RESUMO
Endosomal sorting complex required for transport (ESCRT) proteins can promote extreme membrane deformations, including scission and sealing. New work uncovers a link between these proteins and the early secretory pathway that is functionally important for programmed autophagy during Drosophila development.
Assuntos
Complexos Endossomais de Distribuição Requeridos para Transporte , Endossomos , Autofagia , Complexos Endossomais de Distribuição Requeridos para Transporte/metabolismo , Endossomos/metabolismo , Transporte ProteicoRESUMO
Clearance of misfolded proteins from the secretory pathway often occurs soon after their biosynthesis by endoplasmic reticulum (ER)-associated protein degradation (ERAD). However, certain types of misfolded proteins are not ERAD substrates and exit the ER. They are then scrutinized by ill-defined post-ER quality control (post-ERQC) mechanisms and are frequently routed to the vacuole/lysosome for degradation. Glycosylphosphatidylinositol-anchored proteins (GPI-APs) constitute a class of proteins of the secretory pathway that mostly depends on post-ERQC. How misfolded GPI-APs are detected, transported to the vacuole/lysosome and taken up by this organelle was poorly defined. Originating from the intriguing observation that several misfolded GPI-APs accumulate in the yeast vacuolar membrane in the absence of the major vacuolar protease Pep4, we designed an unbiased genome-wide screen in yeast and followed the trafficking of the misfolded fluorescent GPI-AP Gas1* from the ER to the vacuolar lumen. Our results reveal that post-ERQC of GPI-APs is linked with a novel type of microautophagy.
Assuntos
Microautofagia , Proteínas de Saccharomyces cerevisiae , Ácido Aspártico Endopeptidases/metabolismo , Autofagia , Retículo Endoplasmático/metabolismo , Proteínas Ligadas por GPI/metabolismo , Glicosilfosfatidilinositóis/metabolismo , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismoRESUMO
Glycosylphosphatidylinositol-anchored proteins (GPI-APs) are membrane-conjugated cell-surface proteins with diverse structural, developmental, and signaling functions and clinical relevance. Typically, after biosynthesis and attachment to the preassembled GPI anchor, GPI-APs rapidly leave the endoplasmic reticulum (ER) and rely on post-ER quality control. Terminally misfolded GPI-APs end up inside the vacuole/lysosome for degradation, but their trafficking itinerary to this organelle and the processes linked to their uptake by the vacuole/lysosome remain uncharacterized. In a yeast mutant that is lacking Pep4, a key vacuolar protease, several misfolded model GPI-APs accumulated in the vacuolar membrane. In the same mutant, macroautophagy and the multi-vesicular body (MVB) pathway were intact, hinting at a hitherto-unknown trafficking pathway for the degradation of misfolded GPI-APs. To unravel it, we used a genome-wide screen coupled to high-throughput fluorescence microscopy and followed the fate of the misfolded GPI-AP: Gas1∗. We found that components of the early secretory and endocytic pathways are involved in its targeting to the vacuole and that vacuolar transporter chaperones (VTCs), with roles in microautophagy, negatively affect the vacuolar uptake of Gas1∗. In support, we demonstrate that Gas1∗ internalizes from vacuolar membranes into membrane-bound intravacuolar vesicles prior to degradation. Our data link post-ER degradation with microautophagy.
Assuntos
Glicosilfosfatidilinositóis , Microautofagia , Retículo Endoplasmático/metabolismo , Proteínas Ligadas por GPI/metabolismo , Glicosilfosfatidilinositóis/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismoRESUMO
Endoplasmic reticulum (ER) quality control mechanisms target terminally misfolded proteins for ER-associated degradation (ERAD). Misfolded glycophosphatidylinositol-anchored proteins (GPI-APs) are, however, generally poor ERAD substrates and are targeted mainly to the vacuole/lysosome for degradation, leading to predictions that a GPI anchor sterically obstructs ERAD. Here we analyzed the degradation of the misfolded GPI-AP Gas1* in yeast. We could efficiently route Gas1* to Hrd1-dependent ERAD and provide evidence that it contains a GPI anchor, ruling out that a GPI anchor obstructs ERAD. Instead, we show that the normally decreased susceptibility of Gas1* to ERAD is caused by canonical remodeling of its GPI anchor, which occurs in all GPI-APs and provides a protein-independent ER export signal. Thus, GPI anchor remodeling is independent of protein folding and leads to efficient ER export of even misfolded species. Our data imply that ER quality control is limited for the entire class of GPI-APs, many of them being clinically relevant.
Assuntos
Degradação Associada com o Retículo Endoplasmático/fisiologia , Retículo Endoplasmático/metabolismo , Retículo Endoplasmático/fisiologia , Proteínas Fúngicas/metabolismo , Glicosilfosfatidilinositóis/metabolismo , Ligação Proteica/fisiologia , Dobramento de Proteína , Leveduras/metabolismo , Leveduras/fisiologiaRESUMO
The endoplasmic reticulum (ER) is a major source for the generation of autophagosomes during macroautophagy. Our recent work in yeast shows that particular ER-derived vesicles are generated for the biogenesis of autophagosomes. These vesicles not only incorporate a SNARE protein that is largely ER-resident under nonstarving conditions, but also display COPII requirements for ER-exit that differ from conventional cargo-transporting vesicles. Our results suggest that specific intracellular traffic is launched at the ER for the transport of membranes to sites of autophagosome formation.
Assuntos
Autofagossomos/metabolismo , Vesículas Revestidas pelo Complexo de Proteína do Envoltório/metabolismo , Retículo Endoplasmático/metabolismo , Proteínas SNARE/metabolismo , Animais , Humanos , Modelos BiológicosRESUMO
The de novo formation of autophagosomes for the targeting of cytosolic material to the vacuole/lysosome is upregulated upon starvation. How autophagosomes acquire membranes remains still unclear. Here, we report that, in yeast, the endoplasmic reticulum (ER)-localized Qa/t-SNARE Ufe1 has a role in autophagy. During starvation, Ufe1 is increasingly exported from the ER and targeted to intracellular sites that contain the autophagy markers Atg8 and Atg9. In addition, Ufe1 interacts with non-ER SNARE proteins implicated in autophagosome formation. Loss of Ufe1 function impairs autophagy and results in fewer and smaller autophagosomes. Unlike conventional cargo, the ER export of Ufe1 is significantly reduced in sec23-1 cells, which affects the coat protein (COP)II complex, already at the permissive temperature. Under the same conditions, sec23-1 cells are hypersensitive to starvation and deficient in autophagy. Our data suggest that ER membranes containing Ufe1 are delivered to sites of autophagosome formation in specific COPII vesicles.
Assuntos
Vesículas Revestidas pelo Complexo de Proteína do Envoltório/metabolismo , Retículo Endoplasmático/metabolismo , Regulação Fúngica da Expressão Gênica , Biogênese de Organelas , Fagossomos/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/genética , Autofagia , Família da Proteína 8 Relacionada à Autofagia , Proteínas Relacionadas à Autofagia , Vesículas Revestidas pelo Complexo de Proteína do Envoltório/ultraestrutura , Meios de Cultura/farmacologia , Retículo Endoplasmático/efeitos dos fármacos , Retículo Endoplasmático/ultraestrutura , Lisossomos/efeitos dos fármacos , Lisossomos/metabolismo , Lisossomos/ultraestrutura , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Nitrogênio/deficiência , Fagossomos/efeitos dos fármacos , Fagossomos/ultraestrutura , Transporte Proteico , Proteínas Qa-SNARE/genética , Proteínas Qa-SNARE/metabolismo , Saccharomyces cerevisiae/efeitos dos fármacos , Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/ultraestrutura , Proteínas de Saccharomyces cerevisiae/metabolismo , Transdução de Sinais , Temperatura , Vacúolos/efeitos dos fármacos , Vacúolos/metabolismo , Vacúolos/ultraestruturaRESUMO
Quality control of protein folding inside the endoplasmic reticulum (ER) includes chaperone-mediated assistance in folding and the selective targeting of terminally misfolded species to a pathway called ER-associated protein degradation, or simply ERAD. Once selected for ERAD, substrates will be transported (back) into the cytosol, a step called retrotranslocation. Although still ill defined, retrotranslocation likely involves a protein conducting channel that is in part formed by specific membrane-embedded E3 ubiquitin ligases. Early during retrotranslocation, reversible self-ubiquitination of these ligases is thought to aid in initiation of substrate transfer across the membrane. Once being at least partially exposed to the cytosol, substrates will become ubiquitinated on the cytosolic side of the ER membrane by the same E3 ubiquitin ligases. Ubiquitin on substrates was originally thought to be a permanent modification that (1) promotes late steps of retrotranslocation by recruiting the energy-providing ATPase Cdc48p/p97 via binding to its associated adaptor proteins and that (2) serves to target substrates to the proteasome. Recently it became evident, however, that the poly-ubiquitin chains (PUCs) on ERAD substrates are often subject to extensive remodeling, or processing, at several stages during ERAD. This review recapitulates the current knowledge and recent findings about PUC processing on ERAD substrates and ubiquitination of ERAD machinery components and discusses their functional consequences.
RESUMO
Las enfermedades periodontales y cardiovasculares son condiciones inflama-torias comunes en la población cubana. Con la enfermedad periodontal, los eventos inflamatorios bucales culminan con la destrucción de los tejidos alrededor del diente. De forma similar, con la enfermedad cardiovascular (ECV) el proceso inflamatorio desempeña un papel preponderante en el desarrollo y ruptura de las placas ateromatosas. A través de esta revisión bibliográfica, se explican los posibles mecanismos de asociación entre ambas enfermedades y se presentan evidencias actualizadas que respaldan esta asociación. Estudios de observación y meta-análisis demuestran consistentemente un incremento modesto, pero estadísticamente significativo en el riesgo de ECV entre las personas que presentan enfermedades periodontales. Estos descubrimientos respaldan la hipótesis de que la exposición a la enfermedad periodontal puede promover la aterogénesis por medio de efectos bacterianos directos sobre las plaquetas, respuestas auto-inmune, invasión y/o captación de las bacterias en las células endoteliales y macrófagos, y efectos similares a los endocrinos en los mediadores proinflamatorios.
Periodontal and cardiovascular diseases are both very common diseases in Cuban population. In periodontal diseases, inflammatory events end with the destruction or the tissues surrounding the teeth. As well as in cardiovascular diseases, the inflammatory process plays a main role in the development and rupture of atheromatose plaques. By means of this revision, the possible absorption mechanisms in both diseases are explained and updated evidences back up this association. Observational studies and metanalysis show this consistently modest event, which is significant statistically in cardiovascular risk among people who have these conditions. These discoveries back up the hypothesis that both exposition to periodontal diseases, may predispose atherogenesis by means of bacterial effects directly over platelets, auto immune response, invasion and/or capitation of the bacteria, auto immune response, invasion or capitation of bacteria in endothelium cells and macrophages and similar endocrine effects in proinflamatory mediators.