RESUMO
Intracellular transport among organellar compartments occurs in two general ways, by membrane-bound carriers or membrane contacts. Specific circumstances that involve the coordination of these two modes of transport remain to be defined. Studying Coat Protein I (COPI) transport, we find that phosphatidylcholine with short acyl chains (sPC) is delivered through membrane contact from the endoplasmic reticulum (ER) to sites of COPI vesicle formation at the Golgi to support the fission stage. Phosphatidylinositol transfer protein beta (PITPß) plays a key role in this process, with the elucidation of this role advancing a new understanding of how PITPß acts, providing a mechanistic understanding of a specific circumstance when vesicular transport requires membrane contact, and contributing to a basic understanding of how transport carriers in a model intracellular pathway are formed.
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Cancer cell fate has been widely ascribed to mutational changes within protein-coding genes associated with tumor suppressors and oncogenes. In contrast, the mechanisms through which the biophysical properties of membrane lipids influence cancer cell survival, dedifferentiation and metastasis have received little scrutiny. Here, we report that cancer cells endowed with a high metastatic ability and cancer stem cell-like traits employ ether lipids to maintain low membrane tension and high membrane fluidity. Using genetic approaches and lipid reconstitution assays, we show that these ether lipid-regulated biophysical properties permit non-clathrin-mediated iron endocytosis via CD44, leading directly to significant increases in intracellular redox-active iron and enhanced ferroptosis susceptibility. Using a combination of in vitro three-dimensional microvascular network systems and in vivo animal models, we show that loss of ether lipids also strongly attenuates extravasation, metastatic burden and cancer stemness. These findings illuminate a mechanism whereby ether lipids in carcinoma cells serve as key regulators of malignant progression while conferring a unique vulnerability that can be exploited for therapeutic intervention.
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The epidermal growth factor receptor (EGFR) plays important roles in multiple cellular events, including growth, differentiation, and motility. A major mechanism of downregulating EGFR function involves its endocytic transport to the lysosome. Sorting of proteins into intracellular pathways involves cargo adaptors recognizing sorting signals on cargo proteins. A dileucine-based sorting signal has been identified previously for the sorting of endosomal EGFR to the lysosome, but a cargo adaptor that recognizes this signal remains unknown. Here, we find that phosphoglycerate kinase 1 (PGK1) is recruited to endosomal membrane upon its phosphorylation, where it binds to the dileucine sorting signal in EGFR to promote the lysosomal transport of this receptor. We also elucidate two mechanisms that act in concert to promote PGK1 recruitment to endosomal membrane, a lipid-based mechanism that involves phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] and a protein-based mechanism that involves hepatocyte growth factor receptor substrate (Hrs). These findings reveal an unexpected function for a metabolic enzyme and advance the mechanistic understanding of how EGFR is transported to the lysosome.
Assuntos
Receptores ErbB , Fosfoglicerato Quinase , Fosfoglicerato Quinase/metabolismo , Receptores ErbB/metabolismo , Endossomos/metabolismo , Proteínas/metabolismo , Lisossomos/metabolismo , Transporte Proteico/fisiologia , Complexos Endossomais de Distribuição Requeridos para Transporte/metabolismoRESUMO
The Coat Protein I (COPI) complex forms vesicles from Golgi membrane for retrograde transport among the Golgi stacks, and also from the Golgi to the endoplasmic reticulum (ER). We have been elucidating the mechanistic details of COPI vesicle formation through a reconstitution system that involves the incubation of Golgi membrane with purified components. This approach has enabled us recently to gain new insight into how certain lipids are critical for the fission stage of COPI vesicle formation. Lipid geometry has been proposed to act in the formation of transport carriers by promoting membrane curvature. However, evidence for this role has come from studies using simplified membranes, while confirmation in the more physiologic setting of native membranes has been challenging, as such membranes contain a complex composition of lipids and proteins. We have recently refined the COPI reconstitution system to overcome this experimental obstacle. This has led us to identify an unanticipated type of lipid geometry needed for COPI vesicle fission. This chapter describes the approach that we have developed to enable this discovery. The methodologies include: (i) preparation Golgi membrane from cells that are deficient in a particular lipid enzyme activity and (ii) functional rescue of this deficiency by introducing the product of the lipid enzyme, with experiments being performed at the in vitro level to gain mechanistic clarity and at the in vivo level to confirm physiologic relevance.
Assuntos
Vesículas Revestidas pelo Complexo de Proteína do Envoltório , Complexo de Golgi , Vesículas Revestidas pelo Complexo de Proteína do Envoltório/metabolismo , Complexo de Golgi/metabolismo , Retículo Endoplasmático/metabolismo , Complexo I de Proteína do Envoltório/metabolismo , LipídeosRESUMO
The ADP-Ribosylation Factor (ARF) small GTPases have been found to act in vesicle fission through a direct ability to tubulate membrane. Here, we have used cryo-electron microscopy (EM) to solve the structure of an ARF6 protein lattice assembled on tubulated membrane to 3.9 Å resolution. ARF6 forms tetramers that polymerize into helical arrays to form this lattice. We identify, and confirm functionally, protein contacts critical for this lattice formation. The solved structure also suggests how the ARF amphipathic helix is positioned in the lattice for membrane insertion, and how a GTPase-activating protein (GAP) docks onto the lattice to catalyze ARF-GTP hydrolysis in completing membrane fission. As ARF1 and ARF6 are structurally conserved, we have also modeled ARF1 onto the ARF6 lattice, which has allowed us to pursue the reconstitution of Coat Protein I (COPI) vesicles to confirm more definitively that the ARF lattice acts in vesicle fission. Our findings are notable for having achieved the first detailed glimpse of how a small GTPase bends membrane and having provided a molecular understanding of how an ARF protein acts in vesicle fission.
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G protein-coupled receptors (GPCRs) play crucial roles in numerous physiological and pathological processes. Mutations in GPCRs that result in loss of function or alterations in signaling can lead to inherited or acquired diseases. Herein, studying prokineticin receptor 2 (PROKR2), we initially identify distinct interactomes for wild-type (WT) versus a mutant (P290S) PROKR2 that causes hypogonadotropic hypogonadism. We then find that both the WT and mutant PROKR2 are targeted for endoplasmic reticulum (ER)-associated degradation, but the mutant is degraded to a greater extent. Further analysis revealed that both forms can also leave the ER to reach the Golgi. However, whereas most of the WT is further transported to the cell surface, most of the mutant is retrieved to the ER. Thus, the post-ER itinerary plays an important role in distinguishing the ultimate fate of the WT versus the mutant. We have further discovered that this post-ER itinerary reduces ER stress induced by the mutant PROKR2. Moreover, we extend the core findings to another model GPCR. Our findings advance the understanding of disease pathogenesis induced by a mutation at a key residue that is conserved across many GPCRs and thus contributes to a fundamental understanding of the diverse mechanisms used by cellular quality control to accommodate misfolded proteins.
Assuntos
Estresse do Retículo Endoplasmático/fisiologia , Proteostase/fisiologia , Receptores Acoplados a Proteínas G/metabolismo , Receptores de Peptídeos/metabolismo , Animais , Células COS , Membrana Celular/metabolismo , Chlorocebus aethiops , Retículo Endoplasmático/metabolismo , Degradação Associada com o Retículo Endoplasmático , Complexo de Golgi/metabolismo , Células HEK293 , Células HeLa , Humanos , Hipogonadismo/metabolismo , Mutação de Sentido Incorreto/genética , Transporte Proteico/genética , Transporte Proteico/fisiologia , Receptores Acoplados a Proteínas G/genética , Receptores de Peptídeos/genética , Transdução de SinaisRESUMO
The Golgi apparatus, the main glycosylation station of the cell, consists of a stack of discontinuous cisternae. Glycosylation enzymes are usually concentrated in one or two specific cisternae along the cis-trans axis of the organelle. How such compartmentalized localization of enzymes is achieved and how it contributes to glycosylation are not clear. Here, we show that the Golgi matrix protein GRASP55 directs the compartmentalized localization of key enzymes involved in glycosphingolipid (GSL) biosynthesis. GRASP55 binds to these enzymes and prevents their entry into COPI-based retrograde transport vesicles, thus concentrating them in the trans-Golgi. In genome-edited cells lacking GRASP55, or in cells expressing mutant enzymes without GRASP55 binding sites, these enzymes relocate to the cis-Golgi, which affects glycosphingolipid biosynthesis by changing flux across metabolic branch points. These findings reveal a mechanism by which a matrix protein regulates polarized localization of glycosylation enzymes in the Golgi and controls competition in glycan biosynthesis.
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Glicoesfingolipídeos/metabolismo , Complexo de Golgi/metabolismo , Proteínas da Matriz do Complexo de Golgi/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Autoantígenos/genética , Autoantígenos/metabolismo , Brefeldina A/farmacologia , Ceramidas/metabolismo , Toxina da Cólera/farmacologia , Proteínas do Citoesqueleto/genética , Proteínas do Citoesqueleto/metabolismo , Expressão Gênica , Glicosilação/efeitos dos fármacos , Complexo de Golgi/efeitos dos fármacos , Complexo de Golgi/genética , Proteínas da Matriz do Complexo de Golgi/genética , Células HeLa , Humanos , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Toxina Shiga/farmacologiaRESUMO
Chronic obstructive pulmonary disease (COPD) is a common, complex disease and a major cause of morbidity and mortality. Although multiple genetic determinants of COPD have been implicated by genome-wide association studies (GWASs), the pathophysiological significance of these associations remains largely unknown. From a COPD protein-protein interaction network module, we selected a network path between two COPD GWAS genes for validation studies: FAM13A (family with sequence similarity 13 member A)-AP3D1-CTGF- TGFß2. We find that TGFß2, FAM13A, and AP3D1 (but not CTGF) form a cellular protein complex. Functional characterization suggests that this complex mediates the secretion of TGFß2 through an AP-3 (adaptor protein 3)-dependent pathway, with FAM13A acting as a negative regulator by targeting a late stage of this transport that involves the dissociation of coat-cargo interaction. Moreover, we find that TGFß2 is a transmembrane protein that engages the AP-3 complex for delivery to the late endosomal compartments for subsequent secretion through exosomes. These results identify a pathophysiological context that unifies the biological network role of two COPD GWAS proteins and reveal novel mechanisms of cargo transport through an intracellular pathway.
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Complexo 3 de Proteínas Adaptadoras/metabolismo , Subunidades delta do Complexo de Proteínas Adaptadoras/metabolismo , Proteínas Ativadoras de GTPase/metabolismo , Doença Pulmonar Obstrutiva Crônica/genética , Doença Pulmonar Obstrutiva Crônica/metabolismo , Fator de Crescimento Transformador beta2/metabolismo , Complexo 3 de Proteínas Adaptadoras/genética , Subunidades delta do Complexo de Proteínas Adaptadoras/genética , Linhagem Celular , Exossomos/metabolismo , Proteínas Ativadoras de GTPase/genética , Estudo de Associação Genômica Ampla , Células HEK293 , Humanos , Mapas de Interação de Proteínas/genética , Transporte Proteico , Reprodutibilidade dos Testes , Fator de Crescimento Transformador beta2/genéticaRESUMO
STUDY DESIGN: Prospective, multicenter, partially randomized. OBJECTIVE: Assess rates of complications, revision surgery, and radiation between Mazor robotic-guidance (RG) and fluoro-guidance (FG). SUMMARY OF BACKGROUND DATA: Minimally invasive surgery MIS ReFRESH is the first study designed to compare RG and FG techniques in adult minimally invasive surgery (MIS) lumbar fusions. METHODS: Primary endpoints were analyzed at 1âyear follow-up. Analysis of variables through Cox logistic regression and a Kaplan-Meier Survival Curve of surgical complications. RESULTS: Nine sites enrolled 485 patients: 374 (RG arm) and 111 (FG arm). 93.2% of patients had more than 1âyear f/u. There were no differences for sex, Charlson Comorbidity Index, diabetes, or tumor. Mean age of RG patients was 59.0 versus 62.5 for FG (Pâ=â0.009) and body mass index (BMI) was 31.2 versus 28.1 (P< 0.001). Percentage of smokers was almost double in the RG (15.2% vs. 7.2%, Pâ=â0.029). Surgical time was similar (skin-to-skin time/no. of screws) at 24.9 minutes RG and 22.9 FG (Pâ=â0.550). Fluoroscopy during surgery/no. of screws was 15.5âseconds RG versus 35.4âseconds FG, (15 seconds average reduction). Fluoroscopy time during instrumentation/no. of screws was 3.6âseconds RG versus 17.8âseconds FG showing an 80% average reduction of fluoro time/screw in RG (Pâ<â0.001). Within 1âyear follow-up, there were 39 (10.4%) surgical complications RG versus 39 (35.1%) FG, and 8 (2.1%) revisions RG versus 7 (6.3%) FG. Cox regression analysis including age, sex, BMI, CCI, and no. of screws, demonstrated that the hazard ratio (HR) for complication was 5.8 times higher FG versus RG (95% CI: 3.5-9.6, Pâ<â0.001). HR for revision surgery was 11.0 times higher FG versus RG cases (95% CI 2.9-41.2, Pâ<â0.001). CONCLUSION: Mazor robotic-guidance was found to have a 5.8 times lower risk of a surgical complication and 11.0 times lower risk for revision surgery. Surgical time was similar between groups and robotic-guidance reduced fluoro time per screw by 80% (approximately 1âmin/case).Level of Evidence: 2.
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Parafusos Pediculares , Procedimentos Cirúrgicos Robóticos , Fusão Vertebral , Adulto , Fluoroscopia , Humanos , Vértebras Lombares , Procedimentos Cirúrgicos Minimamente Invasivos , Estudos Prospectivos , Estudos Retrospectivos , Procedimentos Cirúrgicos Robóticos/efeitos adversos , Fusão Vertebral/efeitos adversosRESUMO
The sorting nexin (SNX) family of proteins deform the membrane to generate transport carriers in endosomal pathways. Here, we elucidate how a prototypic member, SNX1, acts in this process. Performing cryoelectron microscopy, we find that SNX1 assembles into a protein lattice that consists of helical rows of SNX1 dimers wrapped around tubular membranes in a crosslinked fashion. We also visualize the details of this structure, which provides a molecular understanding of how various parts of SNX1 contribute to its ability to deform the membrane. Moreover, we have compared the SNX1 structure with a previously elucidated structure of an endosomal coat complex formed by retromer coupled to a SNX, which reveals how the molecular organization of the SNX in this coat complex is affected by retromer. The comparison also suggests insight into intermediary stages of assembly that results in the formation of the retromer-SNX coat complex on the membrane.
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Membrana Celular/metabolismo , Multimerização Proteica , Nexinas de Classificação/metabolismo , Animais , Membrana Celular/química , Membrana Celular/ultraestrutura , Microscopia Crioeletrônica , Camundongos , Estrutura Quaternária de Proteína , Nexinas de Classificação/química , Nexinas de Classificação/ultraestruturaRESUMO
The coat protein I (COPI) complex mediates retrograde trafficking from the Golgi to the endoplasmic reticulum (ER). Five siblings with persistent bacterial and viral infections and defective humoral and cellular immunity had a homozygous p.K652E mutation in the γ1 subunit of COPI (γ1-COP). The mutation disrupts COPI binding to the KDEL receptor and impairs the retrieval of KDEL-bearing chaperones from the Golgi to the ER. Homozygous Copg1K652E mice had increased ER stress in activated T and B cells, poor antibody responses, and normal numbers of T cells that proliferated normally, but underwent increased apoptosis upon activation. Exposure of the mutants to pet store mice caused weight loss, lymphopenia, and defective T cell proliferation that recapitulated the findings in the patients. The ER stress-relieving agent tauroursodeoxycholic acid corrected the immune defects of the mutants and reversed the phenotype they acquired following exposure to pet store mice. This study establishes the role of γ1-COP in the ER retrieval of KDEL-bearing chaperones and thereby the importance of ER homeostasis in adaptive immunity.
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Apoptose/imunologia , Linfócitos B/imunologia , Estresse do Retículo Endoplasmático/imunologia , Ativação Linfocitária , Mutação de Sentido Incorreto , Imunodeficiência Combinada Severa/imunologia , Linfócitos T/imunologia , Substituição de Aminoácidos , Animais , Apoptose/genética , Proteína Coatomer/genética , Retículo Endoplasmático/genética , Retículo Endoplasmático/imunologia , Estresse do Retículo Endoplasmático/genética , Complexo de Golgi/genética , Complexo de Golgi/imunologia , Humanos , Camundongos , Camundongos Mutantes , Receptores de Peptídeos/genética , Receptores de Peptídeos/imunologia , Imunodeficiência Combinada Severa/genéticaRESUMO
ß-Coronaviruses are a family of positive-strand enveloped RNA viruses that includes the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Much is known regarding their cellular entry and replication pathways, but their mode of egress remains uncertain. Using imaging methodologies and virus-specific reporters, we demonstrate that ß-coronaviruses utilize lysosomal trafficking for egress rather than the biosynthetic secretory pathway more commonly used by other enveloped viruses. This unconventional egress is regulated by the Arf-like small GTPase Arl8b and can be blocked by the Rab7 GTPase competitive inhibitor CID1067700. Such non-lytic release of ß-coronaviruses results in lysosome deacidification, inactivation of lysosomal degradation enzymes, and disruption of antigen presentation pathways. ß-Coronavirus-induced exploitation of lysosomal organelles for egress provides insights into the cellular and immunological abnormalities observed in patients and suggests new therapeutic modalities.
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COVID-19/metabolismo , SARS-CoV-2/metabolismo , Via Secretória , Liberação de Vírus , Fatores de Ribosilação do ADP/metabolismo , Animais , COVID-19/patologia , Feminino , Células HeLa , Compostos Heterocíclicos com 2 Anéis/farmacologia , Humanos , Lisossomos , Camundongos , Tioureia/análogos & derivados , Tioureia/farmacologia , Proteínas rab de Ligação ao GTP/antagonistas & inibidores , Proteínas rab de Ligação ao GTP/metabolismo , proteínas de unión al GTP Rab7 , Tratamento Farmacológico da COVID-19RESUMO
The action of guanine nucleotide exchange factors (GEFs) on the ADP-ribosylation factor (ARF) family of small GTPases initiates intracellular transport pathways. This role requires ARF GEFs to be recruited from the cytosol to intracellular membrane compartments. An ARF GEF known as General receptor for 3-phosphoinositides 1 (Grp1) is recruited to the plasma membrane through its pleckstrin homology (PH) domain that recognizes phosphatidylinositol 3,4,5-trisphosphate (PIP3). Here, we find that the phosphorylation of Grp1 induces its PH domain to recognize instead phosphatidylinositol 4-phosphate (PI4P). This phosphorylation also releases an autoinhibitory mechanism that results in the coil-coil (CC) domain of Grp1 engaging two peripheral membrane proteins of the recycling endosome. Because the combination of these actions results in Grp1 being recruited preferentially to the recycling endosome rather than to the plasma membrane, our findings reveal the complexity of recruitment mechanisms that need to be coordinated in localizing an ARF GEF to an intracellular compartment to initiate a transport pathway. Our elucidation is also remarkable for having revealed that phosphoinositide recognition by a PH domain can be switched through its phosphorylation.
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Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/fisiologia , Proteínas de Membrana/metabolismo , Proteínas de Membrana/fisiologia , Fosfatos de Fosfatidilinositol/metabolismo , Células 3T3-L1 , Fatores de Ribosilação do ADP/metabolismo , Sequência de Aminoácidos/genética , Animais , Membrana Celular/metabolismo , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Fosfatos de Inositol/metabolismo , Membranas Intracelulares/metabolismo , Proteínas de Membrana/ultraestrutura , Camundongos , Fosfatidilinositóis/metabolismo , Fosforilação , Receptores Citoplasmáticos e Nucleares/metabolismoRESUMO
Coat proteins have a central role in vesicular transport by binding to cargoes for their sorting into intracellular pathways. Cargo recognition is mediated by components of the coat complex known as adaptor proteins1-3. We previously showed that Arf-GAP with coil-coil, ANK repeat and PH domain-containing protein 1 (ACAP1) functions as an adaptor for a clathrin coat complex that has a function in endocytic recycling4-6. Here, we show that the protein kinase Akt acts as a co-adaptor in this complex, and is needed in conjunction with ACAP1 to bind to cargo proteins to promote their recycling. In addition to advancing the understanding of endocytic recycling, we uncover a fundamentally different function in which a kinase acts, as Akt in this case is an effector rather than a regulator in a cellular event.
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Proteínas Adaptadoras de Transporte Vesicular/metabolismo , Clatrina/metabolismo , Proteínas Ativadoras de GTPase/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Células HEK293 , Células HeLa , Humanos , Integrinas/metabolismo , Ligação Proteica , Receptores da Transferrina/metabolismoRESUMO
The aldehyde dehydrogenase (ALDH) family of metabolic enzymes converts aldehydes to carboxylates. Here, we find that the reductive consequence of ALDH7A1 activity, which generates NADH (nicotinamide adenine dinucleotide, reduced form) from NAD, underlies how ALDH7A1 coordinates a broad inhibition of the intracellular transport pathways. Studying vesicle formation by the Coat Protein I (COPI) complex, we elucidate that NADH generated by ALDH7A1 targets Brefeldin-A ADP-Ribosylated Substrate (BARS) to inhibit COPI vesicle fission. Moreover, defining a physiologic role for the broad transport inhibition exerted by ALDH7A1, we find that it acts to reduce energy consumption during hypoxia and starvation to promote cellular energy homeostasis. These findings advance the understanding of intracellular transport by revealing how the coordination of multiple pathways can be achieved, and also defining circumstances when such coordination is needed, as well as uncovering an unexpected way that NADH acts in cellular energetics.
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Oxirredutases do Álcool/metabolismo , Aldeído Desidrogenase/metabolismo , Proteínas de Ligação a DNA/metabolismo , Metabolismo Energético , Homeostase , Espaço Intracelular/metabolismo , Oxirredutases do Álcool/genética , Aldeído Desidrogenase/genética , Transporte Biológico , Vesículas Revestidas pelo Complexo de Proteína do Envoltório/metabolismo , Hipóxia Celular , Proteínas de Ligação a DNA/genética , Células HEK293 , Células HeLa , Humanos , NAD/metabolismo , Transdução de Sinais , InaniçãoRESUMO
Studies on vesicle formation by the Coat Protein I (COPI) complex have contributed to a basic understanding of how vesicular transport is initiated. Phosphatidic acid (PA) and diacylglycerol (DAG) have been found previously to be required for the fission stage of COPI vesicle formation. Here, we find that PA with varying lipid geometry can all promote early fission, but only PA with shortened acyl chains promotes late fission. Moreover, diacylglycerol (DAG) acts after PA in late fission, with this role of DAG also requiring shorter acyl chains. Further highlighting the importance of the short-chain lipid geometry for late fission, we find that shorter forms of PA and DAG promote the vesiculation ability of COPI fission factors. These findings advance a general understanding of how lipid geometry contributes to membrane deformation for vesicle fission, and also how proteins and lipids coordinate their actions in driving this process.
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Vesículas Revestidas pelo Complexo de Proteína do Envoltório/metabolismo , Diglicerídeos/metabolismo , Ácidos Fosfatídicos/metabolismo , Complexo I de Proteína do Envoltório/metabolismo , Diglicerídeos/química , Complexo de Golgi/metabolismo , Células HeLa , Humanos , Ácidos Fosfatídicos/químicaRESUMO
Studies on the Bin-Amphiphysin-Rvs (BAR) domain have advanced a fundamental understanding of how proteins deform membrane. We previously showed that a BAR domain in tandem with a Pleckstrin Homology (PH domain) underlies the assembly of ACAP1 (Arfgap with Coil-coil, Ankryin repeat, and PH domain I) into an unusual lattice structure that also uncovers a new paradigm for how a BAR protein deforms membrane. Here, we initially pursued computation-based refinement of the ACAP1 lattice to identify its critical protein contacts. Simulation studies then revealed how ACAP1, which dimerizes into a symmetrical structure in solution, is recruited asymmetrically to the membrane through dynamic behavior. We also pursued electron microscopy (EM)-based structural studies, which shed further insight into the dynamic nature of the ACAP1 lattice assembly. As ACAP1 is an unconventional BAR protein, our findings broaden the understanding of the mechanistic spectrum by which proteins assemble into higher-ordered structures to achieve membrane deformation.
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Proteínas Ativadoras de GTPase/metabolismo , Proteínas de Membrana/metabolismo , Membrana Celular/metabolismo , Dimerização , Proteínas Ativadoras de GTPase/química , Humanos , Domínios de Homologia à Plecstrina , Conformação ProteicaRESUMO
Starvation poses a fundamental challenge to cell survival. Whereas the role of autophagy in promoting energy homeostasis in this setting has been extensively characterized1, other mechanisms are less well understood. Here we reveal that glyceraldehyde 3-phosphate dehydrogenase (GAPDH) inhibits coat protein I (COPI) transport by targeting a GTPase-activating protein (GAP) towards ADP-ribosylation factor 1 (ARF1) to suppress COPI vesicle fission. GAPDH inhibits multiple other transport pathways, also by targeting ARF GAPs. Further characterization suggests that this broad inhibition is activated by the cell during starvation to reduce energy consumption. These findings reveal a remarkable level of coordination among the intracellular transport pathways that underlies a critical mechanism of cellular energy homeostasis.
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Metabolismo Energético , Gliceraldeído-3-Fosfato Desidrogenase (Fosforiladora)/metabolismo , Homeostase , Adenilato Quinase/metabolismo , Aminoimidazol Carboxamida/análogos & derivados , Aminoimidazol Carboxamida/metabolismo , Animais , Autofagia , Vesículas Revestidas pelo Complexo de Proteína do Envoltório/metabolismo , Linhagem Celular , Chlorocebus aethiops , Cricetulus , Fibroblastos , Proteínas Ativadoras de GTPase/antagonistas & inibidores , Proteínas Ativadoras de GTPase/metabolismo , Gliceraldeído-3-Fosfato Desidrogenase (Fosforiladora)/química , Humanos , Camundongos , Fosforilação , Ribonucleotídeos/metabolismo , InaniçãoAssuntos
Adenosina Desaminase/deficiência , Agamaglobulinemia/genética , Peptídeos e Proteínas de Sinalização Intercelular/deficiência , Imunodeficiência Combinada Severa/genética , Adenosina Desaminase/sangue , Adenosina Desaminase/genética , Agamaglobulinemia/sangue , Pré-Escolar , Feminino , Glicosilação , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/sangue , Peptídeos e Proteínas de Sinalização Intercelular/genética , Mutação , Imunodeficiência Combinada Severa/sangueRESUMO
Signaling through the Ror2 receptor tyrosine kinase promotes invadopodia formation for tumor invasion. Here, we identify intraflagellar transport 20 (IFT20) as a new target of this signaling in tumors that lack primary cilia, and find that IFT20 mediates the ability of Ror2 signaling to induce the invasiveness of these tumors. We also find that IFT20 regulates the nucleation of Golgi-derived microtubules by affecting the GM130-AKAP450 complex, which promotes Golgi ribbon formation in achieving polarized secretion for cell migration and invasion. Furthermore, IFT20 promotes the efficiency of transport through the Golgi complex. These findings shed new insights into how Ror2 signaling promotes tumor invasiveness, and also advance the understanding of how Golgi structure and transport can be regulated.