Clathrin-mediated vesicular transport is important for normal growth and development of all eukaryotes. Clathrin is required for endocytosis of many nutrients and signal transducing receptors, sorting from the late Golgi to the lysosome, and maturation of regulated secretory granules. Studies in Dr. Lemmon's lab have taken advantage of the powerful molecular genetic methods available in yeast to analyze the function of clathrin. Previous studies have shown that disruption of the clathrin heavy chain gene (CHC1) is lethal in some yeast strains. This observation has permitted the identification of five multicopy suppressors (SCD) genes whose overexpression suppresses the lethality of clathrin-deficient yeast. The SCD genes encode components of the vesicular transport machinery, novel proteins, and known proteins that unexpectedly rescue cells lacking clathrin. The latter have suggested relationships between processes not previously predicted and have provided novel insights into the global effects of clatherin deficiency. This application proposes studies on the SCD5 and SCD3 genes. SCD5 encodes a novel protein with a post Golgi secretory function. Recent evidence suggests SCD5 may be a targeting subunit for type 1 protein phosphatase (PP1), a broad specificity phosphatase with diverse cellular functions. This indicates SCD5 may be involved in regulating a specific phosphorylation event in the late secretory pathway. A combination of genetic, cellular and biochemical approaches will be used to further define the function of SCD5, especially in relation to PP1. SCD3 encodes a member of the 14-3-3 protein family. The 14-3-3 proteins are widely expressed in eukaryotes and appear to function in multiple cellular processes. Dr. Lemmon's genetic analysis in yeast has provided evidence that 14-3-3 proteins may play a role in vesicular transport of regulate a process whose function is affected by alterations in vesicular transport. She will test whether 14-3-3 proteins directly influence membrane sorting and use genetic approaches to identify the players in the membrane associated pathways that require 14-3-3 function. The studies outlined in this proposal will provide insights into the function of SCD5, PP1, and 14-3-3 proteins. This will lead to a fuller understanding of the role of the SCD proteins in vesicular transport and their relationship to clathrin function.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM055796-03
Application #
6180726
Study Section
Microbial Physiology and Genetics Subcommittee 2 (MBC)
Program Officer
Shapiro, Bert I
Project Start
1998-04-01
Project End
2002-03-31
Budget Start
2000-04-01
Budget End
2001-03-31
Support Year
3
Fiscal Year
2000
Total Cost
$245,638
Indirect Cost
Name
Case Western Reserve University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
077758407
City
Cleveland
State
OH
Country
United States
Zip Code
44106
Ding, Jingzhen; Segarra, Verónica A; Chen, Shuliang et al. (2016) Auxilin facilitates membrane traffic in the early secretory pathway. Mol Biol Cell 27:127-36
Boettner, Douglas R; Segarra, Verónica A; Moorthy, Balaji T et al. (2016) Creating a chimeric clathrin heavy chain that functions independently of yeast clathrin light chain. Traffic 17:754-68
Bahnan, Wael; Boettner, Douglas R; Westermark, Linda et al. (2015) Pathogenic Yersinia Promotes Its Survival by Creating an Acidic Fluid-Accessible Compartment on the Macrophage Surface. PLoS One 10:e0133298
Segarra, Verónica A; Boettner, Douglas R; Lemmon, Sandra K (2015) Atg27 tyrosine sorting motif is important for its trafficking and Atg9 localization. Traffic 16:365-78
Mukherjee, Debarati; Sen, Arpita; Boettner, Douglas R et al. (2013) Bem3, a Cdc42 GTPase-activating protein, traffics to an intracellular compartment and recruits the secretory Rab GTPase Sec4 to endomembranes. J Cell Sci 126:4560-71
Chi, Richard J; Torres, Onaidy T; Segarra, Veronica A et al. (2012) Role of Scd5, a protein phosphatase-1 targeting protein, in phosphoregulation of Sla1 during endocytosis. J Cell Sci 125:4728-39
Lemmon, Sandra K; Traub, Linton M (2012) Getting in touch with the clathrin terminal domain. Traffic 13:511-9
Boettner, Douglas R; Friesen, Helena; Andrews, Brenda et al. (2011) Clathrin light chain directs endocytosis by influencing the binding of the yeast Hip1R homologue, Sla2, to F-actin. Mol Biol Cell 22:3699-714
Boettner, Douglas R; Chi, Richard J; Lemmon, Sandra K (2011) Lessons from yeast for clathrin-mediated endocytosis. Nat Cell Biol 14:2-10
Grotsch, Helga; Giblin, Jonathan P; Idrissi, Fatima-Zahra et al. (2010) Calmodulin dissociation regulates Myo5 recruitment and function at endocytic sites. EMBO J 29:2899-914

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