A Modulatory Role for Clathrin Light Chain Phosphorylation in Golgi Membrane Protein Localization during Vegetative Growth and during the Mating Response of<i>Saccharomyces cerevisiae</i>

Chu, Diana S.; Pishvaee, Babak; Payne, Grégory S.

doi:10.1091/mbc.10.3.713

Cited by 13 publications

(14 citation statements)

References 67 publications

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“…Moreover, the assembly properties of pure clathrin (36,41) or of clathrin with AP complexes (42) in vitro are not affected in a significant way by the presence or absence of light chains, although a small inhibitory effect of the light chains has been found with pure clathrin (41). The light chains are thus likely to have regulatory functions in vivo that depend on recruitment of other factors or that affect the trimerization of the triskelion (43)(44)(45)(46).…”

Section: Overall Structure and Molecular Organizationmentioning

confidence: 99%

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Clathrin

Kirchhausen

2000

Annu. Rev. Biochem.

559

512

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Clathrin was discovered nearly 25 years ago. Since then, a large number of other proteins that participate in the process by which clathrin-coated vesicles retrieve synaptic membranes or take up endocytic receptors have been identified. The functional relationships among these disparate components remain, in many cases, obscure. High-resolution structures of parts of clathrin, determined by X-ray crystallography, and lower-resolution images of assembled coats, determined by electron cryomicroscopy, now provide the information necessary to integrate various lines of evidence and to design experiments that test specific mechanistic notions. This review summarizes and illustrates the recent structural results and outlines what is known about coated-vesicle assembly in the context of this information.

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Section: Overall Structure and Molecular Organizationmentioning

confidence: 99%

“…Yeasts express only one light chain (43,44); C. elegans apparently also has only one. In mammalian neurons, there is a further level of complexity caused by differential splicing of the LCa and LCb genes (57,58).…”

Section: Light-chain Sequencesmentioning

confidence: 99%

Clathrin

Kirchhausen

2000

Annu. Rev. Biochem.

559

512

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“…Cells with mutations in clathrin subunits secrete a highly glycosylated precursor form of the ␣-factor mating pheromone (Payne and Schekman, 1989;Seeger and Payne, 1992b;Chu et al, 1996Chu et al, , 1999Huang et al, 1997). This defect is attributed to mislocalization of the Golgi membrane protein Kex2p, which normally initiates proteolytic maturation of the pheromone precursor in the TGN (Fuller et al, 1988).…”

Section: Disruption Of Ap-1 Enhances Effects Of Chc1-tsmentioning

confidence: 99%

Adaptor Complex-independent Clathrin Function in Yeast

Yeung¹,

Phan

Payne

1999

MBoC

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152

166

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Clathrin-associated adaptor protein (AP) complexes are major structural components of clathrin-coated vesicles, functioning in clathrin coat assembly and cargo selection. We have carried out a systematic biochemical and genetic characterization of AP complexes in Saccharomyces cerevisiae. Using coimmunoprecipitation, the subunit composition of two complexes, AP-1 and AP-2R, has been defined. These results allow assignment of the 13 potential AP subunits encoded in the yeast genome to three AP complexes. As assessed by in vitro binding assays and coimmunoprecipitation, only AP-1 interacts with clathrin. Individual or combined disruption of AP-1 subunit genes in cells expressing a temperature-sensitive clathrin heavy chain results in accentuated growth and alpha-factor pheromone maturation defects, providing further evidence that AP-1 is a clathrin adaptor complex. However, in cells expressing wild-type clathrin, the same AP subunit deletions have no effect on growth or alpha-factor maturation. Furthermore, gel filtration chromatography revealed normal elution patterns of clathrin-coated vesicles in cells lacking AP-1. Similarly, combined deletion of genes encoding the beta subunits of the three AP complexes did not produce defects in clathrin-dependent sorting in the endocytic and vacuolar pathways or alterations in gel filtration profiles of clathrin-coated vesicles. We conclude that AP complexes are dispensable for clathrin function in S. cerevisiae under normal conditions. Our results suggest that alternative factors assume key roles in stimulating clathrin coat assembly and cargo selection during clathrin-mediated vesicle formation in yeast.

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“…Aliquots were removed at 0, 10, and 30 min after the addition of unlabeled amino acids. Kex2p was metabolically labeled and immunoprecipitated as described by Chu et al (1999). Vps10p and alkaline phosphatase were labeled and immunoprecipitated using the same protocol as Kex2p except samples were removed at chase times indicated in the figure legend.…”

Section: Metabolic Labeling and Immunoprecipitationmentioning

confidence: 99%

Ric1p and the Ypt6p GTPase Function in a Common Pathway Required for Localization ofTrans-Golgi Network Membrane Proteins

Bensen

Yeung

Payne

2001

MBoC

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In Saccharomyces cerevisiae, clathrin is necessary for localization of trans-Golgi network (TGN) membrane proteins, a process that involves cycling of TGN proteins between the TGN and endosomes. To characterize further TGN protein localization, we applied a screen for mutations that cause severe growth defects in combination with a temperature-sensitive clathrin heavy chain. This screen yielded a mutant allele of RIC1. Cells carrying a deletion of RIC1 (ric1⌬) mislocalize TGN membrane proteins Kex2p and Vps10p to the vacuole. Delivery to the vacuole occurs in ric1⌬ cells also harboring end3⌬ to block endocytosis, indicative of a defect in retrieval to the TGN rather than sorting to endosomes. SYS1, originally discovered as a multicopy suppressor of defects caused by the absence of the Rab GTPase YPT6, was identified as a multicopy suppressor of ric1⌬. Further comparison of ric1⌬ and ypt6⌬ cells demonstrated identical phenotypes. Multicopy plasmids expressing v-SNAREs Gos1p or Ykt6p, but not other v-and t-SNAREs, partially suppressed phenotypes of ric1⌬ and ypt6⌬ cells. SLY1-20, a dominant activator of the cis-Golgi network t-SNARE Sed5p, also functioned as a multicopy suppressor. Because Gos1p and Ykt6p interact with Sed5p, these results raise the possibility that TGN membrane protein localization requires Ric1p-and Ypt6p-dependent retrieval to the cis-Golgi network. INTRODUCTIONLocalization of proteins to appropriate membrane organelles is crucial for the functional compartmentalization of eukaryotic cells. For proteins that function in organelles of the secretory and endocytic pathways, localization requires not only targeting to the proper destination but mechanisms to maintain residence despite extensive membrane and protein flux through each organelle. Continued residence can be achieved through retention mechanisms that restrict incorporation into transport vesicles departing from an organelle and/or retrieval mechanisms that carry out vesicle-mediated return from distal sites in the pathway (Pelham and Munro, 1993;Rothman and Wieland, 1996).The Golgi apparatus in the yeast Saccharomyces cerevisiae, like its mammalian counterpart, is organized into dynamic, functionally distinct subcompartments that pose additional challenges for protein localization. Though not arranged into the cisternal stacks characteristic of the mammalian cell Golgi apparatus, yeast Golgi subcompartments can be considered functionally analogous to the mammalian cis-Golgi network (CGN), medial Golgi, and trans-Golgi (TGN) network (Graham and Emr, 1991;Preuss et al., 1992). The CGN serves as the site where endoplasmic reticulum-derived transport carriers dock and fuse and where mannose residues are first added to the core oligosaccharides of glycoproteins (Gaynor et al., 1994;Graham and Emr, 1991). Accordingly, this compartment is enriched for the t-SNARE Sed5p involved in the fusion of ER transport carriers and in the ␣-1,6 mannosyltransferase Och1p (Gaynor et al., 1994;Hardwick and Pelham, 1992). The medial Golgi compartment carries ...

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A Modulatory Role for Clathrin Light Chain Phosphorylation in Golgi Membrane Protein Localization during Vegetative Growth and during the Mating Response ofSaccharomyces cerevisiae

Cited by 13 publications

References 67 publications

Clathrin

Clathrin

Adaptor Complex-independent Clathrin Function in Yeast

Ric1p and the Ypt6p GTPase Function in a Common Pathway Required for Localization ofTrans-Golgi Network Membrane Proteins

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