scholarly journals Self-Interaction Is Critical for Atg9 Transport and Function at the Phagophore Assembly Site during Autophagy

2008 ◽  
Vol 19 (12) ◽  
pp. 5506-5516 ◽  
Author(s):  
Congcong He ◽  
Misuzu Baba ◽  
Yang Cao ◽  
Daniel J. Klionsky
Keyword(s):  
Membrane Protein ◽  
Membrane Transport ◽  
Integral Membrane Protein ◽  
Human Diseases ◽  
Anterograde Transport ◽  
Membrane Flow ◽  
Phagophore Assembly Site ◽  
And Function ◽  
Assembly Site ◽  
Starvation Conditions

Autophagy is the degradation of a cell's own components within lysosomes (or the analogous yeast vacuole), and its malfunction contributes to a variety of human diseases. Atg9 is the sole integral membrane protein required in formation of the initial sequestering compartment, the phagophore, and is proposed to play a key role in membrane transport; the phagophore presumably expands by vesicular addition to form a complete autophagosome. It is not clear through what mechanism Atg9 functions at the phagophore assembly site (PAS). Here we report that Atg9 molecules self-associate independently of other known autophagy proteins in both nutrient-rich and starvation conditions. Mutational analyses reveal that self-interaction is critical for anterograde transport of Atg9 to the PAS. The ability of Atg9 to self-interact is required for both selective and nonselective autophagy at the step of phagophore expansion at the PAS. Our results support a model in which Atg9 multimerization facilitates membrane flow to the PAS for phagophore formation.

scholarly journals Vacuolar protein Tag1 and Atg1–Atg13 regulate autophagy termination during persistent starvation in S. cerevisiae

2021 ◽  
Vol 134 (4) ◽  
pp. jcs253682
Author(s):  
Shintaro Kira ◽  
Masafumi Noguchi ◽  
Yasuhiro Araki ◽  
Yu Oikawa ◽  
Tamotsu Yoshimori ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Protein Kinase ◽  
Membrane Protein ◽  
Nitrogen Starvation ◽  
Genetic Screen ◽  
Vacuolar Membrane ◽  
Phagophore Assembly Site ◽  
Vacuolar Protein ◽  
Assembly Site ◽  
Starvation Conditions

ABSTRACTUnder starvation conditions, cells degrade their own components via autophagy in order to provide sufficient nutrients to ensure their survival. However, even if starvation persists, the cell is not completely degraded through autophagy, implying the existence of some kind of termination mechanism. In the yeast Saccharomyces cerevisiae, autophagy is terminated after 10–12 h of nitrogen starvation. In this study, we found that termination is mediated by re-phosphorylation of Atg13 by the Atg1 protein kinase, which is also affected by PP2C phosphatases, and the eventual dispersion of the pre-autophagosomal structure, also known as the phagophore assembly site (PAS). In a genetic screen, we identified an uncharacterized vacuolar membrane protein, Tag1, as a factor responsible for the termination of autophagy. Re-phosphorylation of Atg13 and eventual PAS dispersal were defective in the Δtag1 mutant. The vacuolar luminal domain of Tag1 and autophagic progression are important for the behaviors of Tag1. Together, our findings reveal the mechanism and factors responsible for termination of autophagy in yeast.

scholarly journals A 46 kDa integral membrane protein from Mycobacterium leprae resembles a number of bacterial and mammalian membrane transport proteins

Microbiology ◽  
1995 ◽  
Vol 141 (8) ◽  
pp. 1963-1968 ◽  
Author(s):  
L. Oskam ◽  
R. A. Hartskeerl ◽  
C. J. Hermans ◽  
M. Y. L. de Wit ◽  
G. H. Jarings ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Membrane Transport ◽  
Mycobacterium Leprae ◽  
Integral Membrane Protein ◽  
Transport Proteins ◽  
Membrane Transport Proteins

scholarly journals A previously unrecognized membrane protein in the Rhodobacter sphaeroides LH1-RC photocomplex

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Kazutoshi Tani ◽  
Kenji V. P. Nagashima ◽  
Ryo Kanno ◽  
Saki Kawamura ◽  
Riku Kikuchi ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Ring Opening ◽  
Model Organism ◽  
Hypothetical Protein ◽  
Integral Membrane Protein ◽  
Bacterial Photosynthesis ◽  
Core Complex ◽  
Rc Structures ◽  
Complex Forms ◽  
And Function

AbstractRhodobacter (Rba.) sphaeroides is the most widely used model organism in bacterial photosynthesis. The light-harvesting-reaction center (LH1-RC) core complex of this purple phototroph is characterized by the co-existence of monomeric and dimeric forms, the presence of the protein PufX, and approximately two carotenoids per LH1 αβ-polypeptides. Despite many efforts, structures of the Rba. sphaeroides LH1-RC have not been obtained at high resolutions. Here we report a cryo-EM structure of the monomeric LH1-RC from Rba. sphaeroides strain IL106 at 2.9 Å resolution. The LH1 complex forms a C-shaped structure composed of 14 αβ-polypeptides around the RC with a large ring opening. From the cryo-EM density map, a previously unrecognized integral membrane protein, referred to as protein-U, was identified. Protein-U has a U-shaped conformation near the LH1-ring opening and was annotated as a hypothetical protein in the Rba. sphaeroides genome. Deletion of protein-U resulted in a mutant strain that expressed a much-reduced amount of the dimeric LH1-RC, indicating an important role for protein-U in dimerization of the LH1-RC complex. PufX was located opposite protein-U on the LH1-ring opening, and both its position and conformation differed from that of previous reports of dimeric LH1-RC structures obtained at low-resolution. Twenty-six molecules of the carotenoid spheroidene arranged in two distinct configurations were resolved in the Rba. sphaeroides LH1 and were positioned within the complex to block its channels. Our findings offer an exciting new view of the core photocomplex of Rba. sphaeroides and the connections between structure and function in bacterial photocomplexes in general.

scholarly journals A Novel Membrane Protein in the Rhodobacter sphaeroides LH1-RC Photocomplex

2021 ◽  
Author(s):  
K. Tani ◽  
K. V. P. Nagashima ◽  
R. Kanno ◽  
S. Kawamura ◽  
R. Kikuchi ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Ring Opening ◽  
Hypothetical Protein ◽  
Integral Membrane Protein ◽  
Purple Bacterium ◽  
Core Complex ◽  
Rc Structures ◽  
Density Map ◽  
Complex Forms ◽  
And Function

We present a cryo-EM structure of the monomeric light-harvesting-reaction center (LH1-RC) core complex from photosynthetic purple bacterium Rhodobacter (Rba.) sphaeroides at 2.9 Å resolution. The LH1 complex forms a C-shaped structure composed of 14 αβ-polypeptides around the RC with a large ring opening. From the cryo-EM density map, a previously unrecognized integral membrane protein, referred to as protein-U, was identified. Protein-U has a U-shaped conformation near the LH1-ring opening and was annotated as a hypothetical protein in the Rba. sphaeroides genome. Deletion of protein-U resulted in a mutant strain that expressed a much-reduced amount of the dimeric LH1-RC, indicating an important role for protein-U in dimerization of the LH1-RC complex. PufX was located opposite protein-U on the LH1-ring opening, and both its position and conformation differed from that of previous reports of dimeric LH1-RC structures obtained at low-resolution. Twenty-six molecules of the carotenoid spheroidene arranged in two distinct configurations were resolved in the Rba. sphaeroides LH1 and were positioned within the complex to block its pores. Our findings offer a new view of the core photocomplex of Rba. sphaeroides and the connections between structure and function in bacterial photocomplexes in general.

scholarly journals Recruitment of Atg9 to the preautophagosomal structure by Atg11 is essential for selective autophagy in budding yeast

2006 ◽  
Vol 175 (6) ◽  
pp. 925-935 ◽  
Author(s):  
Congcong He ◽  
Hui Song ◽  
Tomohiro Yorimitsu ◽  
Iryna Monastyrska ◽  
Wei-Lien Yen ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Actin Cytoskeleton ◽  
Integral Membrane Protein ◽  
Yeast Two Hybrid ◽  
Anterograde Transport ◽  
Yeast Saccharomyces Cerevisiae ◽  
Selective Autophagy ◽  
Peripheral Membrane Protein ◽  
Developmental Conditions ◽  
Two Hybrid

Autophagy is a conserved degradative pathway that is induced in response to various stress and developmental conditions in eukaryotic cells. It allows the elimination of cytosolic proteins and organelles in the lysosome/vacuole. In the yeast Saccharomyces cerevisiae, the integral membrane protein Atg9 (autophagy-related protein 9) cycles between mitochondria and the preautophagosomal structure (PAS), the nucleating site for formation of the sequestering vesicle, suggesting a role in supplying membrane for vesicle formation and/or expansion during autophagy. To better understand the mechanisms involved in Atg9 cycling, we performed a yeast two-hybrid–based screen and identified a peripheral membrane protein, Atg11, that interacts with Atg9. We show that Atg11 governs Atg9 cycling through the PAS during specific autophagy. We also demonstrate that the integrity of the actin cytoskeleton is essential for correct targeting of Atg11 to the PAS. We propose that a pool of Atg11 mediates the anterograde transport of Atg9 to the PAS that is dependent on the actin cytoskeleton during yeast vegetative growth.

scholarly journals The yeast integral membrane protein Apq12 potentially links membrane dynamics to assembly of nuclear pore complexes

2007 ◽  
Vol 178 (5) ◽  
pp. 799-812 ◽  
Author(s):  
John J. Scarcelli ◽  
Christine A. Hodge ◽  
Charles N. Cole
Keyword(s):  
Membrane Protein ◽  
Integral Membrane Protein ◽  
Membrane Dynamics ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Cold Sensitive ◽  
Low Levels ◽  
Lower Temperature ◽  
Pore Complexes ◽  
And Function

Although the structure and function of components of the nuclear pore complex (NPC) have been the focus of many studies, relatively little is known about NPC biogenesis. In this study, we report that Apq12 is required for efficient NPC biogenesis in Saccharomyces cerevisiae. Apq12 is an integral membrane protein of the nuclear envelope (NE) and endoplasmic reticulum. Cells lacking Apq12 are cold sensitive for growth, and a subset of their nucleoporins (Nups), those that are primarily components of the cytoplasmic fibrils of the NPC, mislocalize to the cytoplasm. APQ12 deletion also causes defects in NE morphology. In the absence of Apq12, most NPCs appear to be associated with the inner but not the outer nuclear membrane. Low levels of benzyl alcohol, which increases membrane fluidity, prevented Nup mislocalization and restored the proper localization of Nups that had accumulated in cytoplasmic foci upon a shift to lower temperature. Thus, Apq12p connects nuclear pore biogenesis to the dynamics of the NE.

Topology and function of the integral membrane protein conferring immunity to colicin A

1989 ◽  
Vol 3 (5) ◽  
pp. 679-687 ◽  
Author(s):  
V. Geli ◽  
D. Baty ◽  
F. Pattus ◽  
C. Lazdunski
Keyword(s):  
Membrane Protein ◽  
Integral Membrane Protein ◽  
And Function

scholarly journals Differential expression and function of two homologous subunits of yeast 1,3-beta-D-glucan synthase.

1995 ◽  
Vol 15 (10) ◽  
pp. 5671-5681 ◽  
Author(s):  
P Mazur ◽  
N Morin ◽  
W Baginsky ◽  
M el-Sherbeini ◽  
J A Clemas ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Function Analysis ◽  
Calcineurin Inhibitors ◽  
Integral Membrane Protein ◽  
Dependent Manner ◽  
Fungal Cell ◽  
Glucan Synthase ◽  
Increased Sensitivity ◽  
And Function ◽  
Null Mutants

1,3-beta-D-Glucan is a major structural polymer of yeast and fungal cell walls and is synthesized from UDP-glucose by the multisubunit enzyme 1,3-beta-D-glucan synthase. Previous work has shown that the FKS1 gene encodes a 215-kDa integral membrane protein (Fks1p) which mediates sensitivity to the echinocandin class of antifungal glucan synthase inhibitors and is a subunit of this enzyme. We have cloned and sequenced FKS2, a homolog of FKS1 encoding a 217-kDa integral membrane protein (Fks2p) which is 88% identical to Fks1p. The residual glucan synthase activity present in strains with deletions of fks1 is (i) immunodepleted by antibodies prepared against FKS2 peptides, demonstrating that Fks2p is also a component of the enzyme, and (ii) more sensitive to the echinocandin L-733,560, explaining the increased sensitivity of fks1 null mutants to this drug. Simultaneous disruption of FKS1 and FKS2 is lethal, suggesting that Fks1p and Fks2p are alternative subunits with essential overlapping function. Analysis of FKS1 and FKS2 expression reveals that transcription of FKS1 is regulated in the cell cycle and predominates during growth on glucose, while FKS2 is expressed in the absence of glucose. FKS2 is essential for sporulation, a process which occurs during nutritional starvation. FKS2 is induced by the addition of Ca2+ to the growth medium, and this induction is completely dependent on the Ca2+/calmodulin-dependent phosphoprotein phosphatase calcineurin. We have previously shown that growth of fks1 null mutants is highly sensitive to the calcineurin inhibitors FK506 and cyclosporin A. Expression of FKS2 from the heterologous ADH1 promoter results in FK506-resistant growth. Thus, the sensitivity of fks1 mutants to these drugs can be explained by the calcineurin-dependent transcription of FKS2. Moreover, FKS2 is also highly induced in response to pheromone in a calcineurin-dependent manner, suggesting that FKS2 may also play a role in the remodeling of the cell wall during the mating process.

Structure and function of integral membrane protein domains resolved by peptide-amphiphiles: Application to phospholamban

Biopolymers ◽  
2003 ◽  
Vol 69 (3) ◽  
pp. 283-292 ◽  
Author(s):  
Nathan A. Lockwood ◽  
Raymond S. Tu ◽  
Zhiwen Zhang ◽  
Matthew V. Tirrell ◽  
David D. Thomas ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Protein Domains ◽  
Integral Membrane Protein ◽  
Structure And Function ◽  
Peptide Amphiphiles ◽  
And Function

scholarly journals The Integral Membrane Protein Snl1p Is Genetically Linked to Yeast Nuclear Pore Complex Function

1998 ◽  
Vol 9 (2) ◽  
pp. 355-373 ◽  
Author(s):  
Albert K. Ho ◽  
Gregory A. Raczniak ◽  
Eric B. Ives ◽  
Susan R. Wente
Keyword(s):  
Membrane Protein ◽  
Transmembrane Domain ◽  
Integral Membrane Protein ◽  
Genetic Screen ◽  
Terminal Region ◽  
Yeast Cells ◽  
Nuclear Pore ◽  
Endoplasmic Reticulum Membrane ◽  
Carboxyl Terminal ◽  
And Function

Integral membrane proteins are predicted to play key roles in the biogenesis and function of nuclear pore complexes (NPCs). Revealing how the transport apparatus is assembled will be critical for understanding the mechanism of nucleocytoplasmic transport. We observed that expression of the carboxyl-terminal 200 amino acids of the nucleoporin Nup116p had no effect on wild-type yeast cells, but it rendered thenup116 null strain inviable at all temperatures and coincidentally resulted in the formation of nuclear membrane herniations at 23°C. To identify factors related to NPC function, a genetic screen for high-copy suppressors of this lethalnup116-C phenotype was conducted. One gene (designatedSNL1 for suppressor of n up116-C lethal) was identified whose expression was necessary and sufficient for rescuing growth. Snl1p has a predicted molecular mass of 18.3 kDa, a putative transmembrane domain, and limited sequence similarity to Pom152p, the only previously identified yeast NPC-associated integral membrane protein. By both indirect immunofluorescence microscopy and subcellular fractionation studies, Snl1p was localized to both the nuclear envelope and the endoplasmic reticulum. Membrane extraction and topology assays suggested that Snl1p was an integral membrane protein, with its carboxyl-terminal region exposed to the cytosol. With regard to genetic specificity, the nup116-C lethality was also suppressed by high-copy GLE2 and NIC96. Moreover, high-copy SNL1 suppressed the temperature sensitivity ofgle2–1 and nic96-G3 mutant cells. Thenic96-G3 allele was identified in a synthetic lethal genetic screen with a null allele of the closely related nucleoporinnup100. Gle2p physically associated with Nup116p in vitro, and the interaction required the N-terminal region of Nup116p. Therefore, genetic links between the role of Snl1p and at least three NPC-associated proteins were established. We suggest that Snl1p plays a stabilizing role in NPC structure and function.

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