共查询到20条相似文献,搜索用时 15 毫秒
1.
Qiao Wang Birong Shen Pengli Zheng Hui Feng Yige Guo Wenyuan Cao Liang Chen Xiao Liu Guodong Zhao Sizheng Xu Weide Shen Jianguo Chen Junlin Teng 《The Journal of biological chemistry》2013,288(37):26649-26657
Silkworm posterior silkgland is a model for studying intracellular trafficking. Here, using this model, we identify several potential cargo proteins of BmKinesin-1 and focus on one candidate, BmCREC. BmCREC (also known as Bombyx mori DNA supercoiling factor, BmSCF) was previously proposed to supercoil DNA in the nucleus. However, we show here that BmCREC is localized in the ER lumen. Its C-terminal tetrapeptide HDEF is recognized by the KDEL receptor, and subsequently it is retrogradely transported by coat protein I (COPI) vesicles to the ER. Lacking the HDEF tetrapeptide of BmCREC or knocking down COPI subunits results in decreased ER retention and simultaneously increased secretion of BmCREC. Furthermore, we find that BmCREC knockdown markedly disrupts the morphology of the ER and Golgi apparatus and leads to a defect of posterior silkgland tube expansion. Together, our results clarify the ER retention mechanism of BmCREC and reveal that BmCREC is indispensable for maintaining ER/Golgi morphology. 相似文献
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Sally E. Thomas Elke Malzer Adriana Ordó?ez Lucy E. Dalton Emily F. A. van ′t Wout Elizabeth Liniker Damian C. Crowther David A. Lomas Stefan J. Marciniak 《The Journal of biological chemistry》2013,288(11):7606-7617
Cell cycle checkpoints ensure that proliferation occurs only under permissive conditions, but their role in linking nutrient availability to cell division is incompletely understood. Protein folding within the endoplasmic reticulum (ER) is exquisitely sensitive to energy supply and amino acid sources because deficiencies impair luminal protein folding and consequently trigger ER stress signaling. Following ER stress, many cell types arrest within the G1 phase, although recent studies have identified a novel ER stress G2 checkpoint. Here, we report that ER stress affects cell cycle progression via two classes of signal: an early inhibition of protein synthesis leading to G2 delay involving CHK1 and a later induction of G1 arrest associated both with the induction of p53 target genes and loss of cyclin D1. We show that substitution of p53/47 for p53 impairs the ER stress G1 checkpoint, attenuates the recovery of protein translation, and impairs induction of NOXA, a mediator of cell death. We propose that cell cycle regulation in response to ER stress comprises redundant pathways invoked sequentially first to impair G2 progression prior to ultimate G1 arrest. 相似文献
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Jared R. Helm Marvin Bentley Kevin D. Thorsen Ting Wang Lauren Foltz Viola Oorschot Judith Klumperman Jesse C. Hay 《The Journal of biological chemistry》2014,289(34):23609-23628
Luminal calcium released from secretory organelles has been suggested to play a regulatory role in vesicle transport at several steps in the secretory pathway; however, its functional roles and effector pathways have not been elucidated. Here we demonstrate for the first time that specific luminal calcium depletion leads to a significant decrease in endoplasmic reticulum (ER)-to-Golgi transport rates in intact cells. Ultrastructural analysis revealed that luminal calcium depletion is accompanied by increased accumulation of intermediate compartment proteins in COPII buds and clusters of unfused COPII vesicles at ER exit sites. Furthermore, we present several lines of evidence suggesting that luminal calcium affected transport at least in part through calcium-dependent interactions between apoptosis-linked gene-2 (ALG-2) and the Sec31A proline-rich region: 1) targeted disruption of ALG-2/Sec31A interactions caused severe defects in ER-to-Golgi transport in intact cells; 2) effects of luminal calcium and ALG-2/Sec31A interactions on transport mutually required each other; and 3) Sec31A function in transport required luminal calcium. Morphological phenotypes of disrupted ALG-2/Sec31A interactions were characterized. We found that ALG-2/Sec31A interactions were not required for the localization of Sec31A to ER exit sites per se but appeared to acutely regulate the stability and trafficking of the cargo receptor p24 and the distribution of the vesicle tether protein p115. These results represent the first outline of a mechanism that connects luminal calcium to specific protein interactions regulating vesicle trafficking machinery. 相似文献
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In the yeast Saccharomyces cerevisiae two alcohol acetyltransferases (AATases), Atf1 and Atf2, condense short chain alcohols with acetyl-CoA to produce volatile acetate esters. Such esters are, in large part, responsible for the distinctive flavors and aromas of fermented beverages including beer, wine, and sake. Atf1 and Atf2 localize to the endoplasmic reticulum (ER) and Atf1 is known to localize to lipid droplets (LDs). The mechanism and function of these localizations are unknown. Here, we investigate potential mechanisms of Atf1 and Atf2 membrane association. Segments of the N- and C-terminal domains of Atf1 (residues 24–41 and 508–525, respectively) are predicted to be amphipathic helices. Truncations of these helices revealed that the terminal domains are essential for ER and LD association. Moreover, mutations of the basic or hydrophobic residues in the N-terminal helix and hydrophobic residues in the C-terminal helix disrupted ER association and subsequent sorting from the ER to LDs. Similar amphipathic helices are found at both ends of Atf2, enabling ER and LD association. As was the case with Atf1, mutations to the N- and C-terminal helices of Atf2 prevented membrane association. Sequence comparison of the AATases from Saccharomyces, non-Saccharomyces yeast (K. lactis and P. anomala) and fruits species (C. melo and S. lycopersicum) showed that only AATases from Saccharomyces evolved terminal amphipathic helices. Heterologous expression of these orthologs in S. cerevisiae revealed that the absence of terminal amphipathic helices eliminates LD association. Combined, the results of this study suggest a common mechanism of membrane association for AATases via dual N- and C-terminal amphipathic helices. 相似文献
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Masato Irisawa Jun Inoue Nozomi Ozawa Kazutoshi Mori Ryuichiro Sato 《The Journal of biological chemistry》2009,284(42):28995-29004
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Kristina Oresic Caroline L. Ng Domenico Tortorella 《The Journal of biological chemistry》2009,284(9):5905-5914
The human cytomegalovirus proteins US2 and US11 have co-opted endoplasmic
reticulum (ER) quality control to facilitate the destruction of major
histocompatibility complex class I heavy chains. The class I heavy chains are
dislocated from the ER to the cytosol, where they are deglycosylated and
subsequently degraded by the proteasome. We examined the role of TRAM1
(translocating chain-associated membrane protein-1) in the dislocation of
class I molecules using US2- and US11-expressing cells. TRAM1 is an ER protein
initially characterized for its role in processing nascent polypeptides.
Co-immunoprecipitation studies demonstrated that TRAM1 can complex with the
wild type US2 and US11 proteins as well as deglycosylated and
polyubiquitinated class I degradation intermediates. In studies using US2- and
US11-TRAM1 knockdown cells, we observed an increase in levels of class I heavy
chains. Strikingly, increased levels of glycosylated heavy chains were
observed in TRAM1 knockdown cells when compared with control cells in a
pulse-chase experiment. In fact, US11-mediated class I dislocation was more
sensitive to the lack of TRAM1 than US2. These results provide further
evidence that these viral proteins may utilize distinct complexes to
facilitate class I dislocation. For example, US11-mediated class I heavy chain
degradation requires Derlin-1 and SEL1L, whereas signal peptide peptidase is
critical for US2-induced class I destabilization. In addition, TRAM1 can
complex with the dislocation factors Derlin-1 and signal peptide peptidase.
Collectively, the data support a model in which TRAM1 functions as a cofactor
to promote efficient US2- and US11-dependent dislocation of major
histocompatibility complex class I heavy chains.HCMV2 can
down-regulate cell surface expression of the immunologically important
molecule major histocompatibility complex class I to avoid immune detection by
cytotoxic T cells (1,
2). More specifically, the HCMV
US2 and US11 gene products alone can target the ER-localized major
histocompatibility complex class I heavy chains for extraction across the ER
membrane by a process referred to as dislocation or retrograde translocation.
The N-linked glycan is then removed upon exposure to the cytosol by
N-glycanase (3),
followed by proteasomal destruction
(4,
5). The HCMV US2 and US11
proteins utilize the ER quality control process to eliminate class I heavy
cells in a similar manner as misfolded or damaged ER proteins (e.g.
genetic mutants of α1-antitrypsin
(6) and the cystic fibrosis
transmembrane conductance regulator protein
(7)) are targeted for
degradation (8). Hence,
analysis of US2- and US11-mediated destruction of class I heavy chains
provides an excellent system to delineate viral protein function as well as
the ER quality control process.ER and cytosolic proteins are required for US2- and US11-mediated
dislocation/degradation of class I heavy chains. Some of these proteins have
also been identified in the processing of aberrant ER polypeptides. The ER
chaperones calnexin, calreticulin, and BiP have been implicated in
US2-mediated class I destruction
(9) as well as in the removal
of some misfolded ER proteins
(10). The ubiquitination
machinery also participates in the extraction of class I heavy chains as
ubiquitinated heavy chains are observed prior to dislocation
(11,
12). For misfolded ER
degradation substrates, ubiquitin conjugation enzymes (e.g. Ubc6p and
Ubc7p/Cue1p) and ubiquitin ligases Hrd1p/Der3p, Doa10p, and Ubc1p have been
implicated in the dislocation reaction
(8). Interestingly, the ER
membrane protein Derlin-1 along with SEL1L are involved in US11-mediated class
I heavy chain degradation
(13-15),
whereas SPP is critical for US2-induced class I destabilization
(16). The ubiquitinated
substrates are dislocated by the AAA-ATPase complex composed of p97-Ufd1-Npl4
(17) while docked to the ER
through its interaction with VIMP
(14) followed by proteasome
destruction. The inhibition of the proteasome causes the accumulation of
deglycosylated class I heavy chain intermediate in US2 and US11 cells,
allowing the dislocation and degradation reactions to be studied as separate
processes (4,
5).Despite the identification of some cellular proteins that assist US2- and
US11-mediated class I dislocation, the dislocation pore and accessory factors
that mediate the efficient extraction of class I through the bilayer have yet
to be completely defined. The current study explores the role of TRAM1
(translocating chain-associated membrane protein-1) in US2- and US11-mediated
class I dislocation. TRAM1 is an ER-resident multispanning membrane protein
that can mediate the lateral movement of select signal peptides and
transmembrane segments from the translocon into the membrane bilayer
(18), a property that makes it
uniquely qualified to participate in the dislocation of a membrane protein.
TRAM1 has been cross-linked to signal peptides as well as transmembrane
domains of nascent polypeptides during the early stages of protein processing
(19-25).
Interestingly, unlike the Sec61 complex and the signal recognition particle
receptor, TRAM1 is not essential for the translocation of all membrane
proteins into the ER (20,
21). Hence, TRAM1 may utilize
its ability to engage hydrophobic domains to assist in the efficient
dislocation of membrane proteins. In fact, association and TRAM1 knockdown
studies demonstrate that TRAM1 participates in US2- and US11-mediated
dislocation of class I heavy chains. Collectively, our data suggest for the
first time that TRAM1 plays a role in the dislocation of a membrane
glycoprotein. 相似文献
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Fu-Chia Yang Ya-Huei Lin Wei-Hao Chen Jing-Yi Huang Hsin-Yun Chang Su-Hui Su Hsiao-Ting Wang Chun-Yi Chiang Pang-Hung Hsu Ming-Daw Tsai Bertrand Chin-Ming Tan Sheng-Chung Lee 《The Journal of biological chemistry》2013,288(47):33861-33872
Salt-inducible kinase 2 (SIK2) is an important regulator of cAMP response element-binding protein-mediated gene expression in various cell types and is the only AMP-activated protein kinase family member known to interact with the p97/valosin-containing protein (VCP) ATPase. Previously, we have demonstrated that SIK2 can regulate autophagy when proteasomal function is compromised. Here we report that physical and functional interactions between SIK2 and p97/VCP underlie the regulation of endoplasmic reticulum (ER)-associated protein degradation (ERAD). SIK2 co-localizes with p97/VCP in the ER membrane and stimulates its ATPase activity through direct phosphorylation. Although the expression of wild-type recombinant SIK2 accelerated the degradation and removal of ERAD substrates, the kinase-deficient variant conversely had no effect. Furthermore, down-regulation of endogenous SIK2 or mutation of the SIK2 target site on p97/VCP led to impaired degradation of ERAD substrates and disruption of ER homeostasis. Collectively, these findings highlight a mechanism by which the interplay between SIK2 and p97/VCP contributes to the regulation of ERAD in mammalian cells. 相似文献
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The Mammalian Protein (rbet1) Homologous to Yeast Bet1p Is Primarily Associated with the Pre-Golgi Intermediate Compartment and Is Involved in Vesicular Transport from the Endoplasmic Reticulum to the Golgi Apparatus 总被引:6,自引:1,他引:6 下载免费PDF全文
Tao Zhang Siew Heng Wong Bor Luen Tang Yue Xu Frank Peter V. Nathan Subramaniam Wanjin Hong 《The Journal of cell biology》1997,139(5):1157-1168
Yeast Bet1p participates in vesicular transport from the endoplasmic reticulum to the Golgi apparatus and functions as a soluble N-ethylmaleimide–sensitive factor attachment protein receptor (SNARE) associated with ER-derived vesicles. A mammalian protein (rbet1) homologous to Bet1p was recently identified, and it was concluded that rbet1 is associated with the Golgi apparatus based on the subcellular localization of transiently expressed epitope-tagged rbet1. In the present study using rabbit antibodies raised against the cytoplasmic domain of rbet1, we found that the majority of rbet1 is not associated with the Golgi apparatus as marked by the Golgi mannosidase II in normal rat kidney cells. Rather, rbet1 is predominantly associated with vesicular spotty structures that concentrate in the peri-Golgi region but are also present throughout the cytoplasm. These structures colocalize with the KDEL receptor and ERGIC-53, which are known to be enriched in the intermediate compartment. When the Golgi apparatus is fragmented by nocodazole treatment, a significant portion of rbet1 is not colocalized with structures marked by Golgi mannosidase II or the KDEL receptor. Association of rbet1 in cytoplasmic spotty structures is apparently not altered by preincubation of cells at 15°C. However, upon warming up from 15 to 37°C, rbet1 concentrates into the peri-Golgi region. Furthermore, rbet1 colocalizes with vesicular stomatitis virus G-protein en route from the ER to the Golgi. Antibodies against rbet1 inhibit in vitro transport of G-protein from the ER to the Golgi apparatus in a dose-dependent manner. This inhibition can be neutralized by preincubation of antibodies with recombinant rbet1. EGTA is known to inhibit ER-Golgi transport at a stage after vesicle docking but before the actual fusion event. Antibodies against rbet1 inhibit ER-Golgi transport only when they are added before the EGTA-sensitive stage. These results suggest that rbet1 may be involved in the docking process of ER- derived vesicles with the cis-Golgi membrane. 相似文献
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Hwa-Young Lee Geum-Hwa Lee Mi-Rin Lee Hye-Kyung Kim Nan-young Kim Seung-Hyun Kim Yong-Chul Lee Hyung-Ryong Kim Han-Jung Chae 《PloS one》2013,8(12)
Eucommia ulmoides Oliver is a natural product widely used as a dietary supplement and medicinal plant. Here, we examined the potential regulatory effects of Eucommia ulmoides Oliver extracts (EUE) on hepatic dyslipidemia and its related mechanisms by in vitro and in vivo studies. EUE and its two active constituents, aucubin and geniposide, inhibited palmitate-induced endoplasmic reticulum (ER) stress, reducing hepatic lipid accumulation through secretion of apolipoprotein B and associated triglycerides and cholesterol in human HepG2 hepatocytes. To determine how EUE diminishes the ER stress response, lysosomal and proteasomal protein degradation activities were analyzed. Although proteasomal activity was not affected, lysosomal enzyme activities including V-ATPase were significantly increased by EUE as well as aucubin and geniposide in HepG2 cells. Treatment with the V-ATPase inhibitor, bafilomycin, reversed the inhibition of ER stress, secretion of apolipoprotein B, and hepatic lipid accumulation induced by EUE or its component, aucubin or geniposide. In addition, EUE was determined to regulate hepatic dyslipidemia by enhancing lysosomal activity and to regulate ER stress in rats fed a high-fat diet. Together, these results suggest that EUE and its active components enhance lysosomal activity, resulting in decreased ER stress and hepatic dyslipidemia. 相似文献
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Leopoldo de Meis Luisa A. Ketzer Rodrigo Madeiro da Costa Ivone Rosa de Andrade Marlene Benchimol 《PloS one》2010,5(3)
Brown adipose tissue (BAT) mitochondria thermogenesis is regulated by uncoupling protein 1 (UCP 1), GDP and fatty acids. In this report, we observed fusion of the endoplasmic reticulum (ER) membrane with the mitochondrial outer membrane of rats BAT. Ca2+-ATPase (SERCA 1) was identified by immunoelectron microscopy in both ER and mitochondria. This finding led us to test the Ca2+ effect in BAT mitochondria thermogenesis. We found that Ca2+ increased the rate of respiration and heat production measured with a microcalorimeter both in coupled and uncoupled mitochondria, but had no effect on the rate of ATP synthesis. The Ca2+ concentration needed for half-maximal activation varied between 0.08 and 0.11 µM. The activation of respiration was less pronounced than that of heat production. Heat production and ATP synthesis were inhibited by rotenone and KCN.Liver mitochondria have no UCP1 and during respiration synthesize a large amount of ATP, produce little heat, GDP had no effect on mitochondria coupling, Ca2+ strongly inhibited ATP synthesis and had little or no effect on the small amount of heat released. These finding indicate that Ca2+ activation of thermogenesis may be a specific feature of BAT mitochondria not found in other mitochondria such as liver. 相似文献
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