scholarly journals Autophagosome Requires Specific Early Sec Proteins for Its Formation and NSF/SNARE for Vacuolar Fusion

2001 ◽  
Vol 12 (11) ◽  
pp. 3690-3702 ◽  
Author(s):  
Naotada Ishihara ◽  
Maho Hamasaki ◽  
Sadaki Yokota ◽  
Kuninori Suzuki ◽  
Yoshiaki Kamada ◽  
...  
Keyword(s):  
Fusion Protein ◽  
Secretory Pathway ◽  
De Novo ◽  
Subcellular Fractionation ◽  
Autophagosome Formation ◽  
Electron Microscopic ◽  
Membrane Flow ◽  
Protein Receptor ◽  
Sec Proteins ◽  
Protein Attachment

Double membrane structure, autophagosome, is formed de novo in the process of autophagy in the yeastSaccharomyces cerevisiae, and many Apg proteins participate in this process. To further understand autophagy, we analyzed the involvement of factors engaged in the secretory pathway. First, we showed that Sec18p (N-ethylmaleimide-sensitive fusion protein, NSF) and Vti1p (solubleN-ethylmaleimide-sensitive fusion protein attachment protein, SNARE), and soluble N-ethylmaleimide-sensitive fusion protein receptor are required for fusion of the autophagosome to the vacuole but are not involved in autophagosome formation. Second, Sec12p was shown to be essential for autophagy but not for the cytoplasm to vacuole-targeting (Cvt) (pathway, which shares mostly the same machinery with autophagy. Subcellular fractionation and electron microscopic analyses showed that Cvt vesicles, but not autophagosomes, can be formed in sec12 cells. Three other coatmer protein (COPII) mutants, sec16, sec23,and sec24, were also defective in autophagy. The blockage of autophagy in these mutants was not dependent on transport from endoplasmic reticulum-to-Golgi, because mutations in two other COPII genes, SEC13 and SEC31, did not affect autophagy. These results demonstrate the requirement for subgroup of COPII proteins in autophagy. This evidence demonstrating the involvement of Sec proteins in the mechanism of autophagosome formation is crucial for understanding membrane flow during the process.

scholarly journals The human SNARE protein Ykt6 mediates its own palmitoylation at C-terminal cysteine residues

2004 ◽  
Vol 384 (2) ◽  
pp. 233-237 ◽  
Author(s):  
Michael VEIT
Keyword(s):  
Fusion Protein ◽  
Binding Site ◽  
Covalent Attachment ◽  
Receptor Protein ◽  
Cysteine Residues ◽  
Snare Protein ◽  
Attachment Protein ◽  
Present Evidence ◽  
Protein Receptor ◽  
Protein Attachment

The yeast SNARE (soluble N-ethylmaleimide-sensitive fusion protein attachment protein receptor) protein Ykt6 was shown to mediate palmitoylation of the fusion factor Vac8 in a reaction essential for the fusion of vacuoles. Here I present evidence that hYkt6 (human Ykt6) has self-palmitoylating activity. Incubation of recombinant hYkt6 with [3H]Pal-CoA ([3H]palmitoyl-CoA) leads to covalent attachment of palmitate to C-terminal cysteine residues. The N-terminal domain of human Ykt6 contains a Pal-CoA binding site and is required for the reaction.

scholarly journals Structural Changes Are Associated with SolubleN-Ethylmaleimide-sensitive Fusion Protein Attachment Protein Receptor Complex Formation

1997 ◽  
Vol 272 (44) ◽  
pp. 28036-28041 ◽  
Author(s):  
Dirk Fasshauer ◽  
Henning Otto ◽  
William K. Eliason ◽  
Reinhard Jahn ◽  
Axel T. Brünger
Keyword(s):  
Complex Formation ◽  
Fusion Protein ◽  
Structural Changes ◽  
Receptor Complex ◽  
Attachment Protein ◽  
Protein Receptor ◽  
Protein Attachment

Regulation of SNAP-25 trafficking and function by palmitoylation

2010 ◽  
Vol 38 (1) ◽  
pp. 163-166 ◽  
Author(s):  
Jennifer Greaves ◽  
Gerald R. Prescott ◽  
Oforiwa A. Gorleku ◽  
Luke H. Chamberlain
Keyword(s):  
Fusion Protein ◽  
Intracellular Trafficking ◽  
Neuroendocrine Cells ◽  
Snare Proteins ◽  
Receptor Protein ◽  
Attachment Protein ◽  
Rich Region ◽  
Protein Receptor ◽  
And Function ◽  
Protein Attachment

The SNARE (soluble N-ethylmaleimide-sensitive fusion protein-attachment protein receptor) protein SNAP-25 (25 kDa synaptosome-associated protein) is essential for regulated exocytosis in neuronal and neuroendocrine cells. Whereas the majority of SNARE proteins contain transmembrane domains, SNAP-25 is instead anchored to membranes by the palmitoylation of a central cysteine-rich region. In this review, we discuss the mechanisms of SNAP-25 palmitoylation and how this modification regulates the intracellular trafficking and exocytotic function of this essential protein.

scholarly journals Unconventional secretion of Pichia pastoris Acb1 is dependent on GRASP protein, peroxisomal functions, and autophagosome formation

2010 ◽  
Vol 188 (4) ◽  
pp. 537-546 ◽  
Author(s):  
Ravi Manjithaya ◽  
Christophe Anjard ◽  
William F. Loomis ◽  
Suresh Subramani
Keyword(s):  
Pichia Pastoris ◽  
Fatty Acyl ◽  
Autophagosome Formation ◽  
Attachment Protein ◽  
Protein Receptor ◽  
Evolutionarily Conserved ◽  
Unconventional Secretion ◽  
Acyl Coenzyme A ◽  
And Function ◽  
Protein Attachment

In contrast to the enormous advances made regarding mechanisms of conventional protein secretion, mechanistic insights into the unconventional secretion of proteins are lacking. Acyl coenzyme A (CoA)–binding protein (ACBP; AcbA in Dictyostelium discoideum), an unconventionally secreted protein, is dependent on Golgi reassembly and stacking protein (GRASP) for its secretion. We discovered, surprisingly, that the secretion, processing, and function of an AcbA-derived peptide, SDF-2, are conserved between the yeast Pichia pastoris and D. discoideum. We show that in yeast, the secretion of SDF-2–like activity is GRASP dependent, triggered by nitrogen starvation, and requires autophagy proteins as well as medium-chain fatty acyl CoA generated by peroxisomes. Additionally, a phospholipase D implicated in soluble N-ethyl-maleimide sensitive fusion protein attachment protein receptor–mediated vesicle fusion at the plasma membrane is necessary, but neither peroxisome turnover nor fusion between autophagosomes and the vacuole is essential. Moreover, yeast Acb1 and several proteins required for its secretion are necessary for sporulation in P. pastoris. Our findings implicate currently unknown, evolutionarily conserved pathways in unconventional secretion.

scholarly journals Homotypic Fusion of Immature Secretory Granules During Maturation Requires Syntaxin 6

2001 ◽  
Vol 12 (6) ◽  
pp. 1699-1709 ◽  
Author(s):  
Franz Wendler ◽  
Lesley Page ◽  
Sylvie Urbé ◽  
Sharon A. Tooze
Keyword(s):  
Fusion Protein ◽  
Indirect Immunofluorescence ◽  
Immunofluorescence Microscopy ◽  
Secretory Granules ◽  
Subcellular Fractionation ◽  
Neuroendocrine Cells ◽  
The Core ◽  
Indirect Immunofluorescence Microscopy ◽  
A Cell ◽  
Protein Attachment

Homotypic fusion of immature secretory granules (ISGs) gives rise to mature secretory granules (MSGs), the storage compartment in endocrine and neuroendocrine cells for hormones and neuropeptides. With the use of a cell-free fusion assay, we investigated which soluble N-ethylmaleimide-sensitive fusion protein attachment receptor (SNARE) molecules are involved in the homotypic fusion of ISGs. Interestingly, the SNARE molecules mediating the exocytosis of MSGs in neuroendocrine cells, syntaxin 1, SNAP-25, and VAMP2, were not involved in homotypic ISG fusion. Instead, we have identified syntaxin 6 as a component of the core machinery responsible for homotypic ISG fusion. Subcellular fractionation studies and indirect immunofluorescence microscopy show that syntaxin 6 is sorted away during the maturation of ISGs to MSGs. Although, syntaxin 6 on ISG membranes is associated with SNAP-25 and SNAP-29/GS32, we could not find evidence that these target (t)-SNARE molecules are involved in homotypic ISG fusion. Nor could we find any involvement for the vesicle (v)-SNARE VAMP4, which is known to be associated with syntaxin 6. Importantly, we have shown that homotypic fusion requires the function of syntaxin 6 on both donor as well as acceptor membranes, which suggests that t–t-SNARE interactions, either direct or indirect, may be required during fusion of ISG membranes.

scholarly journals Prospore Membrane Formation Defines a Developmentally Regulated Branch of the Secretory Pathway in Yeast

1998 ◽  
Vol 140 (1) ◽  
pp. 29-37 ◽  
Author(s):  
Aaron M. Neiman
Keyword(s):  
Cell Division ◽  
Mother Cell ◽  
Plasma Membranes ◽  
Secretory Pathway ◽  
De Novo ◽  
Extracellular Proteins ◽  
Spore Formation ◽  
Electron Microscopic ◽  
Developmentally Regulated ◽  
Prospore Membrane

Spore formation in yeast is an unusual form of cell division in which the daughter cells are formed within the mother cell cytoplasm. This division requires the de novo synthesis of a membrane compartment, termed the prospore membrane, which engulfs the daughter nuclei. The effect of mutations in late-acting genes on sporulation was investigated. Mutation of SEC1, SEC4, or SEC8 blocked spore formation, and electron microscopic analysis of the sec4-8 mutant indicated that this inability to produce spores was caused by a failure to form the prospore membrane. The soluble NSF attachment protein 25 (SNAP-25) homologue SEC9, by contrast, was not required for sporulation. The absence of a requirement for SEC9 was shown to be due to the sporulation-specific induction of a second, previously undescribed, SNAP-25 homologue, termed SPO20. These results define a developmentally regulated branch of the secretory pathway and suggest that spore morphogenesis in yeast proceeds by the targeting and fusion of secretory vesicles to form new plasma membranes in the interior of the mother cell. Consistent with this model, the extracellular proteins Gas1p and Cts1p were localized to an internal compartment in sporulating cells. Spore formation in yeast may be a useful model for understanding secretion-driven cell division events in a variety of plant and animal systems.

The Morphology of the Rat Kidney Following Folic Acid Administration

1970 ◽  
Vol 28 ◽  
pp. 200-201
Author(s):  
Aline Byrnes ◽  
Elsa E. Ramos ◽  
Minoru Suzuki ◽  
E.D. Mayfield
Keyword(s):  
Folic Acid ◽  
De Novo ◽  
Specific Activity ◽  
Rat Kidney ◽  
Present Report ◽  
Intracellular Localization ◽  
Male Rats ◽  
Renal Hypertrophy ◽  
Electron Microscopic ◽  
Biochemical Alterations

Renal hypertrophy was induced in 100 g male rats by the injection of 250 mg folic acid (FA) dissolved in 0.3 M NaHCO3/kg body weight (i.v.). Preliminary studies of the biochemical alterations in ribonucleic acid (RNA) metabolism of the renal tissue have been reported recently (1). They are: RNA content and concentration, orotic acid-c14 incorporation into RNA and acid soluble nucleotide pool, intracellular localization of the newly synthesized RNA, and the specific activity of enzymes of the de novo pyrimidine biosynthesis pathway. The present report describes the light and electron microscopic observations in these animals. For light microscopy, kidney slices were fixed in formalin, embedded, sectioned, and stained with H & E and PAS.

scholarly journals Structural catalog of core Atg proteins opens new era of autophagy research

2021 ◽  
Author(s):  
Kazuaki Matoba ◽  
Nobuo N Noda
Keyword(s):  
De Novo ◽  
Structural Information ◽  
Membrane Dynamics ◽  
Autophagosome Formation ◽  
New Era ◽  
Intracellular Degradation ◽  
Atg Proteins ◽  
Biological Studies ◽  
Evolutionarily Conserved ◽  
Biological Methods

Summary Autophagy, which is an evolutionarily conserved intracellular degradation system, involves de novo generation of autophagosomes that sequester and deliver diverse cytoplasmic materials to the lysosome for degradation. Autophagosome formation is mediated by approximately 20 core autophagy-related (Atg) proteins, which collaborate to mediate complicated membrane dynamics during autophagy. To elucidate the molecular functions of these Atg proteins in autophagosome formation, many researchers have tried to determine the structures of Atg proteins by using various structural biological methods. Although not sufficient, the basic structural catalog of all core Atg proteins was established. In this review article, we summarize structural biological studies of core Atg proteins, with an emphasis on recently unveiled structures, and describe the mechanistic breakthroughs in autophagy research that have derived from new structural information.

Sign in / Sign up

Export Citation Format

Share Document