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1.
《Autophagy》2013,9(10):1639-1641
The role of membrane remodeling and phosphoinositide-binding proteins in autophagy remains elusive. PX domain proteins bind phosphoinositides and participate in membrane remodeling and trafficking events and we therefore hypothesized that one or several PX domain proteins are involved in autophagy. Indeed, the PX-BAR protein SNX18 was identified as a positive regulator of autophagosome formation using an image-based siRNA screen. We show that SNX18 interacts with ATG16L1 and LC3, and functions downstream of ATG14 and the class III PtdIns3K complex in autophagosome formation. SNX18 facilitates recruitment of ATG16L1 to perinuclear recycling endosomes, and its overexpression leads to tubulation of ATG16L1- and LC3-positive membranes. We propose that SNX18 promotes LC3 lipidation and tubulation of recycling endosomes to provide membrane for phagophore expansion. 相似文献
2.
The de novo generation of double-membrane autophagosomes is the hallmark of autophagy. The initial membranous precursor cisterna, the phagophore, is very likely generated by the fusion of vesicles and acts as a membrane seed for the subsequent expansion into an autophagosome. This latter step requires a massive convoy of lipids into the phagophore. In this review, we present recent advances in our understanding of the intracellular membrane sources and lipid delivery mechanisms, which principally rely on vesicular transport and membrane contact sites that contribute to autophagosome biogenesis. In this context, we discuss lipid biosynthesis and lipid remodeling events that play a crucial role in both phagophore nucleation and expansion. 相似文献
3.
The steric repulsion between proteins on biological membranes is one of the most generic mechanisms that cause membrane shape changes. We present a minimal model in which a spontaneous curvature is induced by asymmetric protein crowding. Our results show that the interplay between the induced spontaneous curvature and the membrane tension determines the energy-minimizing shapes, which describes the wide range of experimentally observed membrane shapes, i.e., flat membranes, spherical vesicles, elongated tubular protrusions, and pearling structures. Moreover, the model gives precise predictions on how membrane shape changes by protein crowding can be tuned by controlling the protein size, the density of proteins, and the size of the crowded domain. 相似文献
4.
Hyun JK Accurso C Hijnen M Schult P Pettikiriarachchi A Mitra AK Coulibaly F 《PLoS pathogens》2011,7(9):e1002239
In contrast to most enveloped viruses, poxviruses produce infectious particles that do not acquire their internal lipid membrane by budding through cellular compartments. Instead, poxvirus immature particles are generated from atypical crescent-shaped precursors whose architecture and composition remain contentious. Here we describe the 2.6 Å crystal structure of vaccinia virus D13, a key structural component of the outer scaffold of viral crescents. D13 folds into two jellyrolls decorated by a head domain of novel fold. It assembles into trimers that are homologous to the double-barrel capsid proteins of adenovirus and lipid-containing icosahedral viruses. We show that, when tethered onto artificial membranes, D13 forms a honeycomb lattice and assembly products structurally similar to the viral crescents and immature particles. The architecture of the D13 honeycomb lattice and the lipid-remodeling abilities of D13 support a model of assembly that exhibits similarities with the giant mimivirus. Overall, these findings establish that the first committed step of poxvirus morphogenesis utilizes an ancestral lipid-remodeling strategy common to icosahedral DNA viruses infecting all kingdoms of life. Furthermore, D13 is the target of rifampicin and its structure will aid the development of poxvirus assembly inhibitors. 相似文献
5.
Willenborg C Jing J Wu C Matern H Schaack J Burden J Prekeris R 《The Journal of cell biology》2011,195(1):71-86
During the morphogenesis of the epithelial lumen, apical proteins are thought to be transported via endocytic compartments to the site of the forming lumen, although the machinery mediating this transport remains to be elucidated. Rab11 GTPase and its binding protein, FIP5, are important regulators of polarized endocytic transport. In this study, we identify sorting nexin 18 as a novel FIP5-interacting protein and characterize the role of FIP5 and SNX18 in epithelial lumen morphogenesis. We show that FIP5 mediates the transport of apical proteins from apical endosomes to the apical plasma membrane and, along with SNX18, is required for the early stages of apical lumen formation. Furthermore, both proteins bind lipids, and FIP5 promotes the capacity of SNX18 to tubulate membranes, which implies a role for FIP5 and SNX18 in endocytic carrier formation and/or scission. In summary, the present findings support the hypothesis that this FIP5-SNX18 complex plays a pivotal role in the polarized transport of apical proteins during apical lumen initiation in epithelial cells. 相似文献
6.
Andrzej Mleczak Sarah Millar Sharon A. Tooze Michael F. Olson Edmond Y.W. Chan 《Cellular signalling》2013,25(1):1-11
Macroautophagy, commonly referred to as autophagy, is a protein degradation pathway that functions at a constitutive level in cells, which may become further activated by stressors such as nutrient starvation or protein aggregation. Autophagy has multiple beneficial roles for maintaining normal cellular homeostasis and these roles are related to the implications of autophagy in disease mechanisms including neurodegeneration and cancer. We previously searched for novel autophagy regulators and identified Rho-kinase 1 (ROCK1) as a candidate. Here, we show that activated ROCK1 inhibits autophagy in human embryonic kidney 293 cells. Conversely, ROCK inhibitory compounds enhanced the autophagy response to amino acid starvation or rapamycin treatment. Inhibition of ROCK during the starvation period led to a more rapid response with the production of larger early autophagosomes that matured into enlarged late degradative autolysosomes. Despite the production of enlarged LC3-positive early autophagosomes, membrane precursors containing WD-repeat protein interacting with phosphoinositides 1 (WIPI1) and mammalian Atg9 were not affected by ROCK inhibition, suggesting that phagophore elongation had been unusually extended. However, the enlarged autophagosomes were enriched in ULK1 which was essential to allow progression of autophagy flux. Our results demonstrate a novel role for ROCK in the control of autophagosome size and degradative capacity. 相似文献
7.
Kathrin Krause Kyle Caution Asmaa Badr Kaitlin Hamilton Abdulmuti Saleh Khushbu Patel 《Autophagy》2018,14(11):1928-1942
CASP4/caspase-11-dependent inflammasome activation is important for the clearance of various Gram-negative bacteria entering the host cytosol. Additionally, CASP4 modulates the actin cytoskeleton to promote the maturation of phagosomes harboring intracellular pathogens such as Legionella pneumophila but not those enclosing nonpathogenic bacteria. Nevertheless, this non-inflammatory role of CASP4 regarding the trafficking of vacuolar bacteria remains poorly understood. Macroautophagy/autophagy, a catabolic process within eukaryotic cells, is also implicated in the elimination of intracellular pathogens such as Burkholderia cenocepacia. Here we show that CASP4-deficient macrophages exhibit a defect in autophagosome formation in response to B. cenocepacia infection. The absence of CASP4 causes an accumulation of the small GTPase RAB7, reduced colocalization of B. cenocepacia with LC3 and acidic compartments accompanied by increased bacterial replication in vitro and in vivo. Together, our data reveal a novel role of CASP4 in regulating autophagy in response to B. cenocepacia infection. 相似文献
8.
Donohue E Tovey A Vogl AW Arns S Sternberg E Young RN Roberge M 《The Journal of biological chemistry》2011,286(9):7290-7300
Autophagy enables cells to degrade and recycle cytoplasmic materials both as a housekeeping mechanism and in response to extracellular stress such as nutrient deprivation. Recent studies indicate that autophagy also functions as a protective mechanism in response to several cancer therapy agents, making it a prospective therapeutic target. Few pharmacological inhibitors suitable for testing the therapeutic potential of autophagy inhibition in vivo are known. An automated microscopy assay was used to screen >3,500 drugs and pharmacological agents and identified one drug, verteporfin, as an inhibitor of autophagosome accumulation. Verteporfin is a benzoporphyrin derivative used in photodynamic therapy, but it inhibits autophagy without light activation. Verteporfin did not inhibit LC3/Atg8 processing or membrane recruitment in response to autophagic stimuli, but it inhibited drug- and starvation-induced autophagic degradation and the sequestration of cytoplasmic materials into autophagosomes. Transient exposure to verteporfin in starvation conditions reduced cell viability whereas cells in nutrient-rich medium were unaffected by drug treatment. Analysis of structural analogs indicated that the activity of verteporfin requires the presence of a substituted cyclohexadiene at ring A of the porphyrin core but that it can tolerate a number of large substituents at rings C and D. The existence of an autophagy inhibitor among FDA-approved drugs should facilitate the investigation of the therapeutic potential of autophagy inhibition in vivo. 相似文献
9.
Moreau K 《Médecine sciences : M/S》2011,27(12):1075-1077
10.
Rhesus monkey rhadinovirus (RRV) is a gamma-2 herpesvirus closely related to human herpesvirus 8 (HHV8). RRV encodes viral FLICE inhibitory protein (vFLIP), which has death effector domains. Little is known about RRV vFLIP. This study intended to examine its function in apoptosis. Here we found that RRV vFLIP inhibits apoptosis induced by tumor necrosis factor-α (TNF-α) and cycloheximide. In HeLa cells with vFLIP expression, the cleavage of poly [ADP-ribose] polymerase 1 (PARP-1) and activities of caspase 3, 7, and 9 were much lower than those in controls. Cell viability of HeLa cells with vFLIP expression was significantly higher than control cells after apoptosis induction. However, RRV vFLIP appears unable to induce NF-κB signaling when tested in NF-κB reporter assay. RRV vFLIP was able to enhance cell survival under starved conditions or apoptosis induction. At early time points after apoptosis induction, autophagosome formation was enhanced and LC3-II level was elevated in cells with vFLIP and, when autophagy was blocked with chemical inhibitors, these cells underwent apoptosis. Moreover, RRV latent infection of BJAB B-lymphoblastoid cells protects the cells against apoptosis by enhancing autophagy to maintain cell survival. Knockdown of vFLIP expression in the RRV-infected BJAB cells with siRNA abolished the protection against apoptosis. These results indicate that vFLIP protects cells against apoptosis by enhancing autophagosome formation to extend cell survival. The finding of vFLIP's inhibition of apoptosis via the autophagy pathway provides insights of vFLIP in RRV pathogenesis. 相似文献
11.
Macroautophagy (in this paper referred to as autophagy) and the ubiquitin-proteasome system are the two major catabolic systems in cells. Autophagy involves sequestration of cytosolic contents in double membrane-bounded vesicles called autophagosomes. The membrane source for autophagosomes has received much attention, and diverse sources, such as the plasma membrane, Golgi, endoplasmic reticulum, and mitochondria, have been implicated. These may not be mutually exclusive, but the exact sources and mechanism involved in the formation of autophagosomes are still unclear. In this paper, we identify a positive role for the small G protein Arf6 in autophagosome formation. The effect of Arf6 on autophagy is mediated by its role in the generation of phosphatidylinositol 4,5-bisphosphate (PIP(2)) and in inducing phospholipase D (PLD) activity. PIP(2) and PLD may themselves promote autophagosome biogenesis by influencing endocytic uptake of plasma membrane into autophagosome precursors. However, Arf6 may also influence autophagy by indirect effects, such as either by regulating membrane flow from other compartments or by modulating PLD activity independently of the mammalian target of rapamycin. 相似文献
12.
13.
The formation of autophagosomes is the central part of the macroautophagy pathway. Little is known, however, about how the participants in this process affect the membrane dynamics at the phagophore assembly site (PAS). Recently, we demonstrated that Atg8, a lipid-conjugated ubiquitin-like protein, controls the expansion of the phagophore. In addition, we showed that the autophagosome formation process can be traced and dissected by time-lapse fluorescence microscopy observation of GFP-Atg8. These findings constitute one step further in our understanding of autophagosome formation. Key questions remain open, however, on how the actions of other proteins at the PAS are coordinated with that of Atg8 and on the precise role of Atg8. 相似文献
14.
The conserved Ser/Thr kinase Atg1/ULK1 plays a crucial role in the regulation of autophagy. However, only very few Atg1 targets have been identified, impeding elucidation of the mechanisms by which Atg1 regulates autophagy. In our study, we determined the Saccharomyces cerevisiae Atg1 consensus phosphorylation sequence using a peptide array-based approach. Among proteins containing this sequence we identified Atg9, another essential component of the autophagic machinery. We showed that phosphorylation of Atg9 by Atg1 is required for phagophore elongation, shedding light on the mechanism by which Atg1 regulates early steps of autophagy. 相似文献
15.
《Biophysical journal》2022,121(18):3411-3421
The inner membrane-associated protein of 30 kDa (IM30) is essential in chloroplasts and cyanobacteria. The spatio-temporal cellular localization of the protein appears to be highly dynamic and triggered by internal as well as external stimuli, mainly light intensity. The soluble fraction of the protein is localized in the cyanobacterial cytoplasm or the chloroplast stroma, respectively. Additionally, the protein attaches to the thylakoid membrane as well as to the chloroplast inner envelope or the cyanobacterial cytoplasmic membrane, respectively, especially under conditions of membrane stress. IM30 is involved in thylakoid membrane biogenesis and/or maintenance, where it either stabilizes membranes and/or triggers membrane-fusion processes. These apparently contradicting functions have to be tightly controlled and separated spatiotemporally in chloroplasts and cyanobacteria. IM30’s fusogenic activity depends on Mg2+ binding to IM30; yet, it still is unclear how Mg2+-loaded IM30 interacts with membranes and promotes membrane fusion. Here, we show that the interaction of Mg2+ with IM30 results in increased binding of IM30 to native, as well as model, membranes. Via atomic force microscopy in liquid, IM30-induced bilayer defects were observed in solid-supported bilayers in the presence of Mg2+. These structures differ dramatically from the membrane-stabilizing carpet structures that were previously observed in the absence of Mg2+. Thus, Mg2+-induced alterations of the IM30 structure switch the IM30 activity from a membrane-stabilizing to a membrane-destabilizing function, a crucial step in membrane fusion. 相似文献
16.
SNX9 promotes metastasis by enhancing cancer cell invasion via differential regulation of RhoGTPases
Nawal Bendris Karla C. Williams Carlos R. Reis Erik S. Welf Ping-Hung Chen Bénédicte Lemmers Michael Hahne Hon Sing Leong Sandra L. Schmid 《Molecular biology of the cell》2016,27(9):1409-1419
Despite current advances in cancer research, metastasis remains the leading factor in cancer-related deaths. Here we identify sorting nexin 9 (SNX9) as a new regulator of breast cancer metastasis. We detect an increase in SNX9 expression in human breast cancer metastases compared with primary tumors and demonstrate that SNX9 expression in MDA-MB-231 breast cancer cells is necessary to maintain their ability to metastasize in a chick embryo model. Conversely, SNX9 knockdown impairs this process. In vitro studies using several cancer cell lines derived from a variety of human tumors reveal a role for SNX9 in cell invasion and identify mechanisms responsible for this novel function. We show that SNX9 controls the activation of RhoA and Cdc42 GTPases and also regulates cell motility via the modulation of well-known molecules involved in metastasis, namely RhoA-ROCK and N-WASP. In addition, we find that SNX9 is required for RhoGTPase-dependent, clathrin-independent endocytosis, and in this capacity can functionally substitute to the bona fide Rho GAP, GTPase regulator associated with focal adhesion kinase (GRAF1). Taken together, our data establish novel roles for SNX9 as a multifunctional protein scaffold that regulates, and potentially coordinates, several cellular processes that together can enhance cancer cell metastasis. 相似文献
17.
Organization of the pre-autophagosomal structure responsible for autophagosome formation 总被引:6,自引:1,他引:5
Autophagy induced by nutrient depletion is involved in survival during starvation conditions. In addition to starvation-induced autophagy, the yeast Saccharomyces cerevisiae also has a constitutive autophagy-like system, the Cvt pathway. Among 31 autophagy-related (Atg) proteins, the function of Atg17, Atg29, and Atg31 is required specifically for autophagy. In this study, we investigated the role of autophagy-specific (i.e., non-Cvt) proteins under autophagy-inducing conditions. For this purpose, we used atg11Delta cells in which the Cvt pathway is abrogated. The autophagy-unique proteins are required for the localization of Atg proteins to the pre-autophagosomal structure (PAS), the putative site for autophagosome formation, under starvation condition. It is likely that these Atg proteins function as a ternary complex, because Atg29 and Atg31 bind to Atg17. The Atg1 kinase complex (Atg1-Atg13) is also essential for recruitment of Atg proteins to the PAS. The assembly of Atg proteins to the PAS is observed only under autophagy-inducing conditions, indicating that this structure is specifically involved in autophagosome formation. Our results suggest that Atg1 complex and the autophagy-unique Atg proteins cooperatively organize the PAS in response to starvation signals. 相似文献
18.
Thomas Clairfeuille Suzanne J. Norwood Xiaying Qi Rohan D. Teasdale Brett M. Collins 《The Journal of biological chemistry》2015,290(23):14504-14517
Sorting nexins (SNX) orchestrate membrane trafficking and signaling events required for the proper distribution of proteins within the endosomal network. Their phox homology (PX) domain acts as a phosphoinositide (PI) recognition module that targets them to specific endocytic membrane domains. The modularity of SNX proteins confers a wide variety of functions from signaling to membrane deformation and cargo binding, and many SNXs are crucial modulators of endosome dynamics and are involved in a myriad of physiological and pathological processes such as neurodegenerative diseases, cancer, and inflammation. Here, we have studied the poorly characterized SNX20 and its paralogue SNX21, which contain an N-terminal PX domain and a C-terminal PX-associated B (PXB) domain of unknown function. The two proteins share similar PI-binding properties and are recruited to early endosomal compartments by their PX domain. The crystal structure of the SNX21 PXB domain reveals a tetratricopeptide repeat (TPR)-fold, a module that typically binds short peptide motifs, with three TPR α-helical repeats. However, the C-terminal capping helix adopts a highly unusual and potentially self-inhibitory topology. SAXS solution structures of SNX20 and SNX21 show that these proteins adopt a compact globular architecture, and membrane interaction analyses indicate the presence of overlapping PI-binding sites that may regulate their intracellular localization. This study provides the first structural analysis of this poorly characterized subfamily of SNX proteins, highlighting a likely role as endosome-associated scaffolds. 相似文献
19.