Role of SUMO-1 and SUMO Interacting Motifs in Rhesus TRIM5α-mediated Restriction

Background TRIM5α is a member of the tripartite motif family of proteins that restricts retroviral infection in a species-specific manner. The restriction requires an interaction between the viral capsid lattice and the B30.2/SPRY domain of TRIM5α. Previously, we determined that two SUMO interacting motifs (SIMs) present in the B30.2/SPRY domain of human TRIM5α (huTRIM5α) were important for the restriction of N-tropic Murine Leukemia Virus. Here, we examined whether SUMO expression and the SIM1 and SIM2 motifs in rhesus monkey TRIM5α (rhTRIM5α) are similarly important for Human Immunodeficiency Type 1 (HIV-) restriction. Results We found that mutation of SIM1 and SIM2 of rhTRIM5α abolished the restriction of HIV-1 virus. Further, knockdown of SUMO-1 in rhTRIM5α expressing cells abolished restriction of HIV-1. These results may be due, in part, to the ability of SUMO-1 to stabilize rhTRIM5α protein expression, as SUMO-1 knockdown increased rhTRIM5α turnover and the mutations in SIM1 and SIM2 led to more rapid degradation than the wild type protein. The NF-κB signaling ability of rhTRIM5α was also attenuated by SUMO-1 knockdown. Finally, upon inhibition of CRM1-dependent nuclear export with Leptomycin B (LMB), wild type rhTRIM5α localized to SUMO-1 bodies in the nucleus, while the SIM1 and SIM2 mutants did not localize to SUMO-1. Conclusions Our results suggest that the rhTRIM5α B30.2/SPRY domain is not only important for the recognition of the HIV-1 CA, but it is also important for its association with SUMO-1 or SUMO-1 modified proteins. These interactions help to maintain TRIM5α protein levels and its nuclear localization into specific nuclear bodies

Nucleic acids and growth of \u3cem\u3eCalanus finmarchicus\u3c/em\u3e in the laboratory under different food and temperature conditions

We examined the effects of food concentration and temperature on nucleic acids and protein content of Calanus finmarchicus in order to evaluate the use of RNA as a growth rate index for this species. We measured RNA, DNA, and protein content of copepods reared from egg to adult stage in 5 combinations of food and temperature conditions (25 to 500 µg C l-1, 4 to 12°C). At 8°C, DNA, RNA and protein content and RNA:DNA differed among food treatments during Stages N6 through to adult female. Protein:DNA ratios and RNA:protein ratios were significantly different among food levels for only 3 of the 8 stages examined. At excess food, DNA, RNA, and protein content and RNA:DNA ratios were inversely related to temperature for most stages from C1 onward, but the effect of temperature was relatively small over the range of temperatures investigated. The RNA:DNA and protein:DNA ratios increased with developmental stage whereas the RNA:protein ratio and growth rates (measured in terms of protein, nitrogen, DNA, and carbon content) declined with increasing stage. Although the relationship of RNA:DNA to growth rates was stage-specific, the two were related when standardized for temperature and developmental stage. RNA:protein ratios were directly related to growth rates regardless of stage, and the slope of the relationship increased with increasing temperature in a nonlinear fashion. Our results emphasize the importance of temperature and developmental stage for the relationship of growth rates to RNA concentration and RNA:DNA ratios. We propose 2 ways to estimate in situ growth rates of C. finmarchicus from RNA:DNA or RNA:protein ratios and environmental temperatur

Assembly of the outer retina in the absence of GABA synthesis in horizontal cells

<p>Abstract</p> <p>Background</p> <p>The inhibitory neurotransmitter gamma-amino-butyric acid (GABA) not only modulates excitability in the mature nervous system but also regulates neuronal differentiation and circuit development. Horizontal cells, a subset of interneurons in the outer retina, are transiently GABAergic during the period of cone photoreceptor synaptogenesis. In rodents, both horizontal cells and cone axonal terminals express GABA_Areceptors. To explore the possibility that transient GABA expression in mouse neonatal horizontal cells influences the structural development of synaptic connectivity in the outer retina, we examined a mutant in which expression of GAD67, the major synthesizing enzyme for GABA, is selectively knocked out in the retina.</p> <p>Results</p> <p>Immunocytochemistry and electron microscopy revealed that the assembly of triad synapses involving cone axonal pedicles and the dendrites of horizontal and bipolar cells is unaffected in the mutant retina. Moreover, loss of GABA synthesis in the outer retina did not perturb the spatial distributions and cell densities of cones and horizontal cells. However, there were some structural alterations at the cellular level: the average size of horizontal cell dendritic clusters was larger in the mutant, and there was also a small but significant increase in cone photoreceptor pedicle area. Moreover, metabotropic glutamate receptor 6 (mGluR6) receptors on the dendrites of ON bipolar cells occupied a slightly larger proportion of the cone pedicle in the mutant.</p> <p>Conclusions</p> <p>Together, our analysis shows that transient GABA synthesis in horizontal cells is not critical for synapse assembly and axonal and dendritic lamination in the outer retina. However, pre- and postsynaptic structures are somewhat enlarged in the absence of GABA in the developing outer retina, providing for a modest increase in potential contact area between cone photoreceptors and their targets. These findings differ from previous results in which pharmacological blockade of GABA_Areceptors in the neonatal rabbit retina caused a reduction in cone numbers and led to a grossly disorganized outer retina.</p

Growth and development rates of the copepod \u3cem\u3eCalanus finmarchicus\u3c/em\u3e reared in the laboratory

Development rates, nitrogen- and carbon-specific growth rates, size, and condition were determined for the copepod Calanus finmarchicus reared at 3 temperatures (4, 8, and 12°C) at non-limiting food concentrations and 2 limiting food concentrations at 8°C in the laboratory. Development rates were equiproportional, but not isochronal. Naupliar stage durations were similar, except for non-feeding stages, which were of short duration, and the first feeding stage, which was prolonged, while copepodite stage durations increased with increasing stage of development. Under limiting food concentrations at 8°C, development rates were prolonged but similar relative patterns in stage durations were observed. Body size (length and weight) was inversely related to temperature and positively related to food concentration. Condition measurements were not affected by temperature, but were positively related to food concentration. Growth rates increased with increasing temperature and increased asymptotically with increasing food concentration. At high food concentrations, growth rates of naupliar stages were high (except for individuals molting from the final naupliar stage to the first copepodite stage, in which growth rates were depressed), while growth of copepodites decreased with increasing stage of development. Neither nitrogen nor carbon growth rates, the former a proxy for structural growth, were exponential over the entire life cycle, but rather sigmoidal. Carbon-specific growth rates were greater than nitrogen-specific growth rates, and this difference increased with increasing stage of development, reflecting an augmentation in lipid deposition in the older stages. However, nitrogen and carbon growth rates were more similar under food-limited conditions. Based on this study, we recommend that secondary production rates of Calanus finmarchicus and possibly other lipid-storing copepods not be estimated from egg production measurements alone, as has been suggested for other species of copepods, because growth, including structural growth, is not equivalent for all stages

TRIM5α associates with proteasomal subunits in cells while in complex with HIV-1 virions

<p>Abstract</p> <p>Background</p> <p>The TRIM5 proteins are cellular restriction factors that prevent retroviral infection in a species-specific manner. Multiple experiments indicate that restriction activity requires accessory host factors, including E2-enzymes. To better understand the mechanism of restriction, we conducted yeast-two hybrid screens to identify proteins that bind to two TRIM5 orthologues.</p> <p>Results</p> <p>The only cDNAs that scored on repeat testing with both TRIM5 orthologues were the proteasome subunit PSMC2 and ubiquitin. Using co-immunoprecipitation assays, we demonstrated an interaction between TRIM5α and PSMC2, as well as numerous other proteasome subunits. Fluorescence microscopy revealed co-localization of proteasomes and TRIM5α cytoplasmic bodies. Forster resonance energy transfer (FRET) analysis indicated that the interaction between TRIM5 and PSMC2 was direct. Previous imaging experiments demonstrated that, when cells are challenged with fluorescently-labeled HIV-1 virions, restrictive TRIM5α orthologues assemble cytoplasmic bodies around incoming virion particles. Following virus challenge, we observed localization of proteasome subunits to rhTRIM5α cytoplasmic bodies that contained fluorescently labeled HIV-1 virions.</p> <p>Conclusions</p> <p>Taken together, the results presented here suggest that localization of the proteasome to TRIM5α cytoplasmic bodies makes an important contribution to TRIM5α-mediated restriction.</p

Mental Health Professionals’ Attitudes toward Clients with Antisocial Personality Disorder: An Exploratory Study

This exploratory study examined mental health professionals’ attitudes toward clients with antisocial personality disorder. Specifically, are mental health professionals’ attitudes influenced by (a) personal experiences with criminal victimization, or (b) contact with clients with antisocial personality disorder. A factorial MANOVA and follow-up univariate ANOVAs revealed a statistically significant main effect in relation to participants’ level of clinical contact with clients having antisocial personality disorder. Participants with higher levels of clinical contact were associated with more positive attitudes towards clients. Implications for mental health professionals, supervisors, and counselor educators are discussed, and suggestions for future research are provided

Hubble Space Telescope Planetary Camera images of R136

The Planetary Camera of the Hubble Space Telescope has been used to obtain broad and narrowband images ofR136, the core of the massive star cluster 30 Doradus in the Large Magellanic Cloud. R136a, the brightest component ofR136, is shown to have at least 12 separate components, including the eight originally identified by speckle interferometry. Three of the 12 components are previously unidentified close companions of the speckle components. The stars within R136a are found to have luminosities and colors of normal evolved (Wolf-Rayet and blue supergiants) and zero-age main-sequence (ZAMS) massive stars. A narrowband He II filter was used to investigate the Wolf-Rayet stellar population. We find that three stars in R136a are of the Wolf-Rayet type; of the two identified from ground-based data, one is now resolved into two components. We present color-magnitude diagrams and a luminosity function of the stars within the larger region (~2 pc) defined as R136. We find that the stars in R136 are similar in color and luminosity to those of cluster members that lie outside that crowded inner region. The lower end of the color-magnitude diagram corresponds to ZAMS spectral type B3. No red supergiants have been detected within R136. The luminosity per unit area in the inner 1" (0.25 pc) of R136 is ≥ 50 times that of the center of Orion for a comparable area and seven times that of the core of NGC 3603. The luminosity per unit area of all of R136 is comparable to that of Orion but is sustained over 130 times the area. An F336W surface brightness profile is constructed for R136 based on the stellar photometry. The distribution is found to be consistent with a pure power law with l(r}ɑ r^y with y=-1.72±0.06 or with a small core with r_c 5 X 10^4 M_☉ pc^(-3). The implied upper limit on the relaxation time for the cluster is much smaller than the age of 3.5 X 10^6 yrs required by the presence of Wolf-Rayet stars. This suggests that relaxation effects have been very important in determining the observed structure of the cluster unless a large population of lower mass stars is also present

Imaging of the gravitational lens system PG 1115+080 with the Hubble Space Telescope

This paper is the first of a series presenting observations of gravitational lenses and lens candidates, taken with the Wide Field/Planetary Camera (WFPC) of the Hubble Space Telescope (HST). We have resolved the gravitational lens system PG 1115 + 080 into four point sources and a red, extended object that is presumably the lens galaxy; we present accurate relative intensities, colors, and positions of the four images, and lower accuracy intensity and position of the lens galaxy, all at the epoch 1991.2. Comparison with earlier data shows no compelling evidence for relative intensity variations between the QSO components having so far been observed. The new data agree with earlier conclusions that the system is rather simple, and can be produced by the single observed galaxy. The absence of asymmetry in the HST images implies that the emitting region of the quasar itself has an angular radius smaller than about 10 milliarcsec (100 pc for H_0=50, q_0=0.5)