Geographic divergence of “Sulfolobus islandicus” strains assessed by genomic analyses including electronic DNA hybridization confirms they are geovars

Ten well-annotated genomes of “Sulfolobus islandicus” strains from different geographic locations have been released at the NCBI database. Whole genome based composition vector trees indicate that these strains show the same branching patterns as originally reported by multi-locus sequence analysis. To determine whether the ten strains meet the criteria for separate species, DNA–DNA hybridization (DDH) was performed in silico. DDH values of strains from the same geographic location, i.e., Iceland, Kamchatka and North America, ranged from 82.4 to 95.4 %, clearly qualifying them as members of the same species. The lowest DDH values found between locations ranged from 75.5 to 76.6 %, which exceed the 70 % DDH threshold for a species thereby indicating they are all members of the same species based on the currently accepted definition. The clear divergences of strains from the different geographic locations are sufficiently great to consider them as separate geovars. “S. islandicus” has not yet been validly named and a type strain has not been deposited in culture collections. We urgently recommend that those who study the organism fulfill the criteria of the International Code of Nomenclature of Bacteria in order to designate a type strain and to identify and deposit related strains of this species to make them available to the broader scientific community.     

RhoA/rho kinase signaling reduces connexin43 expression in high glucose-treated glomerular mesangial cells with zonula occludens-1 involvement

RhoA/Rho kinase (ROCK) signaling has been suggested to be involved in diabetic nephropathy (DN) pathogenesis. Altered expression of connexin43 (Cx43) has been found in kidneys of diabetic animals. Both of them have been found to regulate nuclear factor kappa-B (NF-κB) activation in high glucose-treated glomerular mesangial cells (GMCs). The aim of this study was to investigate the relationship between RhoA/ROCK signaling and Cx43 in the DN pathogenesis. We found that upregulation of Cx43 expression inhibited NF-κB p65 nuclear translocation induced by RhoA/ROCK signaling in GMCs. Inhibition of RhoA/ROCK signaling attenuated the high glucose-induced decrease in Cx43. F-actin accumulation and an enhanced interaction between zonula occludens-1 (ZO-1) and Cx43 were observed in high glucose-treated GMCs. ZO-1 depletion or disruption of F-actin formation also inhibited the reduction in Cx43 protein levels induced by high glucose. In conclusion, activated RhoA/ROCK signaling induces Cx43 degradation in GMCs cultured in high glucose, depending on F-actin regulation. Increased F-actin induced by RhoA/ROCK signaling promotes the association between ZO-1 and Cx43, which possibly triggered Cx43 endocytosis, a mechanism of NF-κB activation in high glucose-treated GMCs.     

Mass spectrometry for the biophysical characterization of therapeutic monoclonal antibodies

Monoclonal antibodies (mAbs) are powerful therapeutics, and their characterization has drawn considerable attention and urgency. Unlike small-molecule drugs (150–600 Da) that have rigid structures, mAbs (∼150 kDa) are engineered proteins that undergo complicated folding and can exist in a number of low-energy structures, posing a challenge for traditional methods in structural biology. Mass spectrometry (MS)-based biophysical characterization approaches can provide structural information, bringing high sensitivity, fast turnaround, and small sample consumption. This review outlines various MS-based strategies for protein biophysical characterization and then reviews how these strategies provide structural information of mAbs at the protein level (intact or top-down approaches), peptide, and residue level (bottom-up approaches), affording information on higher order structure, aggregation, and the nature of antibody complexes.     

Expression of NYV1 encoding the negative regulator of Pmc1 is repressed by two transcriptional repressors,Nrg1 and Mig1

ESCRT components function to form multivesicular bodies for sorting of proteins destined to the yeast vacuole. The calcium hypersensitivity of ESCRT mutants is mainly due to repressed expression of PMR1 through the Rim101/Nrg1 pathway in budding yeast. Here, we show that overexpression of PMC1 and its negative regulator gene NYV1 suppresses and increases calcium hypersensitivity of ESCRT mutants, respectively. Consistently, deletion of NYV1 suppresses their calcium hypersensitivity. Expression of NYV1 is dramatically reduced in ESCRT mutants. Promoter analysis demonstrates that both Nrg1 and Mig1 repress NYV1 expression. Deletion of ESCRTs increases Nrg1 binding, but not Mig1-binding, to the NYV1 promoter. Deletion of MIG1 increases calcium sensitivity of ESCRT mutants due to derepression of NYV1 expression.     

Hepatitis C virus core protein regulates NANOG expression via the stat3 pathway

HCV Core plays a role in the development of hepatocellular carcinoma. Aberrant expression of NANOG has been observed in many types of human malignancies. However, relationship between Core and NANOG has not been clarified. In this study, we found that Core is capable of up-regulating NANOG expression. Core-induced NANOG expression was accompanied by enforced expression of phosphorylated stat3 protein and was attenuated by inhibition of stat3 phosphorylation. ChIP showed that phosphorylated stat3 directly binds to the NANOG promoter. Core-induced NANOG expression resulted in enhanced cell growth and cell cycle progression. Knockdown of NANOG blocked the cell cycle at the G0/G1 phases and inhibited the cyclin D1 expression. Our findings provide a new insight into the mechanism of hepatocarcinogenesis by HCV infection.     

Rock2 promotes the invasion and metastasis of hepatocellular carcinoma by modifying MMP2 ubiquitination and degradation

Rho-associated coiled-coil-containing protein kinase 2 (Rock2) is a downstream effector of Rho that plays an important role in the tumorigenesis and progression of hepatocellular carcinoma (HCC). Matrix metalloproteinase 2 (MMP2) is a master regulator of tumor metastasis. In this study, we investigated the collections of Rock2 and MMP2 in HCCs and determined the potential role and molecular mechanism of Rock2 in MMP2-mediated invasiveness and metastasis. We found that Rock2 and MMP2 were markedly overexpressed in HCCs compared with the corresponding adjacent tissues, where a positive correlation in their expression was found. The knockdown of Rock2 significantly decreased MMP2 expression and inhibited the invasion and metastasis of HCC in vitro and in vivo. Additionally, the upregulation of MMP2 rescued the decreased migration and invasion induced by the knockdown of Rock2, whereas the knockdown of MMP2 decreased Rock2-enhanced HCC migration and invasion. Mechanistically, Rock2 stabilized MMP2 by preventing its ubiquitination and degradation. Together, our results link two drivers of invasion and metastasis in HCC and identify a novel pathway for MMP2 control.     

Modulation of cytokine expression in human macrophages by endocrine-disrupting chemical Bisphenol-A

Exposure to environmental endocrine-disrupting chemical Bisphenol-A (BPA) is often associated with dysregulated immune homeostasis, but the mechanisms remain unclear. In the present study, the effects of BPA on the cytokines responses of human macrophages were investigated. Treatment with BPA increased pro-inflammation cytokines tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) production, but decreased anti-inflammation cytokines interleukin-10 (IL-10) and transforming growth factor-β (TGF-β) production in THP1 macrophages, as well as in primary human macrophages. BPA effected cytokines expression through estrogen receptor α/β (ERα/β)-dependent mechanism with the evidence of ERα/β antagonist reversed the expression of cytokines. We also identified that activation of extracellular regulated protein kinases (ERK)/nuclear factor κB (NF-κB) signal cascade marked the effects of BPA on cytokines expression. Our results indicated that BPA effected inflammatory responses of macrophages via modulating of cytokines expression, and provided a new insight into the link between exposure to BPA and human health.     

Aluminum tolerance in Arabidopsis thaliana as affected by endogenous salicylic acid

Endogenous salicylic acid (SA) functions in plant response to an aluminum stress were assessed. We used different Arabidopsis thaliana genotypes including snc1 with a constitutively high content of SA, sid2 and nahG (transgenic lines) both with a low content of SA, SA insensitive mutant npr1-1, and snc1/nahG (i.e., the nahG expression in the snc1 background) with a similar SA content as in wild type (WT) plants. Results show that the snc1 plants displayed obvious growth retardation of roots and shoots under the Al³⁺ stress, whereas the sid2, nahG, and npr1-1 plants exhibited alleviated symptoms in comparison with the WT plants. The Al³⁺ content increased in all the tested genotypes with the increasing AlCl₃ concentration applied, but no significant variations were detected among the tested genotypes. The snc1 had much higher superoxide dismutase and peroxidase activities, and a lower catalase activity and the ratio of reduced to oxidized glutathione accompanied by higher accumulations of H₂O₂ and malondialdehyde compared with the WT plants. These changes were largely reversed by the introduction of nahG; the sid2, nahG, and npr1-1 plants were less affected than WT plants in all the above-mentioned parameters. The Al³⁺ stress significantly enhanced malate exudation in all the tested genotypes, but no significant correlation was observed between the SA-involved response to the Al³⁺ stress and the malate exudation. Based on these data, it was concluded that the SA-related functions in Arabidopsis response to the Al³⁺ stress were associated with the control of oxidative stress, but not of malate exudation.     

High-Density 3D Single Molecular Analysis Based on Compressed Sensing

Single molecule fitting-based superresolution microscopy achieves sub-diffraction-limit image resolution but suffers from a need for long acquisition times to gather enough molecules. Several methods have recently been developed that analyze high molecule density images but most are only applicable to two dimensions. In this study, we implemented a high-density superresolution localization algorithm based on compressed sensing and a biplane approach that provides three-dimensional information about molecules, achieving super-resolution imaging at higher molecule densities than those achieved using the conventional single molecule fitting method.