Characterization of energetically functional inverted membrane vesicles from Corynebacterium glutamicum

AbstractWe show that inverted membrane vesicles from Corynebacteriwn glutamicum, a Gram-positive bacterium, are able to generate and maintain an electrochemical gradient of protons in response to the addition of NADH. This result indicates that the respiratory chain is intact and that the vesicles are reasonably impermeable to protons. These membrane vesicles may be the starting point for in vitro translocation studies of proteins in Gram-positive bacteria

Akt promotes Endocardial-Mesenchyme Transition

Endothelial to mesenchyme transition (EndMT) can be observed during the formation of endocardial cushions from the endocardium, the endothelial lining of the atrioventricular canal (AVC), of the developing heart at embryonic day 9.5 (E9.5). Many regulators of the process have been identified; however, the mechanisms driving the initial commitment decision of endothelial cells to EndMT have been difficult to separate from processes required for mesenchymal proliferation and migration. We have several lines of evidence that suggest a central role for Akt signaling in committing endothelial cells to enter EndMT. Akt1 mRNA was restricted to the endocardium of endocardial cushions while they were forming. The PI3K/Akt signaling pathway is necessary for mesenchyme outgrowth, as sprouting was inhibited in AVC explant cultures treated with the PI3K inhibitor LY294002. Furthermore, endothelial marker, VE-cadherin, was downregulated and mesenchyme markers, N-cadherin and Snail, were induced in response to expression of a constitutively active form of Akt1 (myrAkt1) in endothelial cells. Finally, we isolated the function of Akt1 signaling in the commitment to the transition using a transgenic model where myrAkt1 was pulsed only in endocardial cells and turned off after EndMT initiation. In this way, we determined that increased Akt signaling in the endocardium drives EndMT and discounted its other functions in cushion mesenchymal cells

Developmentally Regulated Post-translational Modification of Nucleoplasmin Controls Histone Sequestration and Deposition

SummaryNucleoplasmin (Npm) is an abundant histone chaperone in vertebrate oocytes and embryos. During embryogenesis, regulation of Npm histone binding is critical for its function in storing and releasing maternal histones to establish and maintain the zygotic epigenome. Here, we demonstrate that Xenopus laevis Npm post-translational modifications (PTMs) specific to the oocyte and egg promote either histone deposition or sequestration, respectively. Mass spectrometry and Npm phosphomimetic mutations used in chromatin assembly assays identified hyperphosphorylation on the N-terminal tail as a critical regulator for sequestration. C-terminal tail phosphorylation and PRMT5-catalyzed arginine methylation enhance nucleosome assembly by promoting histone interaction with the second acidic tract of Npm. Electron microscopy reconstructions of Npm and TTLL4 activity toward the C-terminal tail demonstrate that oocyte- and egg-specific PTMs cause Npm conformational changes. Our results reveal that PTMs regulate Npm chaperoning activity by modulating Npm conformation and Npm-histone interaction, leading to histone sequestration in the egg

Dilepton production in proton-proton collisions at BEVALAC energies

The dilepton production in elementary ${pp\to e^{+}e^{-}X}$ reactions at BEVALAC energies $T_{lab}=1\div 5$ GeV is investigated. The calculations include direct ${e^{+}e^{-}}$ decays of the vector mesons $\rho ^{0}$, $\omega$, and $\phi$, Dalitz decays of the $\pi ^{0}$-, $\eta$-, $$-, $\omega$-, and $\phi$-mesons, and of the baryon resonances $$ $...$ . The subthreshold vector meson production cross sections in $pp$ collisions are treated in a way sufficient to avoid double counting with the inclusive vector meson production. The vector meson dominance model for the transition form factors of the resonance Dalitz decays $R\to e^{+}e^{-}N$ is used in an extended form to ensure correct asymptotics which are in agreement with the quark counting rules. Such a modification gives an unified and consistent description of both $R\to N\gamma$ radiative decays and $R\to N\rho (\omega)$ meson decays. The effect of multiple pion production on the experimental efficiency for the detection of the dilepton pairs is studied. We find the dilepton yield in reasonable agreement with the experimental data for the set of intermediate energies whereas at the highest energy $T_{lab}=4.88$ GeV the number of dilepton pairs is likely to be overestimated experimentally in the mass range $M=300\div 700$ MeV.Comment: 25 pages (IOP style), 5 figures, revised manuscript accepted for publication in JP

Developmentally Regulated Post-translational Modification of Nucleoplasmin Controls Histone Sequestration and Deposition

SummaryNucleoplasmin (Npm) is an abundant histone chaperone in vertebrate oocytes and embryos. During embryogenesis, regulation of Npm histone binding is critical for its function in storing and releasing maternal histones to establish and maintain the zygotic epigenome. Here, we demonstrate that Xenopus laevis Npm post-translational modifications (PTMs) specific to the oocyte and egg promote either histone deposition or sequestration, respectively. Mass spectrometry and Npm phosphomimetic mutations used in chromatin assembly assays identified hyperphosphorylation on the N-terminal tail as a critical regulator for sequestration. C-terminal tail phosphorylation and PRMT5-catalyzed arginine methylation enhance nucleosome assembly by promoting histone interaction with the second acidic tract of Npm. Electron microscopy reconstructions of Npm and TTLL4 activity toward the C-terminal tail demonstrate that oocyte- and egg-specific PTMs cause Npm conformational changes. Our results reveal that PTMs regulate Npm chaperoning activity by modulating Npm conformation and Npm-histone interaction, leading to histone sequestration in the egg

Microparticles from tumors exposed to radiation promote immune evasion in part by PD-L1

Radiotherapy induces immune-related responses in cancer patients by various mechanisms. Here, we investigate the immunomodulatory role of tumor-derived microparticles (TMPs)-extracellular vesicles shed from tumor cells-following radiotherapy. We demonstrate that breast carcinoma cells exposed to radiation shed TMPs containing elevated levels of immune-modulating proteins, one of which is programmed death-ligand 1 (PD-L1). These TMPs inhibit cytotoxic T lymphocyte (CTL) activity both in vitro and in vivo, and thus promote tumor growth. Evidently, adoptive transfer of CTLs pre-cultured with TMPs from irradiated breast carcinoma cells increases tumor growth rates in mice recipients in comparison with control mice receiving CTLs pre-cultured with TMPs from untreated tumor cells. In addition, blocking the PD-1-PD-L1 axis, either genetically or pharmacologically, partially alleviates TMP-mediated inhibition of CTL activity, suggesting that the immunomodulatory effects of TMPs in response to radiotherapy is mediated, in part, by PD-L1. Overall, our findings provide mechanistic insights into the tumor immune surveillance state in response to radiotherapy and suggest a therapeutic synergy between radiotherapy and immune checkpoint inhibitors

Statin Efficacy and Safety for Lipid Modification in Apparently Healthy Male Military Aircrew

Introduction: Military aircrew men represent an elite group of relatively young, fit, and healthy people. The effectiveness of statin treatment in reducing low-density lipoprotein cholesterol (LDL-C) according to the current National Cholesterol Education Program (NCEP) guidelines, its safety, and compliance in this group of people has not yet been determined. Methods: We prospectively evaluated 84 military aircrew men (mean age 43 Ϯ 7 yr) with LDL-C above the current NCEP guidelines. The patients were divided into two groups according to their coronary risk factors: Group 1, LDL-C goal Ͻ 160 mg ⅐ dL Ϫ1 ; Group 2, LDL-C goal Ͻ 130 mg ⅐ dL Ϫ1 . All patients received statins in addition to therapeutic lifestyle changes and were followed for a mean of 3 Ϯ 1 yr according to a simple flow chart. Lipoprotein levels, liver function tests, creatinine phosphokinase, and subjective adverse reactions were checked periodically. Results: LDL-C significantly declined by 32% (p Ͻ 0.0001) within the first month of treatment and 99% of subjects achieved their LDL-C goal within 114 Ϯ 35 d from statin therapy initiation. The Framingham estimated 10-yr coronary risk showed a reduction at an average of 12 mo after statin therapy initiation from a baseline value of 6.54% to 3.95% (p ϭ 0.003). No subjects were grounded or disqualified from duty, there were no cardiovascular events during follow-up, and compliance to therapy was high [82/84 (98%)]. Discussion: Statin treatment in this highly select, relatively young group of aircrew men significantly and safely lowered LDL-C cholesterol levels

RhoB controls coordination of adult angiogenesis and lymphangiogenesis following injury by regulating VEZF1-mediated transcription

Mechanisms governing the distinct temporal dynamics that characterize post-natal angiogenesis and lymphangiogenesis elicited by cutaneous wounds and inflammation remain unclear. RhoB, a stress-induced small GTPase, modulates cellular responses to growth factors, genotoxic stress and neoplastic transformation. Here we show, using RhoB null mice, that loss of RhoB decreases pathological angiogenesis in the ischaemic retina and reduces angiogenesis in response to cutaneous wounding, but enhances lymphangiogenesis following both dermal wounding and inflammatory challenge. We link these unique and opposing roles of RhoB in blood versus lymphatic vasculatures to the RhoB-mediated differential regulation of sprouting and proliferation in primary human blood versus lymphatic endothelial cells. We demonstrate that nuclear RhoB-GTP controls expression of distinct gene sets in each endothelial lineage by regulating VEZF1-mediated transcription. Finally, we identify a small-molecule inhibitor of VEZF1–DNA interaction that recapitulates RhoB loss in ischaemic retinopathy. Our findings establish the first intra-endothelial molecular pathway governing the phased response of angiogenesis and lymphangiogenesis following injury

Novel role for the innate immune receptor toll-like receptor 4 (TLR4) in the regulation of the wnt signaling pathway and photoreceptor apoptosis

Recent evidence has implicated innate immunity in regulating neuronal survival in the brain during stroke and other neurodegenerations. Photoreceptors are specialized light-detecting neurons in the retina that are essential for vision. In this study, we investigated the role of the innate immunity receptor TLR4 in photoreceptors. TLR4 activation by lipopolysaccharide (LPS) significantly reduced the survival of cultured mouse photoreceptors exposed to oxidative stress. With respect to mechanism, TLR4 suppressed Wnt signaling, decreased phosphorylation and activation of the Wnt receptor LRP6, and blocked the protective effect of the Wnt3a ligand. Paradoxically, TLR4 activation prior to oxidative injury protected photoreceptors, in a phenomenon known as preconditioning. Expression of TNFα and its receptors TNFR1 and TNFR2 decreased during preconditioning, and preconditioning was mimicked by TNFα antagonists, but was independent of Wnt signaling. Therefore, TLR4 is a novel regulator of photoreceptor survival that acts through the Wnt and TNFα pathways. © 2012 Yi et al