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A monochromatical gray color scheme doesn't have to be boring as long as you layer, add textures, and bring in bold furniture and decor pieces. The seating above is truly a work of art. A modern chair combined with a rustic tree stump side table is an unexpected pairing but it all works in the space. That is another way you can keep things interesting in a muted palette.
Tag Archives: furniture
Did ya know TJMaxx now offers online shopping?
This isn't a sponsored post it's just TJ Maxx is a store I've loved since working there as a teen. I worked there a year in the service desk area doing returns my freshmen year of college. Once I started having clients, it has been one of my go-to stores for affordable, pretty, and unique decor items. You can buy a whole room filled with stuff from TJ Maxx and it won't feel as if you shopped at one place and it won't break your budget. What I love most is the fact that you can get sizeable mirrors for under $40. I'm talking mirrors big enough to swap out and replace over bathroom vanities like you see in this post. They have nice rugs, chic pet bowls & stuff, drapery, a lot of cool end tables and chairs. Another thing I love is the brand named faux fur throws and pillows and often times they are under $20 like the ones in my collage. I don't know about you but my clients are not trying to spend $400 on two pillows. So if you're in a budget, or simply want more bang for you buck without compromising style, head over to your nearest TJMaxx or online.
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Chairish: a vintage furniture and accessories store
I have been challenged to pick some vintage pieces from Chairish to create a styleboard featuring one of my favorite picks. I loved the vintage Persian Bidjar rug and decided to make a few styleboards around that rug. Then I ended up spending a bunch of hours lost on their website in a black hole of beautiful goodies that made me wish I were rich. I swear there is so much good stuff being sold on that site. Basically Chairish is a site where people can buy and sell vintage decor, furniture and art. The items are carefully curated and it has an etsy/ebay feel where you're buying from trustworthy sources and owners.
So I started compiling pieces and end up creating four different styleboards. The first I envisioned in a corner of a bedroom. You have the armoire and a chair with a lamp to create a nice reading nook. Next I created a sideboard styled area of a living room. I love how the styling is asymmetrical but still feels balanced by the wall sconces. You could even sit a flat screen tv there for an updated more stylish version of tv stands. See more of that here. This board has an eclectic feel…
Then I had to create something girly and glam with a blush pink palette, mirrored and lucite finishes & a pop of gold. This would be a pretty entryway area that is sure to set the tone of anyone who comes in for the first time. You have a pouf to sit on and take shoes off after a long day of work, a mirrored screen to check your hair one last time before stepping out and a place for keys. I added the bust and vintage sign for a little whimsy.
Lastly using the same vintage Persian rug, I styled a sitting area. I forgot to include a coffee table but if I did, I would pick something with glass so that you can see and appreciate the beauty of the rug. There are some Asian influences in this space with the color scheme, the brass fretwork wall sconces and the contemporary Asian latticework buffet. So which styleboard do you like the most? Every single item with the exception of plants were found on Chairish.
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Need some furniture or decor? Live in metro Milwaukee?
These are some of the items being sold this weekend. My uncle let me decorate his entire place 5 years ago so majority of it are things I picked out for him. Everything must go and there are some really amazing deals on quality furniture. If you live in the Milwaukee area and want to inquire about something, email me and I will give you details and send you the flyer.
Just a few colorful spaces…
{Images via Society Social & One Kings Lane}
I just love Zgallerie…
Tiny Tot Thursday: restoration hardware baby and child
The crown canopies make the rooms of Restoration Hardware's baby & child. Just the right amount of charm for baby royalties. The collection uses classic elements found in adult spaces like chandeliers and tufted upholstery and some industrial styled decor.
Not only are tufted cribs visually beautiful but the extra upholstery is sort of like baby proofing as it would soften the blow if a child falls against it.
I like the army styled teepee. Teepees are just so much fun. They literally bring out the kid in you.
Some items are quite pricey because it's a luxury indulgence but the pieces of furniture are beautiful enough to pass down generations or continue to grow with the child. Some items have a historical aged appearance that are timeless and truly a refreshing classic take on children room designs.
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We all sat together as one…
Remember When….I created that last collage…
You can see that collage here. The next day I stumbled upon Bliss At Home which had a knock off DIY tutorial of the Worlds Away Anna bar cabinet and boy did she knock it out. 👊
I don't know what I like best: the leopard lining or the teal hue. I know one thing, it inspires me to want to change my hardware on everything.
The rest of her home and the DIY projects is equally gorgeous.
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Designs and dimensions in tile, doors and furniture
I saw AKDO's tile on pinterest and loved the design. I can see this as backsplash for a stove with gold fixtures. So pretty…
Then I saw this which is making some patio awfully fabulous. See more here…
This door is amazing. Talk about an entrance…
The lines of this desk reminds me of the one Kelly Wearstler used here…
This room has a lot of dimension too in the form of the many textures through out: wood paneled walls, woven headboard, quilted blanket, leather tufted bench. Textures gives interest in an earthy color palette or monochromatic design. Do you like dimension or prefer flush smooth surfaces?
{Other images via Arkpad}
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Two proteins implicated in inherited deafness, myosin IIIa1, a plus end directed motor2, and espin3–5, an actin bundling protein containing the actin-monomer-binding motif WH26, have been shown to influence the length of mechanosensory stereocilia7, 8. Here we report that espin 1, an ankyrin repeat-containing isoform of espin6, colocalizes with myosin IIIa at stereocilia tips and interacts with a unique conserved domain of myosin IIIa. We show that overexpression of these proteins causes elongation of stereocilia greater that when myosin IIIa alone or espin 1 alone are overexpressed. When these two proteins are co-expressed in the fibroblast-like COS-7 cell line they induce a ten-fold elongation of filopodia. This extraordinary filopodia elongation results from the transport of espin 1 to the plus ends of F-actin by myosin IIIa and depends on espin 1 WH2 activity. This study provides the basis for understanding the role myosin IIIa and espin 1 play in regulating stereocilia length, and presents a physiological example where myosins can boost elongation of actin protrusions by transporting actin regulatory factors to the plus ends of actin filaments.
Stereocilia, the prominent actin protrusions on the apical surfaces sensory hair cells, emerge early during development and their lengths are maintained at fixed heights for the lifetime of the organism. The bundle of parallel actin filaments that make up the core of each stereocilium is continuously renewed, with the entire actin bundle constantly assembled at the tip, treadmilling downward, and disassembling at the base9–11. Given that stereocilia can be up to 100 µm in length, it is likely that some form of regulated transport is necessary to localize components of the actin polymerization machinery to the plus end of the actin filaments. Although several myosins have been shown to alter stereocilia lengths and shapes depending on their expression levels9, 12–16, the mechanisms by which these motors or their binding partners regulate actin dynamics and stereocilia lengths remain unclear.
Using antibodies specific to the ankyrin repeat domain (ARD) of espin 1 (Supplementary Information, Fig. S1), we show localization at stereocilia tips with a tip-to-base gradient distribution (Fig. 1) similar to what was previously described for myosin IIIa. The immunofluorescence of espin 1 is more intense in the longer stereocilia and the characteristic tip-to-base fluorescence intensity gradient has a longer decay length (Fig. 1g,h). In contrast to other espin isoforms, which are present inside the actin core and along the entire stereocilia length8, espin 1 is excluded from the actin cores and forms a thimble-like distribution at the tips of stereocilia (Fig. 1). Immunofluorescence in developing hair cells of the rat organ of Corti shows that espin 1 can be detected at the tips of stereocilia during their elongation and maturation phases (Fig. 1k–m). To confirm the localization of espin 1 at the tips of stereocilia, we overexpressed GFP-espin 1 in organotypic cultures of hair cells. Transfected hair cells show that GFP-espin 1 localizes at the tips of stereocilia, exhibiting a tip-to-base gradient of intensity (Fig. 2) comparable to the immunolocalization (Fig. 1). These localization patterns – tip-to-base gradients and thimble-like distributions7, as well as the temporal expression pattern17 – closely match those of myosin IIIa. We hypothesized that targeting espin 1 to stereocilia tips, which is the site of actin polymerization9, 11, influences actin polymerization and stereocilia elongation. An analysis of the heights of stereocilia of cochlear and vestibular hair cells transfected with GFP-espin 1 shows elongation of stereocilia upon overexpression of espin 1 (Fig. 2), consistent with our hypothesis.
The striking similarity of the tip-to-base gradient localization of both espin 1 and myosin IIIa prompted us to investigate whether myosin IIIa helps localize espin 1 to the tips of stereocilia and whether they have a combined role in the regulation of stereocilia length. We compared stereocilia length in hair cells transfected with espin alone, myosin IIIa alone and with a combination of both plasmids (Fig. 2). Hair cells transfected with myosin IIIa and espin 1 show an increase in stereocilia length, higher than the combined increase observed for myosin IIIa alone and espin 1 alone (Fig. 2). It is important to note that any analysis of lengthening due to overexpression of myosin IIIa and espin 1 in hair cells must take into account intrinsic limitations due to natural stereocilia length variations and, importantly, the fact that stereocilia are already quite elongated and express robust amounts of endogenous espin 1 and myosin IIIa.
We tested whether myosin IIIa can effectively interact with and transport espin 1 in COS-7 cells, using filopodia as a model to study actin protrusions. Myosin IIIa and espin 1 are not naturally expressed at detectable levels in COS-7 cells 7. Myosin IIIa has been shown to induce filopodial actin protrusions and localize to their tips in cultured cells particularly well when its kinase domain has been removed (myosin IIIa ΔK)7,7, 18 suggesting that the kinase could serve to down-regulate the functional activity of myosin IIIa. We examined the distribution of co-expressed mCherry-ARD of espin 1 with GFP-tagged myosin IIIa ΔK (Fig. 3). We also co-expressed mCherry-ARD with GFP-tagged myosin X and GFP-tagged myosin XVa. Since all of these myosins accumulate at the tips of filopodia13, 16, 19, they provide a well-defined spatial compartment where any potential interaction can be clearly visualized. Co-transfections showed that mCherry-ARD is efficiently targeted to the tips of filopodia initiated by myosin IIIa ΔK, but not by myosins X or XVa (Fig. 3a), demonstrating a specific colocalization of ARD with myosin IIIa. Live imaging of COS-7 cells transfected with GFP-myosin IIIa ΔK and mCherry-ARD showed dynamic colocalization at the filopodia tips from the early steps of their initiation and elongation (Fig. 3b; Supplementary Information, Video S1). Live imaging also showed matching forward and rearward intra-filopodial movements of the GFP-myosin IIIa ΔK and mCherry-ARD fluorescence puncta while maintaining steady-state tip-to-base distributions (Supplementary Information, Video S2 and Video S3, and Fig. 3c), similar to the distributions of myosin IIIa and espin 1 observed in stereocilia (Fig. 1 and Fig 2). The intensity profiles for mCherry-ARD and GFP-myosin IIIa ΔK within each frame of the video (Fig. 3c) are highly correlated (cross-correlation = ~0.990), supporting the model that these two proteins are trafficking together in the filopodia. The interaction between espin 1 ARD and myosin IIIa ΔK was confirmed with a GST pull-down assay (Fig. 3e). The dynamic localization of espin 1 ARD at filopodia tips when co-transfected with myosin IIIa, but not with myosin X or with myosin XVa, along with our GST pull-down assay results, led us to hypothesize that myosin IIIa transports espin 1 to the tips of stereocilia.
We next asked which specific region of myosin IIIa is involved in the interaction with espin 1. Myosin IIIa has two conserved tail homology domains, designated as 3THDI and 3THDII20. We first co-transfected COS-7 cells with espin 1 and GFP-myosin IIIa and showed that the two proteins colocalize at actin bundles as well as at filopodia tips (Fig. 4a,b). This pattern of colocalization is abolished when we use a GFP-myosin IIIa construct that lacks the portion of the tail containing both 3THDI and 3THDII (GFP-myoIIIa Δ32; Supplementary Information, Table S1, Fig. S2; Fig. 4a,b). We next co-transfected COS-7 cells with espin 1 and with a GFP tagged tail portion lacking 3THDII (GFP-tailΔ3THDII; Fig. 4a,b) and narrowed down the region of interaction to the 3THDI and its immediate flanking regions. We observed colocalization of espin 1 with GFP-myosin IIIa tail that contained only the 3THDI domain (GFP-3THDI; Fig. 4a,b), but not with regions of only the myosin IIIa tail immediately amino-terminal (pre3THDI) or carboxyl-terminal (post3THDI) to the 3THDI domain (Fig. 4a,b). This suggests that the 3THDI domain is necessary for the myosin IIIa:espin 1 interaction. Together these data suggest that espin 1 and myosin IIIa specifically interact via their ARD and 3THDI domains, respectively. We verified this interaction in vitro using a GST pull-down assay and demonstrate that GST-ARD binds to GFP-3THDI, but not to the pre3THDI or post3THDI regions (Fig. 4c).
The fact that stereocilia length can be influenced by either espin 13, 8 or myosin IIIa7, along with the observation that they both localize to the same compartment at stereocilia tips and interact biochemically, suggests a combined functional role for the myosin IIIa:espin 1 complex in the elongation of stereocilia F-actin. We discovered that COS-7 cells co-transfected with myosin IIIa ΔK and espin 1 (Fig. 5a–c) display filopodial actin protrusions that can be up to ten times longer (mean length = 14.3 ± 9.1 µm; number of cells, nc =18; number of filopodia, nf=56) than those transfected with myosin IIIa ΔK alone (1.7 ± 0.83 µm, nc=12, nf=49), or with espin 1 alone (1.3 ± 0.28 µm, nc=13, nf=104). Mean lengths of filopodia of COS-7 cells transfected with empty GFP vector was 1.26 ± 0.7 (nc = 10, nf = 59). The synergistic effect between myosin IIIa and espin 1 is specific for myosin IIIa, since we found no enhanced elongation when espin 1 was co-expressed with either myosin X (2.40 ± 1.50 µm, nc=16, nf=165) or myosin XVa (2.08 ± 1.63 µm, nc=15, nf=134).
We used myosin IIIa without the kinase domain to observe the behavior of the dephosphorylated and more functionally active myosin. To exclude the possibility that the deletion of the kinase domain produces aberrant behavior, we developed a kinase-dead construct, myosin IIIa K50R (Supplementary Information, Table. S1). This construct allowed us to examine the role of autophosphorylation in the regulation of motor function, which in turn enabled us to investigate the role of myosin IIIa motor function in espin 1 tip-localization activity. We have determined that inactivation of the myosin IIIa kinase in a myosin IIIa 2IQ construct reduces the KATPase yet it does not affect maximal ATPase activity (Supplementary Information, Table S2 and Figure S3). We next evaluated the role of the kinase activity in myosin IIIa tip-localization in COS-7 cells using GFP tagged constructs. Full-length myosin IIIa K50R localizes more efficiently to the tips of filopodia in COS-7 cells (39% at tips nc=137) than wild-type (5% at tips nc=200), although not as strikingly as myosin IIIa ΔK (93% at tips n=105). Furthermore, co-expression of myosin IIIa K50R and espin 1 (Fig. 5e) yielded longer filopodia (mean length = 5.93 ± 3.10 µm, nc=15, nf=89) than co-expression of wild-type myosin IIIa and espin 1 (3.7 ± 3.2 µm, nc=15, nf=63; Fig. 5d), although not as long as the myosin IIIa ΔK:espin 1 co-expression. This data shows that myosin IIIa motor ATPase activity parallels the ability of myosin IIIa to localize to filopodia tips and to elongate filopodia when co-expressed with espin 1.
Interestingly, espin 1 co-expressed with a myosin IIIa ΔK lacking the tail domain downstream of exon 32 (myosin IIIa ΔK,33,34; Supplementary Information, Table S1, Fig. S2) resulted in slightly shorter filopodia (10.0 ± 4.74 µm, n=64; Fig. 5f) than co-expression with myosin IIIa ΔK. Using COS-7 cell co-expression and GST pull-down assays, we confirmed that the upstream portion of 3THDI (3THDI Δ33, Supplementary Information, Fig. S4) binds to espin 1. The 3THDII of myosin IIIa has previously been shown to be an actin-binding site18. Previous studies reported that myosin IIIa lacking the 3THDII actin-binding domain does not localize to filopodia tips 7, 18, but here we show that when co-expressed with espin 1 myosin IIIa goes to the tip and promotes filopodia elongation (Fig. 5f). It appears that the association with espin 1, which does have actin-binding sites, compensates for the missing actin-binding site in the myosin IIIa without the 3THDII domain.
Co-expression of espin 1 and myosin IIIa results in enhanced localization of espin 1 at filopodia tips (Supplementary Information, Fig. S5). When myosin IIIa ΔK is co-expressed with espin 1 lacking the ARD domain, we observed that both espin tip localization and filopodia elongation are abolished (Fig. 5g). These results demonstrate that the actin cross-linking activity of espin 1 is not solely responsible for the enhanced filopodia or stereocilia elongation observed in our experiments. We conclude that espin 1 promotes enhanced elongation of filopodia only when transported to the polymerization end of actin filaments by myosin IIIa. The fact that espin 1 elongates filopodia only when localized to the F-actin plus ends by myosin IIIa suggests that WH2-dependent polymerization activity is involved in elongation. We tested this hypothesis by substituting the first two of three highly conserved leucine residues of the espin 1 WH2 motif (L655A, L656A), which have been shown to be essential for its actin-monomer-binding activity21, 22. In COS-7 cells co-transfected with the WH2-mutated espin 1 construct (espin 1 mWH2) and myosin IIIa ΔK (Fig. 5h), the average length of filopodia (2.65 ±1.50 µm, nc=10, nf=75) remains comparable to the protrusions induced by myosin IIIa ΔK alone. The lack of enhanced elongation despite the colocalization of espin 1 mWH2 and myosin IIIa ΔK at the tips of filopodia (Fig. 5h) demonstrates that the WH2 motif is critical for mediating the role of espin 1 in elongation.
The steady-state distribution of myosin IIIa in a tip-to-base gradient is likely dynamically maintained. The length of the myosin IIIa distribution should be inversely proportional to the net velocity of the myosin towards the tip23, which will be slower for faster treadmilling actin cores (i.e. in longer stereocilia9 and filopodia24). This prediction is also consistent with our observation that wild-type myosin IIIa, which has relatively low activity, has decreased tip localization in the filopodia compared to the more active kinase mutant forms of myosin IIIa used in our experiments (Fig. 5). However, in stereocilia where the actin treadmilling is much slower, the wild-type myosin IIIa self-localizes effectively to the tip7 (Fig. 2). Similarly, the observed steady-state tip-to-base gradient distribution of espin 1 is not compatible with a model where espin 1 passively diffuses and binds to myosin IIIa resident at the tip, since this scenario would result in a homogenous distribution along the entire length of the stereocilia with no detectable concentration gradient at steady-state. The gradient distribution of espin 1 at steady-state is reminiscent of a myosin VI-driven gradient for the stereocilia membrane protein PTPRQ, and is best explained by a model that includes binding, directed transport, and diffusion of myosins and their cargo25. A more detailed consideration of this dynamic process that also accounts for actin treadmilling and plus-end directed motors predicts a similar distribution, which can be several microns long for longer stereocilia23. Thus, we favor a model where myosin IIIa:espin 1 complexes are dynamically associated with the treadmilling actin core. This model suggests that espin 1 is transported to the tips of stereocilia by myosin IIIa, whereupon it remains bound to the surface of the actin core for a period of time. Interestingly, abolishing or reducing myosin IIIa kinase activity enhances the affinity of the myosin IIIa for actin, providing further evidence that the kinase domain plays a role in regulating the myosin IIIa motor kinetics and actin-binding properties26, 27. While the myosin IIIa:espin 1 complex is tightly bound to actin, it travels back towards the base of the stereocilia along with the treadmilling actin core. In support of this model, live video imaging in transfected COS-7 cells shows fluorescent puncta of GFP-myosin IIIa ΔK and mCherry-ARD (Supplementary Information, Video S4) that move rearwards at rates matching the rates reported for actin treadmilling in filopodia (~0.5 µm/min)24. We suggest that these puncta are stably bound to the surface of the treadmilling actin filament bundle.
It is noteworthy that the stereocilia tips are also the site of mechanoelectrical transduction (MET)28, that the myosin IIIa developmental expression level is correlated with maturation of MET in stereocilia17, and that myosin IIIa has been shown to transport components of the photoreceptor transduction machinery in Drosophila29, 30. We cannot exclude the possibility that the localization and dynamics of the myosin IIIa:espin 1 complex are also affected by interactions with other proteins at the stereocilia tip. Furthermore, ankyrin repeats have been shown to be promiscuous binders of membrane proteins31. It is possible that the turnover and dynamic localization of the espin 1:myosin IIIa complex are influenced by interactions with components of the MET machinery, and vice-versa.
Affinity-purified polyclonal antibodies (PB538 and PB539) were developed in rabbits immunized with a synthetic peptide (Princeton Biomolecules, Langhorne, PA) corresponding to the amino acid sequence (LDALPVHHAARSGKLHCLR) of the first ankyrin repeat of mouse espin 1. A similarly raised antibody specific for a region conserved in all isoforms of espin (pan-espin, PB127)8 and anti-myosin IIIa (PB638)7 antibodies have been previously described.
Following CO2 anesthesia, rats, mice, and guinea pigs were euthanized in accordance to National Institutes of Health (NIH) guidelines, and their temporal bones fixed by immersion in 4% paraformaldehyde in phosphate buffered saline (PBS; pH 7.4) for 2 h at room temperature. Sensory tissue was dissected in PBS, permeabilized with 0.5% Triton X-100 for 30 min and blocked overnight at 4°C with 4% bovine serum albumin in PBS. Tissue was then incubated with primary antibody for 2 h, rinsed with PBS, stained with Alexa Fluor 488-conjugated secondary antibody (Molecular Probes, Eugene, OR) for 1 h, counterstained with 0.001 U/µl Alexa Fluor 568 phalloidin (Molecular Probes), and mounted using Prolong Antifade (Molecular Probes). Fluorescence confocal images were obtained with a Nikon microscope equipped with a 100× 1.45 numerical aperture (NA) objective and a spinning disk confocal unit (PerkinElmer, Wellesley, MA).
Rat organ of Corti or vestibular tissue were either rapidly frozen by contact with a liquid nitrogen cooled metal block in a LifeCell (The Woodlands, TX) freezing apparatus or fixed, glycerinated, and plunge frozen in Freon 22 cooled in liquid nitrogen prior to freeze-substitution in 1.5% uranyl acetate in absolute methanol at −90°C. Freeze-substituted tissues were infiltrated with Lowicryl HM-20 resin (Electron Microscopy Sciences, Hatfield, PA) at −45°C and polymerized with UV light, thin-sectioned, and immunogold labeled. Samples were viewed and photographed with a Zeiss 922 electron microscope (Peabody, MA). We also used as control for the immunogold labeling the antibody PB288 that is unrelated to espin or to myosin IIIa (Supplementary Figure 1d).
Espin 1 (NCBI accession number NM_031475) in pSPORT1 vector was obtained from imaGenes (Berlin, Germany) and PCR cloned into pEGFP-C2 (Clontech, Mountain View, CA) and pcDNA3.1(−) (Invitrogen, Carlsbad, CA) via EcoRI and KpnI sites. The site-directed leucine-to-alanine mutations in the WH2 motif of espin 1 were generated using a GeneTailor Site-Directed Mutagenesis kit (Invitrogen). The ankyrin repeats domain between amino acid positions 16 and 363 was PCR amplified using the mouse espin 1 template (NM_207687) and subcloned in-frame into the mCherry-C1 (Clontech) expression vector via XhoI and EcoRI sites, and pDEST™ 15 GST expression vector (Invitrogen) via the Gateway LR Clonase cloning method (Invitrogen). The GFP-tagged expression plasmids used were espin 1 ΔARD (a gift from Dr. James Bartles, Northwestern University, Chicago IL), myosin X (a gift of Dr. Richard Cheney, UNC, Chapel Hill NC), myosin XVa (a gift of Dr. Thomas Friedman, NIDCD/NIH), as well as full-length and deletion constructs of myosin IIIa that were generated in our laboratories. Myosin IIIa 2IQ ΔK constructs were generated as previously described (26, 34). Myosin IIIa 2IQ K50R and myosin IIIa K50R constructs were generated by performing site-directed mutagenesis on the myosin IIIa 2IQ and myosin IIIa full length constructs, respectively. All expression plasmids were sequence verified. Further details about the clones used are located in Supplementary Information, Table S1 and Figure 3.
COS-7 (ATCC, Manassas, VA) cells were plated on coverslips and maintained at 37°C in DMEM with 10% FBS. Cultures were transfected using GeneJuice Transfect Reagent (Novagen, San Diego, CA), incubated for 24 h. Time lapse videos of live cells were acquired at maximum nominal laser power and camera gain allowed by the confocal microscope. Samples were also fixed for 20 min in 4% paraformaldehyde in PBS, permeabilized for 30 min in 0.5% Triton X-100 in PBS, and counterstained or processed for immunofluorescence as described above.
Organ of Corti and vestibular tissue were dissected from postnatal day 0–4 rats and attached to coverslips previously coated with 150 µg/µl of Cell-Tak (BD Biosciences, San Jose, CA). Cultures were maintained in DMEM/F12 (Invitrogen) with 5–7% Fetal Bovine Serum (FBS) and 1.5 µg/ml ampicillin (Sigma, St. Louis, MO) and kept at 37°C and 5% CO2. For transfections, 50 µg of DNA was precipitated onto 25 mg of 1 µm gold particles and loaded into the Helios Gene Gun cartridges (BioRad, Hercules, CA). Tissue explants were transfected with the gene gun set at 95 psi of helium and maintained in culture for 18–48 hours. Samples were fixed and counterstained for confocal microscope viewing as described above. The efficiency of transfection ranged from 0 to 9 hair cells per explant.
Image analyses was performed with ImageJ software (NIH). To estimate the relative increase in stereocilia length we compared the heights of the tallest row of well-preserved stereocilia of cochlear and vestibular hair cells transfected (Ht) with the averaged height of all their respective neighboring (usually between 3 to 5) non-transfected cells (Hnt) within the field of view of our camera/confocal setup (30×45 µm). The average ratio of stereocilia length was calculated as . ANOVA analysis was performed using MATLAB (Mathworks, Nantick, MA). Cross-correlation analysis for the intensity plot in figure 3 was performed using Microsoft Excel.
100 mm dishes of transfected semi-confluent COS-7 cells were rinsed in PBS and scraped in 160 µl of lysis buffer: PBS, 1% Triton-X, 5 mM DTT, 1 mM Pefabloc, 5 µg/ml pepstatin A, 5 µg/ml leupeptin, 2 mM EDTA, 0.2 mM PMSF and 1% mammalian protease inhibitor cocktail (Sigma). After addition of 1x loading sample buffer and DTT (Invitrogen), samples were boiled and 10 µl of lysates were loaded in 4–12% Bis-Tris minigel (Invitrogen). Western blots were incubated overnight at 4°C with 4 µg/ml of the primary antibody. Horseradish peroxidase-conjugated goat anti-rabbit antibodies (Santa Cruz) and ECL chemiluminescence system (Amersham) were used for detection.
Protein expressions of glutathione S-transferase (GST) alone or fused to ankyrin repeats domain (GST-ARD) were optimized under L-arabinose induction in BL21-AI bacteria (Invitrogen). GST proteins were purified from bacterial extracts using glutathione–Sepharose 4B beads according to manufacturer's instructions (Amersham Biosciences, Buckinghamshire, UK). GFP-myosin IIIa ΔK, - pre3THDI, -3THDI, and -post3THDI proteins were extracted from 24 h COS-7 transfectants via brief sonication and 20-min ultracentrifugation at 145,000 g in ice-cold lysis buffer (1% Triton X-100, 5 mM DTT, 150 mM NaCl, 50 mM Tris pH 7.4, 2 mM EDTA, 3 mM Pefabloc SC, 1x Pefabloc (Roche, Indianapolis, IN), and 1x Mammalian Protease Inhibitor cocktail (Sigma). To test for myosin IIIa interactions, the same amount of GST-ARD or GST alone was bound to 4B beads for 1 h at 4°C followed by incubation with the GFP-tagged myosin IIIa fragment in CLB for 2 h. The beads were then washed four times with lysis buffer, and bound proteins separated by electrophoresis on NuPAGE Bis-Tris 4–12% gels (Invitrogen), and analyzed by Western blotting using rabbit polyclonal anti-GFP and anti-GST antibodies (Invitrogen).
The steady-state actin-activated ATPase activity of baculovirus expressed myosin IIIa 2IQ K50R and WT were performed with the NADH coupled assay26, 32. ATPase activity of fully phosphorylated myosin IIIa was compared to unphosphorylated myosin IIIa after a 60 minute incubation at room temperature in the presence and absence of 200 µM ATP, respectively. The kinase activity of the myosin IIIa 2IQ constructs were assayed using [32P]ATP or western blotting with antiphosphothreonine antibodies26, 32.
Specificity of the antibodies generated against the ARD of espin 1. (a) Immunoblots of lysates from COS-7 cells transfected with GFP-espin 1, GFP-ARD, and GFP-myoIIIa ΔK as well as lysates from bacteria expressing GST-ARD show that both PB538 and PB539 specifically recognize the ARD of espin 1. (b) GFP-espin 1 (left column) overexpressed in COS-7 cells is recognized by PB539. The same antibody shows no labeling in COS-7 cells with overexpressing GFP-myoIIIa ΔK only (right column). (c) In the negative control, our myosin IIIa specific antibody (PB638) fails to recognize GFP-espin 1 in COS-7 cells. GFP constructs are in green and Alexa Fluor 568-conjugated secondary antibody is red. Scale bar is 5 µm. (d) Control experiments for the post embedding immunogold labeling in directly frozen freeze-substituted adult rat cochlear hair cells shows no gold labeling at the tip of stereocilia when using the PB288 antibody, unrelated to espin 1 and myosin IIIa. Scale bars, 200 nm.
Schematic map of wild-type and deletion constructs. Only the myosin IIIa coding regions are shown. Full-length espin 1 is 871 amino acid-long in mouse and 854 amino acid-long in human (mouse espin 1 is illustrated here). The tail domain of myoIIIa encompasses 3THDI, 3THDII, and the third IQ domain, however; only 3THDI and 3THDII were considered in this study. Myosin IIIa Δ32 lacks exon 32, which causes a frame shift eliminating 3THDI and 3THDII, and introduces 26 new amino acids. Myosin IIIa Δ33,34 lacks exons 33 and 34, which causes a frame shift approximately one third of the way through 3THDI and therefore eliminates the rest of 3THDI and 3THDII, and introduces 72 new amino acids.
Actin-activated ATPase activity of myosin IIIa 2IQ constructs. a) A diagram of the myosin IIIa 2IQ constructs examined. b) The results for myosin IIIa 2IQ K50R (squares) are compared to that of myosin IIIa 2IQ wild-type 37 (open circles) and myosin IIIa 2IQ DKinase28 (diamonds). The steady-state ATPase rate of 0.1 mM myosin was measured using the NADH coupled assay in the presence of 1 mM ATP and a range of actin concentrations. The error bars represent the standard deviation from the mean and include data from 3-4 protein preparations. The enzymatic parameters determined from the fits to the data are summarized in the Table S2. Full phosphorylation of myosin IIIa 2IQ results in a ~ 40% reduction in ATPase activity compared to unphosphorylated myosin IIIa 2IQ, in the presence of 20 µM actin (data not shown). A previous study reported the kinetic mechanisms of the myosin IIIa 2IQ DKinase28 and myosin IIIa 2IQ WT37 constructs, which demonstrated the specific steps in motor ATPase cycle that are altered by the presence of the kinase domain. c) The kinase activity of 1 µM myosin IIIa 2IQ WT was monitored by 32P incorporation (200 µM [32P]ATP) over a 60 minute period (lanes 1-5 are 0, 5, 15, 30, and 60 minute time points, respectively). The results are compared to myosin IIIa 2IQ K50R that demonstrated little or no kinase activity under identical conditions (lanes 6-10 are 0, 5, 15, 30, and 60 minute time points, respectively). The top panel, labeled 32P, is the phosphorimage demonstrating the degree of 32P incorporation while the bottom panel, labeled C, is the same gel comassie stained to demonstrate that the total protein concentration in each lane is similar.
The first 24 amino acids of the 3THDI domain are sufficient for binding espin 1. (a) The mapping of the GFP-3THDI Δ33 fusion protein is illustrated. (b) GFP-3THDI Δ33 (green) colocalizes with espin 1 (red, labeled with PB539). PB539 (red) is labeled with Alexa Fluor 568-conjugated secondary antibody. Scale bar is 5 µm. (c) GST pull-down shows that GFP-3THDI Δ33 interacts with espin 1 ARD but not with GST.
Co-localization of myosin IIIa and espin 1 at filopodia tips. (a) Close up views of COS-7 cells show that espin 1 (red) colocalizes with GFP-myoIIIa ΔK (green) at the filopodia tips when these two proteins are coexpressed in COS-7 cells. (b) Espin 1 (red) colocalizes with GFP-myoIIIa ΔK (green) at the filopodia tips (actin is labeled Alexa fluor 647 shown in blue) when these two proteins are coexpressed in COS-7 cells. (c) Close up views of the regions outlined by rectangles in (b) show the increased concentration of GFP-myosin IIIa ΔK (arrows, green) and espin 1 (arrows, red) but not actin (arrowheads, blue) at the filopodia tips. Bars = 3µm
We thank Chi W. Pak for discussions and for the suggestion of mutations in the WH2 motif, Mark Schneider and Saeeda Latham for initial help with experiments and for discussions related to this work, Martin Horak for advice on cloning procedures, and Dr. Ronald Petralia for comments on the manuscript. This work was supported by NIDCD, DIR, NIH and in part by NIH grants # EY003575 to ACD and EY016419 to CMY.
PubMed Central Canada is a service of the Canadian Institutes of Health Research (CIHR) working in partnership with the National Research Council's Canada Institute for Scientific and Technical Information in cooperation with the National Center for Biotechnology Information at the U.S. National Library of Medicine(NCBI/NLM). It includes content provided to the PubMed Central International archive by participating publishers.

subset: commoncrawl


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Detection of Chlamydia trachomatis in fallopian tube tissue in women with postinfectious tubal infertility.
ABSTRACT Biopsy tissues from women with postinfectious tubal infertility were studied for the presence of Chlamydia trachomatis.
Tubal biopsy specimens from 25 women with postinfectious tubal infertility undergoing laparoscopy for repair of fallopian tubes were evaluated by culture, in situ hybridization. Immunocytochemistry, and transmission electron microscopy for the presence of Chlamydia trachomatis. Serum was also tested for Chlamydia trachomatis antibodies.
Chlamydia trachomatis was detected in postinfectious tubal biopsy specimens in three of 25 patients by culture, 12 of 24 by in situ hybridization, 15 of 22 by immunoperoxidase stain, and two of 10 by transmission electron microscopy. Serum antibody against Chlamydia trachomatis was detected in 15 of 21 patients.
Chlamydia trachomatis deoxyribonucleic acid or antigens were detected at a high percentage (19/24 women) in the biopsy tissues of the fimbrial and peritubal adhesions by in situ hybridization or immunoperoxidase stain, suggesting a persistent infection in these women even after antibiotic treatment.
- SourceAvailable from: Lyndsey Buckner[Show abstract] [Hide abstract]
ABSTRACT: Bacteria have evolved specific adaptive responses to cope with changing environments. These adaptations include stress response phenotypes with dynamic modifications of the bacterial cell envelope and generation of membrane vesicles (MVs). The obligate intracellular bacterium, Chlamydia trachomatis, typically has a biphasic lifestyle, but can enter into an altered growth state typified by morphologically aberrant chlamydial forms, termed persistent growth forms, when induced by stress in vitro. How C. trachomatis can adapt to a persistent growth state in host epithelial cells in vivo is not well understood, but is an important question, since it extends the host-bacterial relationship in vitro and has thus been indicated as a survival mechanism in chronic chlamydial infections. Here, we review recent findings on the mechanistic aspects of bacterial adaptation to stress with a focus on how C. trachomatis remodels its envelope, produces MVs, and the potential important consequences of MV production with respect to host-pathogen interactions. Emerging data suggest that the generation of MVs may be an important mechanism for C. trachomatis intracellular survival of stress, and thus may aid in the establishment of a chronic infection in human genital epithelial cells.Frontiers in Cellular and Infection Microbiology 06/2014; 4:73. DOI:10.3389/fcimb.2014.00073 · 2.62 Impact Factor
Article: Morphologic and molecular evaluation of Chlamydia trachomatis growth in human endocervix reveals distinct growth patterns[Show abstract] [Hide abstract]
ABSTRACT: In vitro models of Chlamydia trachomatis growth have long been studied to predict growth in vivo. Alternative or persistent growth modes in vitro have been shown to occur under the influence of numerous stressors but have not been studied in vivo. Here, we report the development of methods for sampling human infections from the endocervix in a manner that permits a multifaceted analysis of the bacteria, host and the endocervical environment. Our approach permits evaluating total bacterial load, transcriptional patterns, morphology by immunofluorescence and electron microscopy, and levels of cytokines and nutrients in the infection microenvironment. By applying this approach to two pilot patients with disparate infections, we have determined that their contrasting growth patterns correlate with strikingly distinct transcriptional biomarkers, and are associated with differences in local levels of IFNγ. Our multifaceted approach will be useful to dissect infections in the human host and be useful in identifying patients at risk for chronic disease. Importantly, the molecular and morphological analyses described here indicate that persistent growth forms can be isolated from the human endocervix when the infection microenvironment resembles the in vitro model of IFNγ-induced persistence.Frontiers in Cellular and Infection Microbiology 06/2014; 4:71. DOI:10.3389/fcimb.2014.00071 · 2.62 Impact Factor
Article: Commonly prescribed β-lactam antibiotics induce C. trachomatis persistence/stress in culture at physiologically relevant concentrations[Show abstract] [Hide abstract]
ABSTRACT: Chlamydia trachomatis, the most common bacterial sexually transmitted disease agent worldwide, enters a viable, non-dividing and non-infectious state (historically termed persistence and more recently referred to as the chlamydial stress response) when exposed to penicillin G in culture. Notably, penicillin G-exposed chlamydiae can reenter the normal developmental cycle upon drug removal and are resistant to azithromycin-mediated killing. Because penicillin G is less frequently prescribed than other β-lactams, the clinical relevance of penicillin G-induced chlamydial persistence/stress has been questioned. The goal of this study was to determine whether more commonly used penicillins also induce C. trachomatis serovar E persistence/stress. All penicillins tested, as well as clavulanic acid, induced formation of aberrant, enlarged reticulate bodies (RB) (called aberrant bodies or AB) characteristic of persistent/stressed chlamydiae. Exposure to the penicillins and clavulanic acid also reduced chlamydial infectivity by >95%. None of the drugs tested significantly reduced chlamydial unprocessed 16S rRNA or genomic DNA accumulation, indicating that the organisms were viable, though non-infectious. Finally, recovery assays demonstrated that chlamydiae rendered essentially non-infectious by exposure to ampicillin, amoxicillin, carbenicillin, piperacillin, penicillin V, and clavulanic acid recovered infectivity after antibiotic removal. These data definitively demonstrate that several commonly used penicillins induce C. trachomatis persistence/stress at clinically relevant concentrations.Frontiers in Cellular and Infection Microbiology 04/2014; 4:44. DOI:10.3389/fcimb.2014.00044 · 2.62 Impact Factor


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RECENT GALLERIES- PHOTOS: Rollins & Ambrose Debut New Ring Gear!
- PHOTOS: Seth Rollins Debuts New Ring Gear!
- EXCLUSIVE PHOTOS: RAW Script For June 2, 2014!
- PHOTO: Luke Gallows' New Real-Life Wife!
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— When a fan on Twitter asked if Steve Austin The Rock would be appearing on his podcast, Austin replied with the following…
@Gaurav9Kumar Never accepted the invite.
— Steve Austin (@steveaustinBSR) December 17, 2013


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Tvilum Pierce Office Storage Drawers with Two File Drawers in Beech
Tvilum Pierce Office Storage Drawers with Two File Drawers in Beech
Additional Documents
Weights & Dimensions
|Product Type: Drawers
|Country of Manufacture: Denmark
About the ManufacturerTvilum's business concept is to design, manufacture, and market high quality functional board based furniture, at competitive prices.
In order to be responsive to market trends and demands, Tvilum maintains a constant focus on product development. The phrase "two a day" reflects that at Tvilum, they design approx. 400 new models a year! Their range meets the demand for furniture for any room in the home, including home office furniture, storage furniture, wall systems, wardrobes, kitchen, children's furniture, etc. The furniture is supplied as ready-to-assemble kits in boxes containing all the parts necessary to enable the end-user to assemble a complete unit. The Tvilum Group today is Europe's leading supplier in its business area. It is their objective to further strengthen this position, with the aim of making Tvilum "the world's number one supplier of furniture to major retail customers". The Group has eight production plants in Denmark, with a total floor space of 3,000,000 square feet and more than 1,400 employees.
Tvilum Pierce Office Storage Drawers with Two File Drawers in Beech
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OUR SKU # TK2074Pierce Office Storage Drawers with Two File Drawers in Beech is also sometimes listed under the following manufacturer numbers: 8042641


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|Sq ft
|5 beds
|3.0 Ba
Visit us at www.yankeehomes.net
Large split-level home located on quiet residential street off of Lawrenceville Highway. This home features an upgraded kitchen, a beautiful stone fireplace in the living room, and a fully fenced back yard with deck. Three bedrooms and two full bathrooms on upper level with two additional bedrooms and full bath on lower level. Master bath features double vanities and natural stone tile shower. Front and rear decks provide area for entertaining or lounging. Home was recently renovated and includes new paint and appliances.
Close to Wal-Mart Supercenter, Home Depot and other shops nearby. Gwinnett Place Mall is just a 10 minute drive away. Easy access to I-85 (Pleasant Hill Road), Lawrenceville Highway, Ronald Reagan Parkway.
Visit us at www.yankeehomes.net
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Why Has Las Vegas Become a Home for Classic Rock?
For generations of music fans, Las Vegas has always been seen as the place where showbiz, the antithesis of rock, reigned supreme.
For generations of music fans, Las Vegas has always been seen as the place where showbiz, the antithesis of rock, reigned supreme.
Def Leppard will take off on its own cruise in January 2016.
They started the '80s on a relative low note, with the critical and commercial comedown of 1980's poorly received 'Panorama' album, but the Cars bounced back with 'Shake It Up' the following year -- and then they really hit their stride with 'Heartbeat City.'
AXS-TV will premiere a 2014 concert by Styx next month.
Def Leppard is taking a break from work on a long-awaited studio album to tour with Styx and Tesla, but don't expect any concert previews of their new songs.
Anyone who's done any amount of traveling knows it's difficult to stay healthy on the road.
For something that's supposed to be a huge honor, Rock and Roll Hall of Fame induction sure does get a lot of artists riled up.
Fans who turn out to see Def Leppard with Styx and Tesla this summer shouldn't expect to see any big set-closing jams.
The two bands will hit the shed circuit this summer with Tesla as the opener.
While they're waiting for the band's new album, Def Leppard fans in the U.S. can also look forward to summer tour dates -- at least two of them, anyway.
Journey will soon become the latest classic rock band to hit Sin City.
Following a difficult period of health concerns, Vivian Campbell has emerged happy, engaged and ever vigilant.


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Nili Lotan
Halter Maxi Dress
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Nili Lotan sheer black silk halter maxi dress.
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Posted| Updated
Researchers at the University of Canberra are attempting to find out why land use conflicts like coal seam gas and wind turbines cause so much angst in rural Australia.
Topics: rural, alternative-energy, environment, rivers, murray-darling-basin, university-and-further-education, education, agricultural-policy, government-and-politics, environmental-policy, community-and-society, community-development, community-education, mining-environmental-issues, mining-rural, mining-industry, agribusiness, university-of-canberra-2617
Professor Linda Botterill from the University of Canberra tells Lucy Barbour about the historical differences between the Liberal and National parties and why it's no surprise they're divided on the takeover of GrainCorp.
Topics: rural, government-and-politics, federal-government, business-economics-and-finance, agricultural-marketing, agribusiness, markets, industry, agricultural-crops, grain, multinationals, university-of-canberra-2617
Posted| Updated
A professor from the University of Canberra says the cracks with the Coalition over issues like foreign investment in Australian land and agriculture are nothing new.
Topics: rural, business-economics-and-finance, grain, agricultural-crops, agribusiness, agricultural-marketing, markets, industry, university-of-canberra-2617
Posted| Updated
The University of Canberra has announced it will bypass the union representing university academics and take a proposed pay offer direct to staff.
Topics: university-and-further-education, university-of-canberra-2617, act
Posted| Updated
One in every 13 Canberrans are struggling to pay for basics like food and housing, a new study has found.
Topics: poverty, act, university-of-canberra-2617, canberra-2600
Posted| Updated
Dire dry conditions come at a time when there is uncertainty about a national drought policy.
Topics: weather, drought, agricultural-crops, government-and-politics, agricultural-policy, university-of-canberra-2617, melbourne-3000, walgett-2832, qld
Posted| Updated
The University of Canberra is claiming a world-first, after developing a tool that is boosting rehabilitation efforts for hand transplant patients.
Topics: university-and-further-education, health, act, canberra-2600, university-of-canberra-2617
Posted| Updated
The Federal Labor Government has committed $10 million for the next stage of the University of Canberra sports hub project.
Topics: university-and-further-education, alp, act, university-of-canberra-2617
Posted| Updated
University of Canberra student Nathan Beer has returned from spending 20 cold nights homeless on the streets in Melbourne.
Topics: homelessness, act, canberra-2600, melbourne-3000, university-of-canberra-2617
Posted| Updated
ACT Chief Minister Katy Gallagher will travel to China in September to promote Canberra as a study destination.
Topics: university-and-further-education, states-and-territories, act, university-of-canberra-2617, australian-national-university-0200
Posted| Updated
Three language programs will close as the University of Canberra tightens its belt after recent funding cuts in the Federal Budget.
Topics: university-and-further-education, budget, university-of-canberra-2617, act
Posted| Updated
Human rights campaigner Tom Calma has been announced as the new chancellor of the University of Canberra.
Topics: university-and-further-education, human, act, university-of-canberra-2617, canberra-2600
Posted| Updated
The University of Canberra says it has created the first accurate snapshot of how human settlement impacted on bird life in the Pacific Islands.
Topics: research, birds, evolution, university-of-canberra-2617, act
Posted| Updated
The University of Canberra has unveiled an ambitious plan to become a world-ranked institution by 2018.
Topics: university-and-further-education, university-of-canberra-2617, act
Brumbies coach Jake White has been appointed an adjunct professor at the University of Canberra.
Topics: rugby-union, university-and-further-education, university-of-canberra-2617, act
The future of the Canberra Capitals coach remains unclear, as Carrie Graf announces she will be spending time off the court at the University of Canberra.
Topics: basketball, university-and-further-education, act, canberra-2600, university-of-canberra-2617
Posted| Updated
Hundreds of students are celebrating after receiving main round offers for courses at Canberra universities.
Topics: university-and-further-education, act, canberra-2600, university-of-canberra-2617, australian-national-university-0200
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The University of Canberra (UC) is expanding its pharmacy program, providing more options for students.
Topics: pharmaceuticals, university-and-further-education, act, canberra-2600, university-of-canberra-2617
A former University of Canberra (UC) lecturer who has spent years in custody waiting for the outcome of his trial, has now had his bail application delayed.
Topics: courts-and-trials, children, sexual-offences, act, canberra-2600, university-of-canberra-2617
The University of Canberra (UC) says it is coming under attack from a plague of rabbits.
Topics: university-and-further-education, animals, act, bruce-2617, university-of-canberra-2617, canberra-2600
Posted| Updated
An Indigenous woman from Queensland who was abused by a Catholic priest in the 1960s says the Royal Commission into child sexual abuse must examine the widespread abuse of Aboriginal children by members of the Catholic Church.
Topics: indigenous-aboriginal-and-torres-strait-islander, sexual-offences, federal-government, university-of-canberra-2617, act, rockhampton-4700, qld
Posted| Updated
The Australian National University and University of Canberra have increased their international student numbers, bucking the national trend.
Topics: university-and-further-education, australian-national-university-0200, act, university-of-canberra-2617
Posted| Updated
Construction is about to begin on a $50 million student accommodation project at the University of Canberra.
Topics: university-and-further-education, urban-development-and-planning, act, canberra-2600, university-of-canberra-2617
Posted| Updated
The University of Canberra will establish a presence in Melbourne next year in a partnership with a Victorian TAFE.
Topics: university-and-further-education, act, university-of-canberra-2617, chadstone-3148
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A treasure trove of children's classic literature is being dusted off and displayed in libraries across Canberra.
Topics: books-literature, states-and-territories, act, canberra-2600, university-of-canberra-2617


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Fundraisers - Page 3
Taco Fiesta Luncheon
UCM Update
The University of Central Missouri has organized a campus-wide relief effort for those affected by the tragic April 25 earthquake in Nepal. According to Shari Bax, vice provost for student experience and engagement at UCM, cash or check donations will be received from the campus community, as well as the local community, during the Fun Before Finals event planned for Thursday, April 30
MU Grad Donates $1 Million for LGBT Research And Education
Trail's End Western Heritage Days: Photos
Bothwell Regional Health Center Foundation Lub Dub Has Over 500 Participants
Overcast, only slightly cool – it was a great morning for run on Saturday, April 25, when the Bothwell Regional Health Center Foundation held its eighth annual Lub Dub.
More than 540 people registered for the 5K/10K walk and run, which is held annually to raise funds for the Camye Callis Gaspard Memorial Heart Fund for projects to help improve local treatment for cardiovascular disease
Trail's End Western Heritage Day's: Sunday Schedule
I hope you have enjoyed the first two days of the Trail's End Western Heritage Days. Today (Sunday) the first Trail's End Western Heritage Days finishes up.
This area is known for barbeque and the Kiwanis is putting on a competition in front of the Pepsi Grandstand through the morning


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Your little tadpole will spring into Halloween in the Froggy Toddler Costume. This little jumper is king of the leaf pads in this green jumpsuit with frog faced hood.
Look what just landed on a lily pad! Dress your little one in this adorable Lil Froggy baby costume and enjoy the ribbeting croaks and compliments. This cute costume features leg snaps for easy diaper changes and slip-on booties with skid resistant bottoms.


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Dissertations and Theses - Physics
Recent Additions
Epitaxial Aluminum Oxide Thin Films on Niobium (110): A Study of their Growth and Their Use in Superconducting Tunnel-Junctions (2007)Epitaxial hetero-structures have been turned to in recent years in an attempt to eliminate charge fluctuations in Josephson junction-based quantum bits. Amorphous tunnel-barriers have large numbers of two-level fluctuators ...
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(1988)This thesis presents the results of three studies of the excitonic system in Ge. These studies employ near-infrared spectroscopy and imaging techniques to examine the phase and transport properties of the low temperature ...
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Core-Level Photoemission Investigations of the Cadmium-Telluride(100) and Indium-Antimony(100) Surface and Interfacial Structures (1988)Photoemission techniques, utilizing a synchrotron light source, were used to analyze the clean (100) surfaces of the zinc-blende semiconductor materials CdTe and InSb. Several interfacial systems involving the surfaces of ...
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(1988)We consider the thermodynamical and hydrodynamical stability of proposed mechanisms for the dynamical hadronization transition in both ultra-relativistic heavy-ion collisions and the very early universe. We develop very ...
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(1988)In this thesis, light scattering techniques are used to probe the diverse excitation spectra of heavy fermion and high T$\sb{\rm c}$ superconductors.
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Kanie Hedgehog 238m - Kanie Hedgehog 238m - Page 2Kanie Hedgehog 238m
Advantages and Disadvantages While the attractive copper heatsink has it's advantages, it also does have it's fair share of disadvantages. First off there is a BIG problem with this weight spec that all AMD users should take note of; AMD state that their socket a processors can handle a maximum of 300 grams, meaning that this heatsink is too heavy for the processor and would most likely crush it, I wasn't going to try and risk killing a processor when AMD have already told us what would happen. So, even though the Hedgehog-238m can seat a socket a processor perfectly with the clips provided, I wouldn't recommend using it with one unless you have the money to replace your processor. I informed CoolPC Australia of this and they tell me that this heatsink works fine on all AMD socket a processors they have tested this heatsink on without damaging the processor, just be careful. The weight also has two other draw backs that I can think of. First, if using the heatsink with a slotket due it's weight it could have the potential to make the slotket lean and possibly come out of the slot. Second, if the heatsink happened to come lose and fall onto other hardware inside the case during transportation it would easy break, crack or snap chips and PCB material on the video card for example, uhh no what a nightmare! These hedgehog heatsinks are imported into Australia by CoolPC Australia from Kanie who are based in Japan. It is often argued which material is best for heatsinks. Copper can transfer heat faster than aluminum, but takes longer to dissipate heat. Aluminum doesn't transfer heat as fast, but it dissipates heat quicker than copper. So, many companies (including CoolPC Australia) who stock the Hedgehog-238m have been using more powerful fans such as the "black label" 38 CFM Delta fan to help dissipate the heat from the copper quicker thus allowing for better cooling performance at a higher cost to the user. Here we enter another disadvantage thanks to the copper material, because a more powerful fan (Delta black label 38 CFM) compared to the normal YS-Tech 27 CFM fan which is needed to dissipate the heat from the heatsink better. The Delta "black label" fan has hit my bad books, why? First it is VERY loud and it cut my finger which drew blood. This is the first fan which has actually cut my finger, I have touched other spinning fans before but they haven't cut me - I highly encourage CoolPC to include fan guards in the future! In fact, while I was measuring the RPM rating of this fan in the BIOS, it couldn't handle it and started reported lots of weird looking characters after it was spinning faster than 8000 RPM... I'd rather put up with lower cooling performance by using a less powerful, more quieter fan, it's too loud and will annoy you if it is in the same room as you. Below is a picture of the Delta black label fan with 3-pin power connector which means you can monitor the RPM of the fan...
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The road to the State Meet in La Crosse began for high school track and field athletes on Monday.
Regional Championships were decided in Girls Basketball Saturday night.
Local 5's Leslie Spoon features this week's scholar-athlete - Oshkosh West's wrestler Issac Rumbuc, who has recovered from a torn ACL to get back on the mat.
Local 5's Leslie Spoon has highlights from a busy night on the local high school hardwood.
Check out this full court shot from Oshkosh West's Ryan Sorensen vs. Kaukauna
Local 5's Ryan Rodig has highlights from the final week of the high school football regular season on our Xtra Point coverage, presented by Subway.
A big second half propelled the Appleton East Patriots to a 40-16 win over the Oshkosh West Wildcats Friday night.
Wisconsin recently adopted a new concussion bill that does not allow high school athletes to return to their sport after a concussion without written clearance by a health care provider.
Northeastern Wisconsin will be well represented in the WIAA Girls State Basketball Tournament next weekend in Madison.
In girls basketball Thursday night, the Kimberly Papermakers defeated the Oshkosh West Wildcats, 39-27.


text: I was mostly impressed by the very Moroccan and Spanish feel of this hotel. This is a great hotel and represents the culture of its town and country well. I loved where it is laced in the town but outside the square - it is peaceful. The staff is super-friendly, and are mostly willing to go out of their way... More


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Beach Jam for Breast Cancer taking the stage
Beach Jam for Breast Cancer happens on Saturday at the Westbrook Elks Club.
Weekdays. Mon – Friday from 5AM – 7AM and Weekends. Sat and Sunday from 6 AM – 8 AM
Beach Jam for Breast Cancer happens on Saturday at the Westbrook Elks Club.
Road trips with small children can be exhausting, but hopefully these simple screen-free DIY activities will keep your children engaged and …
This week's Report it Recap gets off to a thunderous start thanks to your pictures and videos.
Gil Simmons stopped by Middlebury to chat with some folks there and give his eight-day forecast.
Many people on Twitter voiced extreme opinions after a selfie of a girl smiling at a concentration camp surfaced.
Students at an Atlanta college have created micro-homes as part of an urban housing experiment.
Imagine suddenly finding yourself widowed at the young age of 27, with an 11-month old baby and $3 million dollars in debt…
Asthma Camp
On Good Morning Connecticut Saturday Gary Mendell, the founder of Shatterproof.org, and Len Wolman, chairman of Waterford Group, talked abou…
Tracy Weiss from Right at Home talks about the things people need to look out for with seniors and extreme heat.
Summer sun means summer flowers. Whether you live in an apartment or a home, flowers can really make your environment more enjoyable.
This week's Report It recap is brought to you by the letter S. As in sunrise, sunset, seashore, summer, and spectacular!
What a crowd and what a morning to be On the Green with Gil in North Haven.
A new report says Connecticut and Rhode Island have the highest percentage of poor road conditions in the entire country.
After cranking up our air conditioners during the recent hot weather, we're all sure to see a spike in our electric bills next month. Other …


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Home > News > Nation
Saturday June 21, 2014 MYT 12:00:00 AM
Saturday June 21, 2014 MYT 7:17:17 AM
Compiled by NG SI HOOI, HANIS ZAINAL AND A.RAMAN
ABOUT 50,000 students may be deprived of their National Higher Education Fund Corporation (PTPTN) loans when they enrol for varsity courses between July and December this year, reported Berita Harian.
This was because 183,000 people had defaulted on their PTPTN loans, involving a loss of funds involving RM1.3bil.
"Their failure to pay is serious as they represent 19% of the total number of PTPTN borrowers so the fund is losing RM1.3bil in future loans," a source told the Malay daily.
He said while the Finance Ministry had granted PTPTN a RM4bil allocation for this year, the amount could only cover 75% of the new and existing borrowers.
About 25% would not get the loans.
PTPTN, said the source, had suggested for defaulters to be blacklisted under Bank Negara's Central Credit Reference Information System (CCRIS) but this idea was rejected by the Cabinet.
"However, PTPTN may tighten the requirements for new borrowers to ensure that more people can get the loans," said the source.
> Harian Metro reported that the bungalow of a successful entrepreneur in Sungai Buloh was raided by enforcement authorities for selling sex stimulants containing banned substances.
The man, who had appeared several times on television and penned a few books, was widely considered to be an icon for entrepreneurship and believed to have earned tens of thousands of ringgit a month from selling the stimulants.
Selangor enforcement chief Adnan Salimin said the stimulants sold by the man, who is in his 50s, were found to contain sildenafil and steroid.
"The man has been selling the stimulant for over four years, most of which are aggressively marketed online via a network of distributors around the country," he said.
Raids were also carried out at the homes of two distributors in Shah Alam and Cheras, where officers seized RM200,000 worth of products.
Tags / Keywords:
Entertainment, Found in Translation, bookies, PTPTN loans, Berita Harian, Kosmo
Fish farm suspected of using animal carcasses as feed raided
'Live frugally to repay loan'
Gamblers turn to illegal bookies since gambling dens have been shut, says SAC
Rude shock for students eyeing PTPTN loans
Many vehicle owners fooled into thinking they are well-insured
Event celebrates the tasty ritual food of Malaysian communities
Najib: Lawyers looking at WSJ accusations
Najib: Would I steal and put money in country?
Najib to file suit against The Wall Street Journal on Tuesday
Celebrate Raya in style with GEMFIVE
The 3 unexpected life events that leave us in debt
50 students compete in three-on-three basketball contest
French, German presidents tried to influence World Cup votes - Blatter in paper
Soundcloud announces significant update for iOS app
Copyright © 1995-2015 Star Media Group Berhad (ROC 10894D)(Formerly known as Star Publications (Malaysia) Berhad)


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cARELESS MISTAKES
The Tech Savvy 2015 Nissan Sentra.
Choose Nissan
cARELESS MISTAKES
I really want to upload and edit the photos I took on my camera because I think I took some nice ones, but my dumb camera doesn't have the usb or the sd card! D: f this, I'll just stick to doing nothing for now. OTL
vocab quiz studying
antigone qs
ap bio studying
ap bio objectives
chinese hwk
bio ec
flashcards for wh
two math assignments
it is currently… 10:54 pm.
Choose Nissan
Okay sure that's fine if you're having a bad day, but if it's like that all the time? What, do you walk around with a stick up your booty? Dang, can't even talk to you or ask questions without some condescending remark.
I know you got things to say about me and pretty much everyone else too… why are you so hypocritical? You say everyone should get along blah blah blah but you go around talking about them. Oh ok makes sense. If there's anything I've learned this year, it's that no one likes a gossiper.


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Dr. Albert Krueger specializes in family practice in Meeker, Colorado.
Dr. Albert Krueger has been compared 33 times. Details of Dr. Krueger's experience as an MD are available on your free doctor report.
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Pioneers Medical Center
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Meeker, CO 81641
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A collection of news and information related to Gore Vidal published by this site and its partners.
Top Gore Vidal Articles
Displaying items 67-77
A letter to the class of 2016 (that I wrote in 2007)
Theaters On Political Wave? Not This Election in ConnecticutStory | Aug 29, 2012 | 8:34 AMWith the closing of Gore Vidal's "The Best Man" last week, the most politically direct show on Broadway has left the stage. It has also got me to thinking about theater and politics during these last few weeks of a major presidential campaign. I look...
Michael Wilson To Direct "Talley's Folly" In New York
Local places third in world in equestrian eventStory | Sep 11, 2012 | 7:51 PMBits and pieces from the local sports scene. Christine Hocutt-Senteney does well in equestrian event in Texas: Burbank resident Christine Hocutt-Senteney captured the third-place Farnam Select All-Around Amateur title at the 2012 Adequan Select...
Michael McKean makes 'break a leg' jokes after breaking leg
Watching the Tonys? Root For The Home Players
Tony Awards 2012: James Corden, Steve Kazee score lead actor upsets
Connie Wald dies at 96; Hollywood wife known for low-key parties
Great Old Broad Series: Elizabeth Ashley
Gore Vidal says nice things about women in the new Vanity Fair
Carnett: Speech by any other accent would sound as sweetStory | May 13, 2013 | 11:11 AMEver used a phony accent to fool somebody? Yeah, me too. When I was an 18-year-old college student, in 1963, my buddy Steve and I tried to impress girls at the beach with our English accents. We'd recently appeared in a Shakespearean production on...


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AquaCraft Motor Coupler
With your purchase, receive at no cost:
This is a replacement AquaCraft Motor Coupler. Package includes the motor connector, and three 3x3mm set screws.
NOTE: This motor coupler will not work with motors that have shafts larger than 3mm.
This product was added to our catalog on October 21, 2013.
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Available in select areas
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Bach Flower Remedies Essence Cherry Plum -- 0.7 fl oz
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Helps you act rationally & think clearly with a calm and balanced mind when you fear losing control.
"Fear of mind being over-strained, of reason giving away, of doing fearful and dreaded things, not wished and known wrong, yet there comes the through and impulse to do them"


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1 dead, 1 injured after robbery and Durham officer-involved shooting
DURHAM, N.C. (WNCN) – Durham Police are investigating an officer-involved shooting that happened early Saturday morning. Police responded to…
Skateboarder hit, killed by truck in Durham
DURHAM, NC. (WNCN) – Durham Police are investigating after a skateboarder was hit and killed while crossing the street early Saturday mornin…
5-year-old girl dies after being hit by sturgeon jumping into boat in Fla.
A 5-year-old Florida girl riding in a boat has died after a sturgeon leaped from the Suwannee River and struck her.
NC man throws 7-foot shark back in the surf
A viewer sent a WNCN sister station photos of a shark being thrown back out into the surf on Friday afternoon.
NC WEATHER: Mainly dry for fireworks Saturday
Here is the daily weather forecast from the WNCN meteorologists.
NC lawmakers may repeal cussing on highways ban
In 1913, North Carolina passed a law that banned cursing on a public road or highway within earshot of at least two people.
Parents banished teen girl to tent after she ate a Pop-Tart, SC deputies say
DALZELL, SC (WNCN) — The Sumter County Sheriff's Office arrested a husband and wife on child neglect charges Friday night after learning the…
UNC doctoral candidate looks to change perception of sharks
Sharks are in the news with recent attacks along the Carolina coast and the headlines are not positive for the sea creatures.
Bill Reh attempts to eat a hot dog in 5 seconds
While covering July Fourth festivities in Wake Forest, WNCN's Bill Reh decided to channel his inner-competitive eater while live on the air.
Black woman denied Raleigh library card in 1942 finally gets one
Pearl Thompson got a Wake County library card on Thursday in Raleigh.
Bubba Watson will remove Rebel flag on 'Dukes of Hazzard' car
Golfer Bubba Watson says he will paint an American flag where the Confederate flag is on the roof of the 'Dukes of Hazzard' car.
Pets more prone to run away July 4 weekend
Fireworks and loud sounds during the 4th of July weekend can be tough on your pets. Animal shelters say the days after 4th of July are the b…
Ft. Bragg solider arrested after carrying rifle into Fayetteville mall
A Fort Bragg sergeant faces charges after police said he walked through the Cross Creek mall with a rifle in his hand Thursday.
Hundreds celebrate Independence in Morrisville
Hundreds celebrate Independence Day a night early in Morrisville.
Oregon fines bakery owners for denying service to same-sex couple
Owners of a Portland-area bakery that denied service to a same-sex couple must pay $135,000 in damages.
Rolesville firefighter's simple idea could help locate a missing child
A Rolesville firefighter has come up with an idea to help identify and locate missing children.
Huge tree down in Raleigh near Cameron Village
A huge tree fell Friday on Raleigh's Van Dyke Avenue.
Pilot dies as medical center helicopter crashes in Colorado
A medical center helicopter crashed Friday in the Rocky Mountains just west of Denver.
2 Cuban baseball players defect while in Durham
Two Cuban baseball players have defected to the United States during a competition in Durham.
Photo shows 'Enjoy your ride, cuz we sure will' sign in Baltimore police van
A photo of a sticker allegedly posted in a Baltimore city police transport van reads, "Enjoy your ride, cuz we sure will!"
Official: Fayetteville man left in closet for 2 days before strangled, burned
Authorities say a 49-year-old Fayetteville man was beaten, tied up, and was left in a closet for as long as two days before he was strangled…
New leadership at Sweet Briar College
Changing of the guard at Sweet Briar College in Virginia
No makeup selfies go viral to stop cyberbullying
No-makeup selfies are going viral on social media.


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Police outside Kingsmeadow during yesterday's AFC Wimbledon match
/ News
A police officer is believed to have been assaulted as violence broke out at yesterday's football match between AFC Wimbledon and Bury at Kingsmeadow.
The fight is understood to have happened before the game kicked off, but a heavy police presence remained in and outside the stadium during the match as a result.
Two men were arrested - one for assaulting a police officer, and another man for affray.
Both were being held in custody yesterday.
A spokesman for Scotland Yard said an update would be provided tomorrow (Monday February 24).
Were you at the match? Did you see what happened?
Call the newsdesk on 020 87444255 or email firstname.lastname@example.com
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Two arrested as violence breaks out at AFC Wimbledon match
AFC Wimbledon promptly banned for life the 6 individuals who were involved in this minor fracas. They are to be congratulated for doing so. They highly value their reputation as one of the safest places to watch football in the league.
It's a shame that the Surrey Comet didn't rush to report this with as much haste as the original article.
It's a shame that the Surrey Comet didn't rush to report this with as much haste as the original article.


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Remembers when they let the AOL crowd onto USENET. His awesome wife also let him buy a Symbolics LispM and use a Sun 3/470MP as the T.V. stand in the living room in grad school.
The light purple never did match the decor.
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