Cytochrome c dysregulation induced by HIV infection of astrocytes results in bystander apoptosis of uninfected astrocytes by an IP3 and calcium‐dependent mechanism

HIV entry into the CNS is an early event after peripheral infection, resulting in neurologic dysfunction in a significant number of individuals despite successful anti‐retroviral therapy. The mechanisms by which HIV mediates CNS dysfunction are not well understood. Our group recently demonstrated that HIV infection of astrocytes results in survival of HIV infected cells and apoptosis of surrounding uninfected astrocytes by the transmission of toxic intracellular signals through gap junctions. In the current report, we characterize the intracellular signaling responsible for this bystander apoptosis. Here, we demonstrate that HIV infection of astrocytes results in release of cytochrome C from the mitochondria into the cytoplasm, and dysregulation of inositol trisphosphate/intracellular calcium that leads to toxicity to neighboring uninfected astrocytes. Blocking these dysregulated pathways results in protection from bystander apoptosis. These secondary messengers that are toxic in uninfected cells are not toxic in HIV infected cells, suggesting that HIV protects these cells from apoptosis. Thus, our data provide novel mechanisms of HIV mediated toxicity and generation of HIV reservoirs. Our findings provide new potential therapeutic targets to reduce the CNS damage resulting from HIV infection and to eradicate the generation of viral reservoirs.

     

Modulation of Lipid Kinase PI4KIIα Activity and Lipid Raft Association of Presenilin 1 Underlies γ-Secretase Inhibition by Ginsenoside (20S)-Rg3

Amyloid β-peptide (Aβ) pathology is an invariant feature of Alzheimer disease, preceding any detectable clinical symptoms by more than a decade. To this end, we seek to identify agents that can reduce Aβ levels in the brain via novel mechanisms. We found that (20S)-Rg3, a triterpene natural compound known as ginsenoside, reduced Aβ levels in cultured primary neurons and in the brains of a mouse model of Alzheimer disease. The (20S)-Rg3 treatment induced a decrease in the association of presenilin 1 (PS1) fragments with lipid rafts where catalytic components of the γ-secretase complex are enriched. The Aβ-lowering activity of (20S)-Rg3 directly correlated with increased activity of phosphatidylinositol 4-kinase IIα (PI4KIIα), a lipid kinase that mediates the rate-limiting step in phosphatidylinositol 4,5-bisphosphate synthesis. PI4KIIα overexpression recapitulated the effects of (20S)-Rg3, whereas reduced expression of PI4KIIα abolished the Aβ-reducing activity of (20S)-Rg3 in neurons. Our results substantiate an important role for PI4KIIα and phosphoinositide modulation in γ-secretase activity and Aβ biogenesis.     

Glycine and Folate Ameliorate Models of Congenital Sideroblastic Anemia

Sideroblastic anemias are acquired or inherited anemias that result in a decreased ability to synthesize hemoglobin in red blood cells and result in the presence of iron deposits in the mitochondria of red blood cell precursors. A common subtype of congenital sideroblastic anemia is due to autosomal recessive mutations in the SLC25A38 gene. The current treatment for SLC25A38 congenital sideroblastic anemia is chronic blood transfusion coupled with iron chelation. The function of SLC25A38 is not known. Here we report that the SLC25A38 protein, and its yeast homolog Hem25, are mitochondrial glycine transporters required for the initiation of heme synthesis. To do so, we took advantage of the fact that mitochondrial glycine has several roles beyond the synthesis of heme, including the synthesis of folate derivatives through the glycine cleavage system. The data were consistent with Hem25 not being the sole mitochondrial glycine importer, and we identify a second SLC25 family member Ymc1, as a potential secondary mitochondrial glycine importer. Based on these findings, we observed that high levels of exogenous glycine, or 5-aminolevulinic acid (5-Ala) a metabolite downstream of Hem25 in heme biosynthetic pathway, were able to restore heme levels to normal in yeast cells lacking Hem25 function. While neither glycine nor 5-Ala could ameliorate SLC25A38 congenital sideroblastic anemia in a zebrafish model, we determined that the addition of folate with glycine was able to restore hemoglobin levels. This difference is likely due to the fact that yeast can synthesize folate, whereas in zebrafish folate is an essential vitamin that must be obtained exogenously. Given the tolerability of glycine and folate in humans, this study points to a potential novel treatment for SLC25A38 congenital sideroblastic anemia.     

Differences between integrin α5β1 and EGRF receptor in signal pathways controlling proliferation and apoptosis of MCF-7/Dox human breast carcinoma cells

In MCF-7/Dox human breast carcinoma cells, down-regulation of integrin α5β1 and inhibition of epidermal growth factor receptor (EGFR) markedly reduced cell proliferation. Cell cycle analysis showed that α5β1 down-regulation resulted in cycle arrest at the S-phase, followed by a significant increase in the population of apoptotic cells (subG₁ population). Inhibition of EGFR activity also caused cell cycle arrest at the S-phase but without any increase in the subG₁ population. Down-regulation of α5β1 and EGFR inhibition resulted in a significant decrease of cell content of the active (phosphorylated) forms of FAK and Erk protein kinases. The data obtained suggest that α5β1 integrin is implicated in cell growth control via inhibition of apoptotic cell death and through EGFR activation.     

Cold hardiness and range of the earthworm <Emphasis Type="Italic">Eisenia sibirica</Emphasis> (Oligochaeta,Lumbricidae)

A hypothesis of range formation of the earthworm Eisenia sibirica Perel et Graphodatsky 1984, which is an endemic species of the Altai–Sayan mountain system and is also found on the adjacent plains of Siberia across the valleys of the rivers, is suggested. The limited distribution of the species can be connected with the insufficient cold hardiness of the worm stage (–10 to–12°C). The plains of Western Siberia lie in an area of minimum soil temperature isotherms at a depth of 3 cm:–12 to–14°C, i.e., on average 2–4°C below the tolerable limits for this species. Foothill and mountain soils are warmer, since they obtain much more solid precipitation. Low soil temperatures of the plains apparently “lock up” this species within the Altai–Sayan system. At the same time, there are reasons to consider the northernmost locations of E. sibirica to be relict.     

Analyzing the Long Term Cohesive Effect of Sector Specific Driving Forces

Financial markets are partially composed of sectors dominated by external driving forces, such as commodity prices, infrastructure and other indices. We characterize the statistical properties of such sectors and present a novel model for the coupling of the stock prices and their dominating driving forces, inspired by mean reverting stochastic processes. Using the model we were able to explain the market sectors’ long term behavior and estimate the coupling strength between stocks in financial markets and the sector specific driving forces. Notably, the analysis was successfully applied to the shipping market, in which the Baltic dry index (BDI), an assessment of the price of transporting the major raw materials by sea, influences the shipping financial market. We also present the analysis of other sectors—the gold mining market and the food production market, for which the model was also successfully applied. The model can serve as a general tool for characterizing the coupling between external forces and affected financial variables and therefore for estimating the risk in sectors and their vulnerability to external stress.     

A high‐throughput capillary isoelectric focusing immunoassay for fingerprinting protein sialylation

The serum half‐life, biological activity, and solubility of many recombinant glycoproteins depend on their sialylation. Monitoring glycoprotein sialylation during cell culture manufacturing is, therefore, critical to ensure product efficacy and safety. Here a high‐throughput method for semi‐quantitative fingerprinting of glycoprotein sialylation using capillary isoelectric focusing immunoassay on NanoPro (Protein Simple) platform was developed. The method was specific, sensitive, precise, and robust. It could analyze 2 μL of crude cell culture samples without protein purification, and could automatically analyze from 8 samples in 4 h to 96 samples in 14 h without analyst supervision. Furthermore, its capability to detect various changes in sialylation fingerprints during cell culture manufacturing process was indispensable to ensure process robustness and consistency. Moreover, the changes in the sialylation fingerprints analyzed by this method showed strong correlations with intact mass analysis using liquid chromatography and mass spectrometry. © 2015 American Institute of Chemical Engineers Biotechnol. Prog., 32:235–241, 2016     

Heteroresistance to the model antimicrobial peptide polymyxin B in the emerging Neisseria meningitidis lineage 11.2 urethritis clade: mutations in the pilMNOPQ operon

Clusters of Neisseria meningitidis (Nm) urethritis among primarily heterosexual males in multiple US cities have been attributed to a unique non‐encapsulated meningococcal clade (the US Nm urethritis clade, US_NmUC) within the hypervirulent clonal complex 11. Resistance to antimicrobial peptides (AMPs) is a key feature of urogenital pathogenesis of the closely related species, Neisseria gonorrhoeae. The US_NmUC isolates were found to be highly resistant to the model AMP, polymyxin B (PmB, MICs 64–256 µg ml^–1). The isolates also demonstrated stable subpopulations of heteroresistant colonies that showed near total resistant to PmB (MICs 384–1024 µg ml^–1) and colistin (MIC 256 µg ml^–1) as well as enhanced LL‐37 resistance. This is the first observation of heteroresistance in N. meningitidis. Consistent with previous findings, overall PmB resistance in US_NmUC isolates was due to active Mtr efflux and LptA‐mediated lipid A modification. However, whole genome sequencing, variant analyses and directed mutagenesis revealed that the heteroresistance phenotypes and very high‐level AMP resistance were the result of point mutations and IS1655 element movement in the pilMNOPQ operon, encoding the type IV pilin biogenesis apparatus. Cross‐resistance to other classes of antibiotics was also observed in the heteroresistant colonies. High‐level resistance to AMPs may contribute to the pathogenesis of US_NmUC.     

Measuring the repertoire of age-related behavioral changes in Drosophila melanogaster

Aging affects almost all aspects of an organism—its morphology, its physiology, its behavior. Isolating which biological mechanisms are regulating these changes, however, has proven difficult, potentially due to our inability to characterize the full repertoire of an animal’s behavior across the lifespan. Using data from fruit flies (D. melanogaster) we measure the full repertoire of behaviors as a function of age. We observe a sexually dimorphic pattern of changes in the behavioral repertoire during aging. Although the stereotypy of the behaviors and the complexity of the repertoire overall remains relatively unchanged, we find evidence that the observed alterations in behavior can be explained by changing the fly’s overall energy budget, suggesting potential connections between metabolism, aging, and behavior.