Monocyte and macrophage activation by lipoteichoic Acid is independent of alanine and is potentiated by hemoglobin

Lipoteichoic acids (LTAs) are Gram-positive bacterial cell wall components that elicit mononuclear cell cytokine secretion. Cytokine-stimulating activity is thought to be dependent on retaining a high level of ester-linked D-alanine residues along the polyglycerol phosphate backbone. However, Streptococcus pyogenes LTA essentially devoid of D-alanine caused human and mouse cells to secrete as much IL-6 as LTA with a much higher D-alanine content. Furthermore, hemoglobin (Hb) markedly potentiates the stimulatory effect of various LTAs on mouse macrophages or human blood cells, regardless of their d-alanine content. LTA and Hb appear to form a molecular complex, based on the ability of each to affect the other's migration on native acrylamide gels, their comigration on these gels, and the ability of LTA to alter the absorption spectra of Hb. Because S. pyogenes is known to release LTA and secrete at least two potent hemolytic toxins, LTA-Hb interactions could occur during streptococcal infections and might result in a profound alteration of the local inflammatory response.     

Force spectroscopy of barnase-barstar single molecule interaction

Results of the single molecule force spectroscopy study of specific interactions between ribonuclease barnase and its inhibitor barstar are presented. Experimental data obtained for the force loading rate ranging 2-70 nN/s are well approximated by a single straight line, from which the dissociation barrier of the width of 0.12 nm and height of 0.75-0.85 × 10(-19)J can be inferred. The measured value of specific interaction does not depend on the NaCl concentration. This apparently contradicts the well-known dependence of the binding energy of this pair on the salt concentration, but such a "contradiction" is explained by the insensitivity of the force spectroscopy data to the relatively long-range electrostatic interaction. The latter essentially contributes to the value of barnase-barstar binding energy revealed by biochemical measurements, and it is exactly this electrostatic interaction which is influenced by the salt concentration.     

More than just sugars: Conserved oligomeric Golgi complex deficiency causes glycosylation‐independent cellular defects

The conserved oligomeric Golgi (COG) complex controls membrane trafficking and ensures Golgi homeostasis by orchestrating retrograde vesicle trafficking within the Golgi. Human COG defects lead to severe multisystemic diseases known as COG‐congenital disorders of glycosylation (COG‐CDG). To gain better understanding of COG‐CDGs, we compared COG knockout cells with cells deficient to 2 key enzymes, Alpha‐1,3‐mannosyl‐glycoprotein 2‐beta‐N‐acetylglucosaminyltransferase and uridine diphosphate‐glucose 4‐epimerase (GALE), which contribute to proper N‐ and O‐glycosylation. While all knockout cells share similar defects in glycosylation, these defects only account for a small fraction of observed COG knockout phenotypes. Glycosylation deficiencies were not associated with the fragmented Golgi, abnormal endolysosomes, defective sorting and secretion or delayed retrograde trafficking, indicating that these phenotypes are probably not due to hypoglycosylation, but to other specific interactions or roles of the COG complex. Importantly, these COG deficiency specific phenotypes were also apparent in COG7‐CDG patient fibroblasts, proving the human disease relevance of our CRISPR knockout findings. The knowledge gained from this study has important implications, both for understanding the physiological role of COG complex in Golgi homeostasis in eukaryotic cells, and for better understanding human diseases associated with COG/Golgi impairment.      

The Fibroblast Growth Factor 14·Voltage-gated Sodium Channel Complex Is a New Target of Glycogen Synthase Kinase 3 (GSK3)

The FGF14 protein controls biophysical properties and subcellular distribution of neuronal voltage-gated Na⁺ (Nav) channels through direct binding to the channel C terminus. To gain insights into the dynamic regulation of this protein/protein interaction complex, we employed the split luciferase complementation assay to screen a small molecule library of kinase inhibitors against the FGF14·Nav1.6 channel complex and identified inhibitors of GSK3 as hits. Through a combination of a luminescence-based counter-screening, co-immunoprecipitation, patch clamp electrophysiology, and quantitative confocal immunofluorescence, we demonstrate that inhibition of GSK3 reduces the assembly of the FGF14·Nav channel complex, modifies FGF14-dependent regulation of Na⁺ currents, and induces dissociation and subcellular redistribution of the native FGF14·Nav channel complex in hippocampal neurons. These results further emphasize the role of FGF14 as a critical component of the Nav channel macromolecular complex, providing evidence for a novel GSK3-dependent signaling pathway that might control excitability through specific protein/protein interactions.     

Completing the family portrait of the anti-apoptotic Bcl-2 proteins: crystal structure of human Bfl-1 in complex with Bim

Evasion of apoptosis is recognized as a characteristic of malignant growth. Anti-apoptotic B-cell lymphoma-2 (Bcl-2) family members have therefore emerged as potential therapeutic targets due to their critical role in proliferating cancer cells. Here, we present the crystal structure of Bfl-1, the last anti-apoptotic Bcl-2 family member to be structurally characterized, in complex with a peptide corresponding to the BH3 region of the pro-apoptotic protein Bim. The structure reveals distinct features at the peptide-binding site, likely to define the binding specificity for pro-apoptotic proteins. Superposition of the Bfl-1:Bim complex with that of Mcl-1:Bim reveals a significant local plasticity of hydrophobic interactions contributed by the Bim peptide, likely to be the basis for the multi specificity of Bim for anti-apoptotic proteins.     

Slowing of the HIV Epidemic in Ukraine: Evidence from Case Reporting and Key Population Surveys, 2005–2012

Background

Ukraine developed Europe''s most severe HIV epidemic due to widespread transmission among persons who inject drugs (PWID). Since 2004, prevention has focused on key populations; antiretroviral therapy (ART) coverage has increased. Recent data show increases in reported HIV cases through 2011, especially attributed to sexual transmission, but also signs of potential epidemic slowing. We conducted a data triangulation exercise to better analyze available data and inform program implementation.

Methods and Findings

We reviewed data for 2005 to 2012 from multiple sources, primarily national HIV case reporting and integrated biobehavioral surveillance (IBBS) studies among key populations. Annually reported HIV cases increased at a progressively slower rate through 2011 with recent increases only among older, more immunosuppressed individuals; cases decreased 2.7% in 2012. Among women <25 years of age, cases attributed to heterosexual transmission and HIV prevalence in antenatal screening declined after 2008. Reported cases among young PWID declined by three-fourths. In 2011, integrated biobehavioral surveillance demonstrated decreased HIV prevalence among young members of key populations compared with 2009. HIV infection among female sex workers (FSW) remains strongly associated with a personal history of injecting drug use (IDU).

Conclusions

This analysis suggests that Ukraine''s HIV epidemic has slowed, with decreasing reported cases and older cases predominating among those diagnosed. Recent decreases in cases and in prevalence support decreased incidence among young PWID and women. Trends among heterosexual men and men who have sex with men (MSM) are less clear; further study and enhanced MSM prevention are needed. FSW appear to have stable prevalence with risk strongly associated with IDU. Current trends suggest the Ukrainian epidemic can be contained with enhanced prevention among key populations and increased treatment access.     

γ-Tubulin 2 nucleates microtubules and is downregulated in mouse early embryogenesis

γ-Tubulin is the key protein for microtubule nucleation. Duplication of the γ-tubulin gene occurred several times during evolution, and in mammals γ-tubulin genes encode proteins which share ～97% sequence identity. Previous analysis of Tubg1 and Tubg2 knock-out mice has suggested that γ-tubulins are not functionally equivalent. Tubg1 knock-out mice died at the blastocyst stage, whereas Tubg2 knock-out mice developed normally and were fertile. It was proposed that γ-tubulin 1 represents ubiquitous γ-tubulin, while γ-tubulin 2 may have some specific functions and cannot substitute for γ-tubulin 1 deficiency in blastocysts. The molecular basis of the suggested functional difference between γ-tubulins remains unknown. Here we show that exogenous γ-tubulin 2 is targeted to centrosomes and interacts with γ-tubulin complex proteins 2 and 4. Depletion of γ-tubulin 1 by RNAi in U2OS cells causes impaired microtubule nucleation and metaphase arrest. Wild-type phenotype in γ-tubulin 1-depleted cells is restored by expression of exogenous mouse or human γ-tubulin 2. Further, we show at both mRNA and protein levels using RT-qPCR and 2D-PAGE, respectively, that in contrast to Tubg1, the Tubg2 expression is dramatically reduced in mouse blastocysts. This indicates that γ-tubulin 2 cannot rescue γ-tubulin 1 deficiency in knock-out blastocysts, owing to its very low amount. The combined data suggest that γ-tubulin 2 is able to nucleate microtubules and substitute for γ-tubulin 1. We propose that mammalian γ-tubulins are functionally redundant with respect to the nucleation activity.     

The role of some donor-host cell interactions in conditions of the regenerative process microenvironment

The review is devoted to the analysis of experimental data about possible mechanisms of transdifferentiation or plasticity of tissue specific stem cells. Considerable attention is focused on the mechanisms and genetic consequences of fusion of different types of donor cells with the cells of recipient tissues which investigated on the models of cellular therapy of liver and heart diseases. The role of various kinds of cell contacts and their role in stem cells integration, reparation and regeneration of injured tissue and horizontal genes transfer are considered.