Carbohydrate modifications in the Golgi function in a variety of biological roles ranging from the stabilization of protein structure to the regulation of cell surface properties. The overall goal of this research is to understand how glycosylation is regulated and how carbohydrate modifications mediate their biological roles at the cell surface. Glycosyl transferases catalyze a set of ubiquitous reactions that contribute to and are affected by the ionic equilibrium of the Golgi lumen. As a byproduct of glycosylation, vast quantities of inorganic phosphate are generated. This phosphate must be removed to prevent an overly acidic lumen that would otherwise inhibit the activity of the resident enzymes.
Our first aim i s to determine how this phosphate is removed from the Golgi, using S. cerevisiae as an experimental model system. Our past studies identified ERD1 as an important, highly conserved regulator of phosphate efflux from the Golgi. To better understand how ERD1 regulates the lumenal environment, experiments are proposed to generate new mutants and analyze mutants already identified that interact with ERD1 or that are similarly defective in regulating the lumenal environment of the Golgi. In a complementary approach we will apply biochemical assays to identify and characterize the Gotgi phosphate transporter.
Our second aim i s to determine how the byproducts of sugar consumption contribute to other metabolic pathways in yeast, specifically phosphate homeostasis. To achieve this aim, we will apply sensitive biochemical assays to directly measure phosphate production from glycosylation. The proposed experiments will reveal important information about the establishment and maintenance of an optimal Golgi environment and how the reactions that occur in the Golgi intersect with other pathways.
Our third aim i s to examine the role of the Golgi and the glycans themselves in contributing to the cell surface properties of Candida albicans, the most frequently isolated human fungat pathogen. Our past studies demonstrated that C. albicans has evolved an alternate strategy for establishing polarity during hyphal growth, in which the entire Golgi complex redistributes sub-apicality during filamentation. Specifically, we will use cell biological and molecular techniques to study the regulation and cytoskeletal requirements of Golgi movement to the hyphal tip.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM048467-12
Application #
7118902
Study Section
Microbial Physiology and Genetics Subcommittee 2 (MBC)
Program Officer
Shapiro, Bert I
Project Start
1995-09-01
Project End
2009-05-31
Budget Start
2006-09-01
Budget End
2009-05-31
Support Year
12
Fiscal Year
2006
Total Cost
$389,357
Indirect Cost
Name
State University New York Stony Brook
Department
Biochemistry
Type
Schools of Medicine
DUNS #
804878247
City
Stony Brook
State
NY
Country
United States
Zip Code
11794
Noffz, Christine; Keppler-Ross, Sabine; Dean, Neta (2009) Hetero-oligomeric interactions between early glycosyltransferases of the dolichol cycle. Glycobiology 19:472-8
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Keppler-Ross, Sabine; Noffz, Christine; Dean, Neta (2008) A new purple fluorescent color marker for genetic studies in Saccharomyces cerevisiae and Candida albicans. Genetics 179:705-10
Averbeck, Nicole; Gao, Xiao-Dong; Nishimura, Shin-Ichiro et al. (2008) Alg13p, the catalytic subunit of the endoplasmic reticulum UDP-GlcNAc glycosyltransferase, is a target for proteasomal degradation. Mol Biol Cell 19:2169-78
Averbeck, Nicole; Keppler-Ross, Sabine; Dean, Neta (2007) Membrane topology of the Alg14 endoplasmic reticulum UDP-GlcNAc transferase subunit. J Biol Chem 282:29081-8
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Gao, Xiao-Dong; Wang, Ji; Keppler-Ross, Sabine et al. (2005) ERS1 encodes a functional homologue of the human lysosomal cystine transporter. FEBS J 272:2497-511
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Nishikawa, Akiko; Poster, Jay B; Jigami, Yoshifumi et al. (2002) Molecular and phenotypic analysis of CaVRG4, encoding an essential Golgi apparatus GDP-mannose transporter. J Bacteriol 184:29-42
Cipollo, J F; Trimble, R B; Chi, J H et al. (2001) The yeast ALG11 gene specifies addition of the terminal alpha 1,2-Man to the Man5GlcNAc2-PP-dolichol N-glycosylation intermediate formed on the cytosolic side of the endoplasmic reticulum. J Biol Chem 276:21828-40

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