to characterize tumor-specific (DNA repair gene) mutations, such as whole genome sequencing approaches, should then bring truly personalized medicine for cancer treatment within reach.
Competing interests
The authors declare that hey have no competing interests.
Authors' contributions
IB wrote the first draft. IB and DvG wrote the final text. Both authors read and approved the final manuscript.
The research leading to these results has received funding from the European Community's Seventh Framework Programme (FP7/2007-2013) under grant agreement No. HEALTH-F2-2010-259893.
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text:
RemoteGroup Resource - MonitorOnly
I created a RemoteGroup resource last night, in VCS 5.1 SP1 RP1 on Solaris 10, to do some testing and validation. I needed a resource which monitors a remote service group (Oracle database service group). It doesn't need to cause the remote service group to go online or offline. It simply needs to show an online status when the remote service group is online, and show an offline (or faulted) status when the remote service group is not online.
I created the local RemoteGroup resource, and set the ControlMode attribute to "MonitorOnly". I thought that by setting up this resource in this way, it would behave much like a NIC resource. I was wrong. The NIC resource doesn't have to be brought online or offline. It just automatically is online when the NIC is working, and is offline (or faulted) when the NIC isn't working. Just what you'd expect.
When I first created the local RemoteGroup resource, it turned blue after probing the remote service group, and indicated an online status. This is what I expected. Then the remote service group was manually taken offline, and the local RemoteGroup resource became faulted. Again, just what I expected.
However, when the remote service group was manually brought online again, the local RemoteGroup resource did not automatically clear its fault, and become online again (like a NIC resource would). It just remained faulted. I don't think this is the expected behavior.
Another thing that was odd: When the local service group (with local RemoteGroup resource) was taken offline, the local RemoteGroup resource went offline. I would have expected the local RemoteGroup resource to remain online when the local group was offlined......like a NIC or NFS (Server) resource does.
My question is this: Is this the expected behavior of the MonitorOnly setting for the RemoteGroup resource? If so, is anyone aware of some settings that will allow this resource to behave much like a NIC resource, when the ControlMode attribute is set to MonitorOnly?
Thanks, -John
Comments 1 Comment • Jump to latest comment
The behavior you describe is as expected as it is an OnOff resource - its operations are not "None" like NIC. If it were in a failover service groups and it faulted and onlined on another service group (as target DB service group being monitored came back on line) then if resource cleared and onlined, like a NIC on faulted system, then you would have an OnOff resource online on 2 systems causing a concurrency violation. If the database was in the same cluster then if it faulted you would have to manually clear it and there is no way round this apart from scripted, so really it is the same using the remote group resource.
If you want it to act like a NIC then you would need to write your own agent with operations of None. You could actually clone the RemoteGroup agent and make it operations of OnOnly, but this only part solves your problem as it would mean the remote groiup resource would not offline if you offlined the service group it was contained in, but I am not sure what, if anything, you gain from this. I am pretty sure you can't clone and make opeations of None as I thing the online entry point creates a lock file that the monitor entry point monitors.
Another alternative would be to put the RemoteGroup agent in a separate parallel service group (this is only supported from 5.1 SP1) and use an Infinate (high number which is effectively infinate) for the OnlineRetryLimit on the servicegroup (NOT the resource) and use a PreOnline for the service group to clear the fault.
UK Symantec Consultant in VCS, GCO, SF, VVR, VxAT on Solaris, AIX, HP-ux, Linux & Windows
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text:
Ng, F.S.L. and Zuber, M.T. (2006) Patterning instability on the Mars polar ice caps. Journal of Geophysical Research - Plants, 111 (E2). Art no.E02005 . ISSN 0148-0227
Full text available as:
ng_zuber_2006[1].pdf Download (3227Kb) |
We present a mathematical theory to study the origin of large-scale spiral troughs on the Mars residual polar caps, starting with the hypothesis that atmospheric circulation governs the planform of the troughs via an instability that operates in the flow direction of surface winds. This concept can explain why the troughs spiral at each pole in an opposite sense to that expected for Coriolis-deflected winds. The instability arises from interactions on water ice, assumed to contain dust, and depends on how the exchange of atmospheric dust and moisture (H2O) with the polar cap surface controls its albedo and mass and energy balance. Our model predicts spatial patterns to form when moisture is carried by wind over the surface, owing to unstable coupling between the albedo and the H2O-vapor pressure. The resulting albedo pattern causes an alternating "accumulation-ablation'' mass balance, so that an undulating topography develops which resembles the (dark) troughs and their adjacent (bright) smooth terrains on the polar caps. Because the albedo patterning process is fast, whereas topographic evolution is slow, we suggest that an ancient imprint in the surface albedo preconditions today's trough morphology.
Item Type: | Article |
Copyright, Publisher and Additional Information: | © 2006 American Geophysical Union. Reproduced in accordance with the publisher's self-archiving policy. |
Keywords: | Orbiter Laser Altimeter; Layered Deposits; Water Ice; Flow; Topography; Sublimation; Variability; Features; History; Regions |
Institution: | The University of Sheffield |
Academic Units: | The University of Sheffield > Faculty of Social Sciences (Sheffield) > Department of Geography (Sheffield) |
Depositing User: | Miss Anthea Tucker |
Date Deposited: | 14 Dec 2010 09:19 |
Last Modified: | 04 Jun 2014 19:42 |
Published Version: | http://dx.doi.org/10.1029/2005JE002533 |
Status: | Published |
Publisher: | American Geophysical Union |
Refereed: | Yes |
Identification Number: | 10.1029/2005JE002533 |
URI: | http://eprints.whiterose.ac.uk/id/eprint/42721 |


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text:
Jack Klugman's son Adam has slammed the late Cory Monteith — because the Glee star is getting an extended Emmys tribute and his Odd Couple actor dad isn't.
Adam Klugman criticized Emmy organizers for extending a special honor to Monteith, who died in July of heroin and alcohol poisoning at age 31, while putting his father into the routine "In Memoriam" reel.
Referring to Monteith's overdose, Adam Klugman said in an Associated Press interview: "They're celebrating this self-inflicted tragedy instead of celebrating the life of my father, who won three Emmys…. Cory Monteith never won an Emmy."
PHOTOS: Celebs Who Have Battled Drinking Problems
Adam, whose Quincy M.E. star father died in December at age 90, said he has nothing against Monteith but "to compare the contribution he made to the contribution my father made — it doesn't compare.
"It's an insult and it really seems typical of this youth-centric culture that has an extremely short attention span and panders to only a very narrow demographic" of young adults, he said.
This year, for the first time, Emmy organizers have decided to beef up the "In Memoriam" section with special tributes to five notable TV figures who died this year. The other honorees are writer-producer Gary David Goldberg, comic legend Jonathan Winters, and actors James Gandolfini and Jean Stapleton.
PHOTOS: Heroin Addicts, Coke Fiends & Celebrity Stoners — Hollywood Drug Dealers Tell All!
In addition to leaving Klugman off of the extended tribute list, organizers also aren't doing anything special for Larry Hagman, who played iconic J.R. Ewing on the original "Dallas."
In a statement, Emmy executive producer Ken Ehrlich explained, "To a younger generation, Cory Monteith's portrayal of Finn Hudson [on 'Glee'] was highly admired, and the producers felt that he should be included along with the four other individuals we have singled out."


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Embracing the shift: Edupunk, Teaching Naked, Social Media and making it all work in the classroom[download PDF]Howard Cook
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Fall Out BoyFri, September 6, 2013 - 7:00 PMTsongas Center in Lowell, MA |
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Congratulations to the following people:
Winners were decided with a random number generator.
Thanks for entering, everyone! If you didn't win, better luck next time!
In addition, if you have any interest in journalism, writing, publishing or the video gaming industry and you have Facebook, I'd appreciate it if anyone could like my Facebook page: www.facebook.com/pages/Alex-Ph…
I currently write everything from articles for music magazines to columns about video games, to freelance work for publishers and news articles for universities. Recently, I'm branching out into interviewing developers for video games that many of you surely enjoy playing.
If you're also entering the premium giveaway, feel free to mention in your comment with your link to your journal that you also liked the Facebook page.
Have a happy time!
WOW, I'm SO surprised that the winners are Premium Members. Urgh.... Anyways, that's just my bad side. Congrats to the winners who won. (I never win anything)


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Book your room at Century Royal Hotel
Exclusive Double Room
Max Occupancy: 2 guestsEconomy Single Room
Max Occupancy: 1 guestEconomy Twin Room
Max Occupancy: 2 guestsDouble Room Non Smoking
Max Occupancy: 3 guests (up to 1 child)Twin Room Non Smoking
Max Occupancy: 2 guestsDeluxe Twin Room
Max Occupancy: 2 guestsCorner Twin Room with Extra bed (Non-Smoking)
Max Occupancy: 3 guestsExclusive Twin Room
Max Occupancy: 2 guestsSemidouble Room, Non-Smoking
Max Occupancy: 2 guests (up to 1 child)Large Twin Room
Max Occupancy: 2 guestsLuxury Twin Room
Max Occupancy: 4 guests (up to 2 children)4-star romantic hotel, walk to Sapporo JR Tower
Located in the heart of Sapporo, this romantic hotel is within a 15-minute walk of Sapporo JR Tower, Former Hokkaido Government Office Building, and Clock Tower. Odori Park and Sapporo TV Tower are also within 1 mi (2 km).
Hotel Features
This hotel features 3 restaurants, a coffee shop/café, and self parking. WiFi in public areas is free. Additionally, a business center, a meeting room, and dry cleaning are onsite.
Room Amenities
All 300 rooms boast private spa tubs and deep soaking tubs, while also providing free WiFi, free wired Internet, and flat-screen TVs with satellite channels. Guests will also find refrigerators, minibars, and room service.
Information missing or incorrect? Tell us!Hotel Amenities
Hotel Amenities
Dining is available at one of the hotel's 3 restaurants and guests can grab coffee at the coffee shop/café. The property also has a grocery/convenience store. A computer station is located on site and high-speed wired and wireless Internet access is complimentary. Business-related amenities at this 4-star property consist of a business center and a meeting room. This romantic hotel also offers a terrace, multilingual staff, and gift shops/newsstands. Onsite parking is available (surcharge). A total renovation of this property was completed in December 2013.
Century Royal Hotel has designated areas for smoking.
- 24-hour front desk
- Business center
- Elevator/lift
- Gift shops or newsstand
- Designated smoking areas
- Grocery/convenience store
- Free WiFi
- Free wired high-speed Internet
- Computer station
- Total number of rooms - 300
- Number of floors - 23
- One meeting room
- Terrace
- Porter/bellhop
- Number of restaurants - 3
- Long-term parking (surcharge)
- Offsite parking discounted rates available
- Self parking (surcharge)
- Breakfast available (surcharge)
- Coffee shop or café
- Free newspapers in lobby
- Dry cleaning/laundry service
- Multilingual staff
- Safe-deposit box at front desk
- Number of buildings/towers - 1
- Wedding services
Family Friendly Amenities
- Free Wi-Fi
- Grocery/convenience store
- Refrigerator
- Cribs/infant beds (complimentary)
Available in all rooms: Free WiFi , Free wired high-speed Internet
Available in some public areas: Free WiFi , Free wired high-speed Internet
Long-term parking (surcharge), Self parking (surcharge), Offsite parking discounted rates available
Room Amenities
- Pay movies
- Free newspaper
- Refrigerator
- Daily housekeeping
- Shower/tub combination
- Iron/ironing board (on request)
- Climate control
- Phone
- Minibar
- Satellite TV service
- Free WiFi
- Private bathroom
- Free toiletries
- Hair dryer
- In-room massage available
- Slippers
- Deep soaking bathtub
- Room service (limited hours)
- Rollaway/extra beds (surcharge)
- Free cribs/infant beds
- Desk
- Flat-panel TV
- Blackout drapes/curtains
- Private spa tub
- Free wired high-speed Internet
- Pillow menu
Where to Eat
Buffet breakfasts are available for a surcharge and are served each morning between 6:30 AM and 10 AM.
YUUYOO TERRACE - Overlooking the garden, this buffet restaurant specializes in fusion cuisine and serves breakfast and lunch.
Sky Restaurant RONDO - This restaurant specializes in French cuisine and serves lunch and dinner.
KITANOJI - This restaurant specializes in Japanese cuisine and serves breakfast, lunch, and dinner.
Room service (during limited hours) is available.
Spa Services
- Massages
Hotel Policies
Check-in time starts at 2 PM
Check-in time ends at 11 PM
Check-out time is 11 AM
Payment types
You need to know
Extra-person charges may apply and vary depending on hotel policy.
Government-issued photo identification and a credit card or cash deposit are required at check-in for incidental charges.
Special requests are subject to availability upon check-in and may incur additional charges. Special requests cannot be guaranteed.
Parking height restrictions apply
- Fee for buffet breakfast: JPY 2484 per person (approximately)
- Self parking fee: JPY 1300.00 per night
- Rollaway bed fee: JPY 5500 per day
We should mention
Reservations are required for massage services and spa treatments. Reservations can be made by contacting the hotel prior to arrival, using the contact information on the booking confirmation. The Japanese Ministry of Health, Labour, and Welfare requires all international visitors to submit their passport number and nationality when registering at any lodging facility (inns, hotels, motels, etc. ). Additionally, lodging proprietors are required to photocopy passports for all registering guests and keep the photocopy on file.


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Released: Feb 01st, 2002
Running Time: 125
Director: Shayla LaVeaux
Company: Shayla Productions
Distribution Companies: Jill Kelly
Shayla LaVeaux, Peter Colin, Anthony (I), Chloe (I), Samantha (I), Taiga, James (I), Jazz (I), Tony (I) (Hungarian)
Critical Rating: AAA 1/2
Genre: All-Sex
Shayla fans, naughty travelogue devotees.
Shayla LaVeaux goes for a romp in Australia and engages a series of locals in sundry sorts of sleazy fun. More a pro-am reel than anything else, this title is at its best when contrasting LaVeaux's consummate professionalism with the horny enthusiasm of local Aussie would-be porn stars. The two highlights here are LaVeaux's tussle with paid escort Samantha in a scene which cooks with lots of real-deal estrogen-driven g/g eroticism, and LaVeaux's athletic little triad with a pair of male strippers who may arguably be more into each other than they are her, but who are in any case buff, bronzed and quite pretty.


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MAYBE ONCE OR TWICE A YEAR, a game comes along that everyone finds time to play. Last year, it was Battlefield 1942, a game that even rival publishers and developers happily admitted an addiction to. This year? That game might not have arrived yet, but Call of Duty is making an inspired bid for the title -- it seems everyone we know has played the demo multiple times and is aching for more.
That's why we were more than happy to make the trip to Fort Mason in San Francisco this past Thursday, where Activision and Infinity Ward were showing off more of the game than ever before: seven single-player levels and the first-ever look at the game's multiplayer. We spent a few hours playing through both, and, well, let's just say we're ready. Is it out yet?
To start off the evening, Activision brought in retired Army Colonel Hank Keirsey, one of two military advisors consulting on the project. After a bit of background (among numerous accomplishments, Keirsey spearheaded one of the earlier missions during the Gulf War), as well as a bit of schtick mixed with briefings for a few single-player missions, Keirsey gave the crowd their marching orders, and off we went to fight it out.
The single-player missions on display were a mix of levels from the American, British and Russian campaigns, some of which we've covered in earlier previews: "Dawnville," "Brecourt Manor," "Pegasus Bridge" and "Pavlov's House." To start, we replayed through Dawnville (which is the level immediately following the demo in St. Mere Eglise), and immediately noticed one big change: there's now a large green health bar in the lower right. Apparently, a lot of people were confused by the original red health meter in the lower left, prompting Infinity Ward to create something they thought would be a bit more intuitive.
Of the remaining three levels, two were vehicle-based, extremely similar in concept to those found in recent Medal of Honor games. The first was a ridealong in the back of a truck, where your soldier is armed with a Panzerfaust and tasked with shooting down enemies chasing from behind. The second put you in the driver's seat of a tank, where we rolled along laying waste to everything in front of us.
The highlight of the evening for us, however, was the "Red Square" mission, one of the levels from the Russian campaign. Similar to the "Stalingrad" mission that kicks off the campaign, the mission starts with a stirring speech from your commanding officer, who reminds the troops that they're never to retreat -- "never one step back." After a few moments (with some dramatic music added for full effect), the army rushes the yard in one of the most epic videogame scenes in recent memory ... and Call of Duty already has a few. Soldiers scramble forward and scatter in all directions, and after your character finally finds a weapon, he finds a good flanking position and starts taking out mounted guns across the yard.
(Note: I was so impressed with this level that we took footage of the entire scene and made the FilePlanet guys promise to get it out to the public ASAP, so go grab it if you want to see it in action. If you can watch this movie and you're still not interested in Call of Duty ... well, PC action games may not be for you. :)


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[ Bars & Clubs Index | Bars & Clubs | Metro | Metroactive Central | Archives ]
Bars & Clubs
Live Salsa & Latin
Club Caribe
Club Miami
Copa Cabana
Jose's Caribbean Nightclub
The Latin Village
Towne Oak Supper Club
[ Metro | Metroactive Central | Archives ]
Copyright © Metro Publishing Inc. Maintained by Boulevards New Media.
Live Rock & Pop | Live Jazz & Blues | Cocktail Lounges
Live Salsa & Latin | Country & Western | Coffeehouses
Comedy & Vaudeville | Dance Clubs | Hangouts | Pubs
Sports Bars | Microbreweries | Billiard Parlors | Juice Bars
Go-Go & Strip Clubs | Wineries
736 W. Dana St., Mountain View (415/968-3007)
"Discotheque" is a French word, but at Alberto's, it is pronounced with a Spanish accent, especially on Sunday, which is officially Disco Night at this downtown Mountain View institution. The decor recalls a low-budget movie version of Havana, Miami or East L.A. The ambiance varies night to night: Tuesday is salsa night (dance lessons begin at 7pm); Wednesday is Caribbean night (look for steel-drum bands or reggae). Cover ranges from $10 to $15, with a discount for those holding a monthly pass, available for $25.
1001 S. First St., San Jose (408/297-7272)
The people are friendly, the dance floor huge and the music loud (but it's a good kind of loud). Club Caribe features both live DJs and live bands, with tunes ranging from ranchera to banda to tejano. This is a clean, well-designed club, with seating around the dance floor from which patrons can watch the dancers twirl and still see the band. High ceilings and good ventilation keep the smoke at bay and the climate cool. Open 8pm2am, except Sunday.
177 W. Santa Clara St., San Jose (408/279-3670)
Hot, hot, hot sums up Club Miami. Steamy salsa music blasts through the strobe- and neon-lighted club. It is both upstairs and upscale. Sharply dressed dancers move, spin and sway to the unending thump, thump of the merengue music. Thirsts are quenched with beer, wine and the occasional shot of tequila. Tight clothes are required for both sexes to advertise the goods. The patio provides a place to talk, scam and watch the hip Latinas smoke cigars. Cover runs $5$10, ThursdaySaturday.
2618 Alum Rock Ave., San Jose (408/272-3481)
For 16 years, the Copa Cabana has been bringing live music and entertainment to south San Jose. Located in the Alum Rock Plaza, the club features both live DJs and live bands playing everything from salsa and cumbia to ranchera and tejano. The flashing lights and glitter ball radiate '70s, but turn away from the crowded dance floor, and it seems more like an earthy local drinking spot. The manager describes the dress code as "formal," and patrons do seem to dress up for the evening. Open 2pm2am, seven days a week.
2275 El Camino Real, Palo Alto (415/326-6522)
There are very few places in the valley where you can eat Cuban food and dance to live reggae till 2am. Jose's Caribbean Nightclub and Restaurant is one such place. With bright pink and blue place mats on the tables and a well-stocked indoor tiki bar, Jose's attracts an eclectic clientele--mostly dancers--from Latinos to yuppies to youths. At Jose's, you can munch on fried plantains and Creole alligator, wash that down with a pink Hawaiian Hurricane, polish your meal off with a smooth Cuban coffee and then hit the floor for salsa classes. (A little Dramamine might be in order, first.) Call for the somewhat erratic schedule at Jose's ahead of time, and then be prepared for something entirely new. Open till 2am weekends.
1899 Alum Rock Ave., San Jose (408/259-7712)
The Latin Village is the place to find the traditional Mexican charro (cowboy) in his full regalia: jeans, boots, a wide brim Stetson and, of course, holding it all together, buckles the size of the East Side. The large band of caballeros playing cumbias and rancheras pack the dance floor while lonely Latinas wait to be asked to join in the fun. The room is smoky, the beer is cold and there are plenty of waitresses to serve up brews. Parking is free, although the cover charge comes and goes depending on the mood of the doorman.
407 Town and Country Village, Sunnyvale (408/732-5500)
By day, a quiet ivy and brick lunch restaurant with a glassed-in atrium, and a quiet clientele on the other side of 40 munching on the Towne Oak Steak Bomb and the Towne Sizzler. There's a quiet, upscale bar, flanked by portraits of Marilyn Monroe and Mae West, serving Gordon Biersch Märzen, Talisker single-malt and cocktails. On Saturday nights, the Towne Oak comes to life with live salsa, cumbia and merengue with such bands as Orquestra Borinquen and Orquestra Cadencia; free appetizers and CD giveaways are offered to lure in dance fans. Open daily, 11:30am2am.
From the June 12-18, 1997 issue of Metro.
Foreclosures - Real Estate Investing |
San Jose.com Real Estate |


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You need the latest Flash player to see this video.Click here to get Adobe Flash player
wow i want a nosh off her fit
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she looks like my new cock i wanna fuck
hot scene!
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who is she???? i want to get more videos!!! someoen tell me pls


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Portsmouth County, Va., funds by Energy, Department of
Listing $1,002,000.00 in stimulus funds from Energy, Department of for Portsmouth
Note: For some programs where states do not report where money will be distributed across the state, we do not have the allocation for individual counties. Those programs include: Medicaid, unemployment benefits and food stamps. Those amounts are included in the totals for where the state agency receiving that money is located.
Amount refers to both the amount of stimulus funding going toward the project and the face value of the loan.
Recipient | Amount | Description | Federal Dept./Agency | Date |
PORTSMOUTH, CITY OF | $1,002,000 | Energy Efficiency and Conservation Block Grant Program (EECBG) The City Portsmouth plans to: 1) retrofit four city owned parking garages with energy efficient lighting and lighting control systems to reduce energy usage and associated carbon emissions by 35%. 2) synchronize the operation of the city'y's 94 signalized i... Show more | Assistant Secretary for Energy Efficiency and Renewable Energy | 9/30/2009 |


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Telos GSA Schedule GS-35F-4315D
Blanket Purchase Agreement FA8771-09-A-0301
The DoD Enterprise Software Initiative (ESI) is a joint effort in coordination with the federal-wide SmartBUY program to maximize DoD buying power for commercially available information technology and selected services on a DoD-wide basis. Telos offerings through the DoD ESI include:
Xacta® IA Manager
Xacta IA Manager establishes a centralized security management platform that facilitates compliance assessment, continuous risk and compliance management, and security process enforcement. In addition to identifying risks, Xacta IA Manager automatically initiates corrective actions to protect systems. It automates the C&A process, reduces time to produce an SSAA, and lowers cost to conduct a C&A. Quick procedures and full compliance with DFARS reduces acquisition costs.
Telos AMHS Telos Automated Message Handling System (AMHS) allows users to manage their respective classified and unclassified defense messages in a more efficient and cost-effective manner and provides agency users with precise control over the large volume of messaging information they already handle every day. Further, AMHS enables agencies to easily create, coordinate, release, archive, and search massive volumes of messages. Armed with AMHS, users can route mission-critical messaging information in real-time to appropriate personnel regardless of their location whether they are in the office or on the battlefield.
Support Services and Training Support services and training are also available at discounted rates.
Defense Federal Acquisition Regulation Supplement (DFARS) Section 208.74 requires use of an ESA before using another method of acquisition. Telos DoD ESI Portal: https://esi.telos.com/ Contact Information
General Inquiries
For general information on Telos' DoD ESI contract, please contact us at 1-800-70TELOS (1-800-708-3567)
Contact our sales staff for assistance with your requirements by calling toll free 1-800-70TELOS (1-800-708-3567) or via their contact information below.
Sales Angela Robinson 703-726-2267 703-724-3862 (FAX) E-mail: firstname.lastname@example.com
VP, Business Development/Sales Rinaldi Pisani 703-724-4681 703-724-3862 (FAX) E-mail: email@example.com
Program Manager Doug Luckett 703-858-4833 703-724-3862 (FAX) E-mail: firstname.lastname@example.com
DoD ESI


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Franklin D. Roosevelt (FDR)
Franklin D. Roosevelt (FDR): Death
Although Franklin Delano Roosevelt led the country for twelve years, he never saw the peaceful prosperity he worked to create. Years of disease, stress and constant cigarette-smoking took their toll. In March of 1945, Roosevelt traveled to his resort in Warm Springs, Georgia, to rest. On 12 April, he suffered a massive brain hemorrhage and died at the age of 63.
FDR's legacy is immense. He led the nation through two of the greatest crises in its history, he expanded the power of the presidency, and he created new agencies that transformed the federal government and, in the process, its relationship with its citizens. Alongside Abraham Lincoln and George Washington, many consider him to be one of the influential presidents in American history.
Ultimately, FDR was not trapped by the privileges and opportunities of his patrician upbringing. Instead, he found the confidence and optimism with which to lead the nation through two great crises. A consummate politician, he pulled the strings of government by giving everyone what they needed—the press, the people, and the Congress. He was not without his shortcomings—the internment of Japanese-Americans, and the failure to prevent the Holocaust chief among them—but he is remembered as a great president because he became what Americans needed most: an optimistic, decisive, and able leader, who could both inspire and get things done.
In the end, much of his success seems to have stemmed from his upbeat personality and from his ability to play the political game. As a friend observed, Roosevelt picked politics for his career, and enjoyed it, "just as one enjoys a game that one has always liked and learned to play well."45 The fruits of this game were beyond measure. Shortly after his death, the New York Times said, "Men will thank God on their knees a hundred years from now that Franklin D. Roosevelt was in the White House."46


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USA v. Talgar General Partnership, et al
Defendant: | Talgar General Partnership, H. William Gardner and Bruce Talbot |
Plaintiff: | United States of America |
Case Number: | 1:2012cv00219 |
Filed: | June 11, 2012 |
Court: | New Hampshire District Court |
Office: | Concord Office |
Presiding Judge: | Landya B. McCafferty |
Nature of Suit: | Civil Rights: Accommodations |
Cause of Action: | 28:1331 Federal Question: Other Civil Rights |
Jury Demanded By: | Plaintiff |
Available Case Documents
The following documents for this case are available for you to view or download:
Date Filed | # | Document Text |
August 15, 2012 | 12 | ORDER. Attorney Rosenbaum shall register for Electronic Case Filing (ECF) on or before August 29, 2012. Failure to register by that date shall result in attorney Rosenbaum being removed as counsel of record. So Ordered by Judge Paul J. Barbadoro.(vln) |
Access additional case information on PACER
Use the links below to access additional information about this case on the US Court's PACER system. A subscription to PACER is required.
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St Louis County Library
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Library Foundation Hosts Holiday Bazaar & Open House
Thu, 11/07/2013 (All day)
The St. Louis County Library Foundation will host a holiday bazaar and open house at Library Headquarters, 1640 South Lindbergh Blvd. in Ladue on Saturday, November 23 from 12:00 to 2:00 p.m. Festivities include special sales items, author signings, children's activities and more.
Artist Mary Engelbreit will attend and sign a variety of her holiday merchandise. Items will be available for purchase at the event.
Additional activities include a book signing with John Oldani, author of "Christmas in St. Louis: Traditions, Displays, and Celebrations." Baked goods from Companion will also be available. Bebe the Clown and face painting will keep the kids entertained.
All sales from the Open House will benefit the St. Louis County Library Foundation, which is a 501 c 3 tax-exempt organization that raises money for and awareness of the St. Louis County Library.
For more information please call 314 994-3300.
Program sites are accessible. Upon two weeks' notice, accommodations will be made for persons with disabilities. Contact St. Louis County Library by phone 314-994-3300, or visit www.slcl.org.
© Copyright 2015. St. Louis County Library. All rights reserved. 1640 S. Lindbergh Blvd., St. Louis MO. 314-994-3300
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Gummy toolbox is giving me an error when I try and access it from terminal. Its saying that busybox isn't installed. I've verified that it is, however, and still get the error. Screenshot coming. Sent from my SCH-I510 using DroidForums
Thank you! Also, it appeared that Busybox needed to be updated. The installer updated three or four times, then it worked.


text:
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Slam the Stands event at White River High Jan. 30
Slam the Stands will take place at 7 p.m. Jan. 30th at White River High School.
This will be the second Slam the Stand event.
This is a game wtih Enumclaw.
This is an opportunity to come out and support all of these athletes. There will also be T-shirts available for $7 at the event.
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Register for an Edmodo Webinar
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As we gear up for another great month on our Webinar calendar, we wanted to call attention to a couple of things that you might not be aware of with getting just-in-time professional development on Edmodo.
First, you can sign up for all of your favorite Edmodo webinars from a single page.
Second, you can tweet, like on Facebook, or post to LinkedIn that you are coming to a webinar with a single click from each one of our webinar registration pages:
Third, if you cannot attend a Webinar at the times they are offered, you can watch one in the Webinar Archives.
To further add to the options, we can also set up a personalized webinar for your district. If you have an Edmodo district subdomain (or would like one) and you can get at least 10 teachers and administrators together at a single time (doesn't have to be in the same space because they can connect to the webinar from anywhere), we would love to conduct a webinar just for you.
If you are interested in this option, please send an email to our Online Community Manager, Ben Wilkoff: ben(at)edmodo.com
One Response to "Register for an Edmodo Webinar"
l like this web site……
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African New Testament Geloof Kom Deur Gehoor Die Bybel NuweTestament
Original, contemporary, gospel, Christian, Blues and Country combine for an easy listening experience that is a powerful and safe musical excursion while you are working in on your computer in your office or homw.
inspirational music source for contemporary smooth jazz/christian
Kids New Testament read by Jack the Wonder Dog funded this year in honor of Dr. John Ohara
The Social Radio Network
Showcasing the Pioneers of Contemporary Gospel Music
#1 For Christian Hip Hop and R&P
Offering the best in Contemporary Jazz, Inspirational, and Instrumental- Not your everyday jazz station! Playing artist like Kirk Whalum, Gerald Albright, Kim Burrell, 3Central, Ben Tankard, Partners in Christ, Eric Copeland
The broadcast ministry of David Michael Carrillo & Friends
La Biblia Nuevo Testamento- Audio Drama con Musica y Efectos de Sonido
Jewish Rock Radio
Uplifting Music, (R&P) Rhythm & Praise,Gospel House and More
Playing the best in Gospel/Christen Music tune in daily to Spiritual Soul Radio.
A Piercing Mix of Reggae, Blues, Soul, Groove, Acid Jazz, Jazz, Latin plus a coupla weird ones


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Summary: 2040 Reports Ecology, 86(8), 2005, pp. 2040Ł2046 2005 by the Ecological Society of America TREE SWALLOWS TRADE OFF IMMUNE FUNCTION AND REPRODUCTIVE EFFORT DIFFERENTLY ACROSS THEIR RANGE DANIEL R. ARDIA1 Department of Ecology and Evolutionary Biology, Cornell University, Ithaca, New York 14853 USA Abstract. Latitudinal differences in life histories are believed to be underlain by dif- ferences in trade-offs between current and future reproduction. I report differences in trade- offs between parent and offspring across the range of a widespread avian species, the Tree Swallow (Tachycineta bicolor). I manipulated parental effort and found that in Alaska, where yearly adult return rates are low, breeding females increase their reproductive in- vestment to maintain offspring quality, whereas in Tennessee, where yearly adult return rates are high, breeding females are either unable or unwilling to increase reproductive investment and, consequently, raise offspring of lower quality. I further investigated a critical mechanism of self-maintenance that may underlie differences in survival among sites: immunocompetence. Females breeding in Alaska mounted weaker immune responses when raising enlarged broods, whereas females in Tennessee did not, further suggesting a strategy in Alaskan females to incur costs (i.e., reduce future reproduction) in order to
Source: Ardia, Dan - Department of Biology, Franklin and Marshall College
Collections: Biology and Medicine