Escherichia coli TatA and TatB Proteins Have N-out, C-in Topology in Intact Cells

The twin arginine protein transport (Tat) system translocates folded proteins across the cytoplasmic membrane of prokaryotes and the thylakoid membrane of chloroplasts. In Escherichia coli, TatA, TatB, and TatC are essential components of the machinery. A complex of TatB and TatC acts as the substrate receptor, whereas TatA is proposed to form the Tat transport channel. TatA and TatB are related proteins that comprise an N-terminal transmembrane helix and an adjacent amphipathic helix. Previous studies addressing the topological organization of TatA have given conflicting results. In this study, we have addressed the topological arrangement of TatA and TatB in intact cells by labeling of engineered cysteine residues with the membrane-impermeable thiol reagent methoxypolyethylene glycol maleimide. Our results show that TatA and TatB share an N-out, C-in topology, with no evidence that the amphipathic helices of either protein are exposed at the periplasmic side of the membrane. We further show that the N-out, C-in topology of TatA is fixed and is not affected by the absence of other Tat components or by the overproduction of a Tat substrate. These data indicate that topological reorganization of TatA is unlikely to accompany Tat-dependent protein transport

Influences of shot-peening on fine grain steels

Recently, fine grain steel has been actively researched. However, as we know, the influence concerning the shot peening processing to fine grain steels has been few reported up to now. Therefore residual stress distribution near the surface of fine grain steel after shot-peening was measured by X-ray diffraction technology. Moreover, the hardening effect the distributed near the shot-peened surface was also estimated. The relationships between the effects of shot-peening and the grain size of material were discussed. Copyright © 2007 by The International Society of Off shore and Polar Engineers(ISOPE)

Concerted loop motion triggers induced fit of FepA to ferric enterobactin

Spectroscopic analyses of fluorophore-labeled Escherichia coli FepA described dynamic actions of its surface loops during binding and transport of ferric enterobactin (FeEnt). When FeEnt bound to fluoresceinated FepA, in living cells or outer membrane fragments, quenching of fluorophore emissions reflected conformational motion of the external vestibular loops. We reacted Cys sulfhydryls in seven surface loops (L2, L3, L4, L5, L7 L8, and L11) with fluorophore maleimides. The target residues had different accessibilities, and the labeled loops themselves showed variable extents of quenching and rates of motion during ligand binding. The vestibular loops closed around FeEnt in about a second, in the order L3 > L11 > L7 > L2 > L5 > L8 > L4. This sequence suggested that the loops bind the metal complex like the fingers of two hands closing on an object, by individually adsorbing to the iron chelate. Fluorescence from L3 followed a biphasic exponential decay as FeEnt bound, but fluorescence from all the other loops followed single exponential decay processes. After binding, the restoration of fluorescence intensity (from any of the labeled loops) mirrored cellular uptake that depleted FeEnt from solution. Fluorescence microscopic images also showed FeEnt transport, and demonstrated that ferric siderophore uptake uniformly occurs throughout outer membrane, including at the poles of the cells, despite the fact that TonB, its inner membrane transport partner, was not detectable at the poles

Gene expression and matrix turnover in overused and damaged tendons

Chronic, painful conditions affecting tendons, frequently known as tendinopathy, are very common types of sporting injury. The tendon extracellular matrix is substantially altered in tendinopathy, and these changes are thought to precede and underlie the clinical condition. The tendon cell response to repeated minor injuries or “overuse” is thought to be a major factor in the development of tendinopathy. Changes in matrix turnover may also be effected by the cellular response to physical load, altering the balance of matrix turnover and changing the structure and composition of the tendon. Matrix turnover is relatively high in tendons exposed to high mechanical demands, such as the supraspinatus and Achilles, and this is thought to represent either a repair or tissue maintenance function. Metalloproteinases are a large family of enzymes capable of degrading all of the tendon matrix components, and these are thought to play a major role in the degradation of matrix during development, adaptation and repair. It is proposed that some metalloproteinase enzymes are required for the health of the tendon, and others may be damaging, leading to degeneration of the tissue. Further research is required to investigate how these enzyme activities are regulated in tendon and altered in tendinopathy. A profile of all the metalloproteinases expressed and active in healthy and degenerate tendon is required and may lead to the development of new drug therapies for these common and debilitating sports injuries

Spin dynamics in semiconductors

This article reviews the current status of spin dynamics in semiconductors which has achieved a lot of progress in the past years due to the fast growing field of semiconductor spintronics. The primary focus is the theoretical and experimental developments of spin relaxation and dephasing in both spin precession in time domain and spin diffusion and transport in spacial domain. A fully microscopic many-body investigation on spin dynamics based on the kinetic spin Bloch equation approach is reviewed comprehensively.Comment: a review article with 193 pages and 1103 references. To be published in Physics Reports

Micro-patterned culture of iPSC-derived alveolar and airway cells distinguishes SARS-CoV-2 variants.

iPS細胞から作った肺胞や気道の細胞によりSARS-CoV-2変異株の病原性を比較評価する. 京都大学プレスリリース. 2024-03-29.Micro-patterning a new system to induce alveolar and airway epithelial cells. 京都大学プレスリリース. 2024-03-29.The emergence of severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) variants necessitated a rapid evaluation system for their pathogenesis. Lung epithelial cells are their entry points; however, in addition to their limited source, the culture of human alveolar epithelial cells is especially complicated. Induced pluripotent stem cells (iPSCs) are an alternative source of human primary stem cells. Here, we report a model for distinguishing SARS-CoV-2 variants at high resolution, using separately induced iPSC-derived alveolar and airway cells in micro-patterned culture plates. The position-specific signals induced the apical-out alveolar type 2 and multiciliated airway cells at the periphery and center of the colonies, respectively. The infection studies in each lineage enabled profiling of the pathogenesis of SARS-CoV-2 variants: infection efficiency, tropism to alveolar and airway lineages, and their responses. These results indicate that this culture system is suitable for predicting the pathogenesis of emergent SARS-CoV-2 variants

KF-1 Ubiquitin Ligase: An Anxiety Suppressor

Anxiety is an instinct that may have developed to promote adaptive survival by evading unnecessary danger. However, excessive anxiety is disruptive and can be a basic disorder of other psychiatric diseases such as depression. The KF-1, a ubiquitin ligase located on the endoplasmic reticulum (ER), may prevent excessive anxiety; kf-1−/− mice exhibit selectively elevated anxiety-like behavior against light or heights. It is surmised that KF-1 degrades some target proteins, responsible for promoting anxiety, through the ER-associated degradation pathway, similar to Parkin in Parkinson's disease (PD). Parkin, another ER-ubiquitin ligase, prevents the degeneration of dopaminergic neurons by degrading the target proteins responsible for PD. Molecular phylogenetic studies have revealed that the prototype of kf-1 appeared in the very early phase of animal evolution but was lost, unlike parkin, in the lineage leading up to Drosophila. Therefore, kf-1−/− mice may be a powerful tool for elucidating the molecular mechanisms involved in emotional regulation, and for screening novel anxiolytic/antidepressant compounds

Regulation of integrin-mediated cellular responses through assembly of a CAS/Crk scaffold

AbstractThe molecular coupling of CAS and Crk in response to integrin activation is an evolutionary conserved signaling module that controls cell proliferation, survival and migration. However, when deregulated, CAS/Crk signaling also contributes to cancer progression and developmental defects in humans. Here we highlight recent advances in our understanding of how CAS/Crk complexes assemble in cells to modulate the actin cytoskeleton, and the molecular mechanisms that regulate this process. We discuss in detail the spatiotemporal dynamics of CAS/Crk assembly and how this scaffold recruits specific effector proteins that couple integrin signaling networks to the migration machinery of cells. We also highlight the importance of CAS/Crk signaling in the dual regulation of cell migration and survival mechanisms that operate in invasive cells during development and pathological conditions associated with cancer metastasis

カンケイ ドウブツ アオゴカイ Perinereis aibuhitensis キョダイ ヘモグロビン ノ グロビンサ ニ フクマレル ケウナ SSケツゴウ

The extracellular hemoglobin (Hb) from the polychaete Perinereis aibuhitensis consists of four types of 144 globins and two types of 36 linker chains，having a molecular mass of about 3,500 kDa. There are two types of globin subunits: monomer chain a and disulfide-bondedt rimer AbB. The amino acid sequences of the four globin chains (a，A ，b ，B ) were already reported，previously (Yamanaka，M. et al. Natural Science Research of Tokushima University, 19, 63-92, 2005). The site of disulfide bonds in the globin subunits have been investigated. Each globin chain contains an intrachain disulfide bond between N-terminal and C-terminal Cys residues. In addition，the interchain disulfide bonds were found between chains A and b，and b and B. Therefore，it is elucidated that the chain b is situated at the center of disulfide-bonded trimer，such as A-b-B.The sites of disulfide-bonds determined all could be suitably fitted to the tree dimensional structure of each subunit in a model without stretching or twisting. It was also confirmed that there is no free Cys residue in Peinereis globins. The positions of Cys residues of Perinereis globin sequences were compared wIth those of other 27 chains derived from the homologous Hbs. Among 31 sequences，Cys residues were distributed in six sites. The sites 1 and 2 are located at the N-terminal region of amino acid sequences，the sItes 3 and 4 at the central region，and the sites 5 and 6 at C-terminal region. Furthermore，the Cys distribution was categorized into eight patterns. Perinereis Hb has four patterns I，IV，VII，VIII，be ing lack of the central sites 3 and 4. It should be noted that the pattern II includes the unique globins from Hbs of Lamellibrachia，Riftia and Oligobrachia that carry H2S to the symbiotic bacteria，suggesting that these globin chains might carry H2S in vivo. The phylogenetic tree of 31 globin chains derived from the giant Hbs is divided into two families A and B， as already poited out by us Previously. The family A indudes pattern I-V，whereas the family B includes V- VIII