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51.
Cell-to-cell transfer of glial proteins to the squid giant axon: The glia- neuron protein transfer hypothesis
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The hypothesis that glial cells synthesize proteins which are transferred to adjacent neurons was evaluated in the giant fiber of the squid (Loligo pealei). When giant fibers are separated from their neuron cell bodies and incubated in the presence of radioactive amino acids, labeled proteins appear in the glial cells and axoplasm. Labeled axonal proteins were detected by three methods: extrusion of the axoplasm from the giant fiber, autoradiography, and perfusion of the giant fiber. This protein synthesis is completely inhibited by puromycin but is not affected by chloramphenicol. The following evidence indicates that the labeled axonal proteins are not synthesized within the axon itself. (a) The axon does not contain a significant amount of ribosomes or ribosomal RNA. (b) Isolated axoplasm did not incorporate [(3)H]leucine into proteins. (c) Injection of Rnase into the giant axon did not reduce the appearance of newly synthesized proteins in the axoplasm of the giant fiber. These findings, coupled with other evidence, have led us to conclude that the adaxonal glial cells synthesize a class of proteins which are transferred to the giant axon. Analysis of the kinetics of this phenomenon indicates that some proteins are transferred to the axon within minutes of their synthesis in the glial cells. One or more of the steps in the transfer process appear to involve Ca++, since replacement of extracellular Ca++ by either Mg++ or Co++ significantly reduces the appearance of labeled proteins in the axon. A substantial fraction of newly synthesized glial proteins, possibly as much as 40 percent, are transferred to the giant axon. These proteins are heterogeneous and range in size from 12,000 to greater than 200,000 daltons. Comparisons of the amount of amino acid incorporation in glia cells and neuron cell bodies raise the possibility that the adaxonal glial cells may provide an important source of axonal proteins which is supplemental to that provided by axonal transport from the cell body. These findings are discussed with reference to a possible trophic effect of glia on neurons and metabolic cooperation between adaxonal glia and the axon. 相似文献
52.
Orlando E. Martinez Brendan J. Mahoney Andrew K. Goring Sung-Wook Yi Denise P. Tran Duilio Cascio Martin L. Phillips Musleh M. Muthana Xi Chen Michael E. Jung Joseph A. Loo Robert T. Clubb 《The Journal of biological chemistry》2022,298(2)
Wall teichoic acid (WTA) polymers are covalently affixed to the Gram-positive bacterial cell wall and have important functions in cell elongation, cell morphology, biofilm formation, and β-lactam antibiotic resistance. The first committed step in WTA biosynthesis is catalyzed by the TagA glycosyltransferase (also called TarA), a peripheral membrane protein that produces the conserved linkage unit, which joins WTA to the cell wall peptidoglycan. TagA contains a conserved GT26 core domain followed by a C-terminal polypeptide tail that is important for catalysis and membrane binding. Here, we report the crystal structure of the Thermoanaerobacter italicus TagA enzyme bound to UDP-N-acetyl-d-mannosamine, revealing the molecular basis of substrate binding. Native MS experiments support the model that only monomeric TagA is enzymatically active and that it is stabilized by membrane binding. Molecular dynamics simulations and enzyme activity measurements indicate that the C-terminal polypeptide tail facilitates catalysis by encapsulating the UDP-N-acetyl-d-mannosamine substrate, presenting three highly conserved arginine residues to the active site that are important for catalysis (R214, R221, and R224). From these data, we present a mechanistic model of catalysis that ascribes functions for these residues. This work could facilitate the development of new antimicrobial compounds that disrupt WTA biosynthesis in pathogenic bacteria. 相似文献
53.
Christopher K. Sue Nicole A. Cheung Brendan J. Mahoney Scott A. McConnell Jack M. Scully Janine Y. Fu Chungyu Chang Hung Ton-That Joseph A. Loo Robert T. Clubb 《Biopolymers》2024,115(1):e23539
Many species of pathogenic gram-positive bacteria display covalently crosslinked protein polymers (called pili or fimbriae) that mediate microbial adhesion to host tissues. These structures are assembled by pilus-specific sortase enzymes that join the pilin components together via lysine-isopeptide bonds. The archetypal SpaA pilus from Corynebacterium diphtheriae is built by the CdSrtA pilus-specific sortase, which crosslinks lysine residues within the SpaA and SpaB pilins to build the shaft and base of the pilus, respectively. Here, we show that CdSrtA crosslinks SpaB to SpaA via a K139(SpaB)-T494(SpaA) lysine-isopeptide bond. Despite sharing only limited sequence homology, an NMR structure of SpaB reveals striking similarities with the N-terminal domain of SpaA (NSpaA) that is also crosslinked by CdSrtA. In particular, both pilins contain similarly positioned reactive lysine residues and adjacent disordered AB loops that are predicted to be involved in the recently proposed “latch” mechanism of isopeptide bond formation. Competition experiments using an inactive SpaB variant and additional NMR studies suggest that SpaB terminates SpaA polymerization by outcompeting NSpaA for access to a shared thioester enzyme–substrate reaction intermediate. 相似文献
54.
Old World psittacines experienced an acute fatal illness in outdoor breeding collections in South Florida. Toxoplasma-like organisms were found histologically in pulmonary capillaries and elsewhere. Because the organisms underwent schizogony and could not be transmitted to mice, we looked for a cause other than Toxoplasma gondii. An opossum was trapped on the premises of 1 facility and was found to be shedding sporocysts similar to Sarcocystis falcatula in its feces. Cockroaches were prevalent and suspected as transport hosts. Cockroaches that had ingested opossum feces and subsequently were fed to cockatoos induced an identical fatal illness. Obstruction of pulmonary capillaries by developing schizonts and pulmonary edema were the most important pathologic findings. The epidemic was stopped by biological insect control employing flightless chickens to reduce cockroach populations and by an electric fence restricting access of opossums to these outdoor psittacine breeding facilities. 相似文献
55.
56.
Calcitonin gene-related peptide is not essential for the development of pressure overload-induced hypertrophy in vivo 总被引:2,自引:0,他引:2
Young Martin E. Okerberg Kristen A. Wilson Christopher R. Deferrari Dario A. Ying Jun Guthrie Patrick Razeghi Peter Clubb Fred J. Jr. Taegtmeyer Heinrich 《Molecular and cellular biochemistry》2001,225(1-2):43-49
The regulatory neuropeptide calcitonin-gene related peptide (CGRP) has been shown to evoke a hypertrophic response in isolated cardiomyocytes in vitro, an effect which was attributed to PKC activation. Activation of PKC has previously been implicated in the development of cardiac hypertrophy. We therefore investigated the role of CGRP in pressure overload-induced hypertrophy in vivo, which has not previously been reported. Constriction of the ascending aorta of rats resulted in an increase in the heart weight to body weight ratio, increased myocyte diameter, re-expression of the fetal genes ANF, MHC and skeletal -actin, and decreased expression of the adult genes GLUT4 and SERCA2a. Treatment of neonatal rat pups (1–2 days old) with capsaicin (50 mg/kg), resulted in the permanent de-afferentation of small-diameter unmyelinated CGRP-containing sensory C-fibres. Such treatment caused a 68% decrease in the CGRP-like immunoreactivity of hearts isolated from 10 week old rats (p < 0.001). Contrary to expectations, aortic constriction of capsaicin treated rats had no effect on the development of hypertrophy at the trophic, morphometric or gene expression levels. The results suggest that the development of pressure overload-induced hypertrophy in vivo does not require the regulatory neuropeptide CGRP. 相似文献
57.
Clubb BH Chou YH Herrmann H Svitkina TM Borisy GG Goldman RD 《Biochemical and biophysical research communications》2000,273(1):183-187
Plectin is a high-molecular-weight cytoskeleton-associated protein that was initially identified in intermediate filament (IF)-enriched fractions of rat C6 glioma cells. At the cellular level, plectin has been found to associate with IF networks and IF-associated structures that are involved in cell-cell and cell-substrate adhesions. IFAP300 is an IF-associated protein that was initially identified in hamster cells by a monoclonal antibody directed against a high molecular weight protein present in IF-enriched cytoskeletal preparations. Plectin and IFAP300 display similar distribution patterns within cells as determined by immunofluorescence. Based upon this and the finding that their biochemical properties are similar, it has been suggested that they may actually be orthologous proteins. In this paper we demonstrate that this is the case. Cloning and sequencing of most of the hamster plectin cDNA demonstrates that plectin is found in hamster cells and that its sequence is highly conserved between species. Using immunological cross-reactivity, epitope mapping, and immunoelectron microscopy, we show that IFAP300 is actually the hamster ortholog of plectin. 相似文献
58.
59.
Renal failure was diagnosed in an 11-mo-old male domestic shorthair cat from a colony with mucopolysaccharidosis type I lysosomal storage disease. Grossly, the kidneys were enlarged and bulged on cut section. Histology revealed tubular necrosis and regeneration with severe interstitial macrophage accumulation. Tubular epithelial cells and interstitial macrophages were distended by abundant, large cytoplasmic vacuoles. Electron microscopy demonstrated severe tubular epithelial vacuolar degeneration with lysosomes distended by granular debris and mineral precipitates. Interstitial macrophages contained similarly distended lysosomes. Although the initial cause of the tubular injury was not identified, the presence of macrophages laden with storage product most likely exacerbated the disease. The macrophage infiltrate may have caused tubular ischemia by compressing peritubular capillaries and separating tubules from their blood supply. Because the kidney is not normally affected in MPS I, this case is an unusual presentation of a well-characterized disease. Furthermore, this report documents the diagnostic workflow used to investigate a single case of feline acute renal failure in the setting of numerous at-risk laboratory animals. 相似文献
60.
Duan L Raja SM Chen G Virmani S Williams SH Clubb RJ Mukhopadhyay C Rainey MA Ying G Dimri M Chen J Reddi AL Naramura M Band V Band H 《The Journal of biological chemistry》2011,286(1):620-633
The E3 ubiquitin ligase Casitas B lymphoma protein (Cbl) controls the ubiquitin-dependent degradation of EGF receptor (EGFR), but its role in regulating downstream signaling elements with which it associates and its impact on biological outcomes of EGFR signaling are less clear. Here, we demonstrate that stimulation of EGFR on human mammary epithelial cells disrupts adherens junctions (AJs) through Vav2 and Rac1/Cdc42 activation. In EGF-stimulated cells, Cbl regulates the levels of phosphorylated Vav2 thereby attenuating Rac1/Cdc42 activity. Knockdown of Cbl and Cbl-b enhanced the EGF-induced disruption of AJs and cell motility. Overexpression of constitutively active Vav2 activated Rac1/Cdc42 and reorganized junctional actin cytoskeleton; these effects were suppressed by WT Cbl and enhanced by a ubiquitin ligase-deficient Cbl mutant. Cbl forms a complex with phospho-EGFR and phospho-Vav2 and facilitates phospho-Vav2 ubiquitinylation. Cbl can also interact with Vav2 directly in a Cbl Tyr-700-dependent manner. A ubiquitin ligase-deficient Cbl mutant enhanced the morphological transformation of mammary epithelial cells induced by constitutively active Vav2; this effect requires an intact Cbl Tyr-700. These results indicate that Cbl ubiquitin ligase plays a critical role in the maintenance of AJs and suppression of cell migration through down-regulation of EGFR-Vav2 signaling. 相似文献