Burston

Thursday, May 26, 2011, 12:30 pm. Room 200, Graduate Student Centre

“Genome-wide Analysis of Endocytic Recycling in S. cerevisiae”

 

EXAMINING COMMITTEE

Chair:
    Dr. Artem Cherkasov (Experimental Medicine)

Supervisory Committee:
    Dr. Elizabeth Conibear, Research Supervisor (Medical Genetics)
    Dr. Phil Hieter (Medical Genetics)
    
University Examiners:
    Dr. Lacey Samuels (Botany)
    Dr. Masayuki Numata (Biochemistry and Molecular Biology)

External Examiner:
    Dr. Gary Eitzen
    Department of Cell Biology
    University of Alberta
    Edmonton, Alberta
    Canada


SUPERVISORY COMMITTEE
    Dr. Elizabeth Conibear (Supervisor),
    Dr. Phil Hieter,
    Dr. Pamela Hoodless,
    Dr. Michael Kobor



ABSTRACT
The process of endocytic recycling, in which cell surface proteins are internalized and re-delivered to the plasma membrane, is essential in all eukaryotes for maintaining plasma membrane composition, and regulating the surface levels of signaling receptors.  The applicability of yeast as a model to study endocytic recycling is a subject of debate, as there appears to be critical differences in yeast and mammalian cells.  For example, while clathrin and its adaptors are critical for uptake in mammals, they do not seem to be essential in yeast.  Endocytic recycling has not been comprehensively studied on a genetic level in yeast, however, and only limited cargo have been considered, making it difficult to accurately assess the similarity between the two systems.  Furthermore, the transport of SNARE proteins is poorly understood, but appears to involve specialized mechanisms.  In this study, a genome-wide screening approach was used to systematically and quantitatively identify genes required for the endocytic recycling of the yeast SNARE protein Snc1, which is homologous to the mammalian VAMP2/synaptobrevin.
Endocytic defects for mutants of many yeast homologs of mammalian endocytosis genes were identified, for the first time.  Significantly, a cargo-selective and partially-redundant role for clathrin and its adaptors yAP1801 and yAP1802 was identified.  The lipid phosphatase Inp52 was found to mediate AP180 release from endocytic vesicles.  Furthermore, the previously uncharacterized protein Ldb17, homologous to the mammalian endocytic protein SPIN90, was identified as a new component of the endocytic machinery, and regulates both coat and actin dynamics at endocytic sites.
Factors regulating Snc1 recycling were also identified, including the variant clathrin adaptor AP-1R.  This is the first reported function for this complex.  The previously uncharacterized protein Ima1 was found to be a putative enzyme that specifically binds to AP-1R, and may have activity related to AP-1R function.
Overall, this study demonstrates that endocytic recycling in yeast and mammals is more similar than previously appreciated, and identifies new factors in this process.  Furthermore, it raises awareness of the degree of cargo-selectivity underlying this pathway, and demonstrates quantitative methods that can be further applied to future studies in both systems.



PUBLICATIONS
Burston HE, Maldonado-Báez L, Davey M, Montpetit B, Schluter C, Wendland B, and Conibear E. (2009). Regulators of Yeast Endocytosis Identified by Systematic Quantitative Analysis. Journal of Cell Biology. 185(6):1097-110.

Burston HE, Davey M, Conibear E. (2008). Genome-wide Analysis of Membrane Transport Using Yeast Knockout Arrays. Methods in Molecular Biology, vol. Membrane Trafficking, ed. A. Vancura: Humana Press. 457:29-39.



ORAL PRESENTATIONS
Burston HE, Maldonaldo-Baez L, Montpetit B, Davey M, Schluter C, Wendland B, and Conibear E. (2008). Genome-wide analysis of Snc1 endocytosis. 2008 Yeast Genetics and Molecular Biology Meeting, University of Toronto, Ontario, July 2008. Invited Platform Presentation

Burston HE, Maldonaldo-Baez L, Montpetit B, Davey M, Schluter C, Wendland B, and Conibear E. (2008). Genome-wide analysis of Snc1 endocytosis. Cambridge Institute for Medical Research, University of Cambridge, Cambridge, UK, July 2008. Invited Oral Presentation.


POSTER PRESENTATIONS
Burston HE, Davey M, Raghuram N, and Conibear E. (2010). Regulation of Snc1 Recycling by the Alternate AP-1R Clathrin Adaptor and Ima1. 2010 Yeast Genetics and Molecular Biology Meeting, University of British Columbia, Vancouver BC, August 2010.

Burston HE, Maldonaldo-Baez L, Montpetit B, Davey M, Schluter C, Wendland B, and Conibear E. (2008). Genome-wide analysis of Snc1 endocytosis. 2009 Excitatory Synapses & Brain Function, Gordon Research Conference.  Les Diablerets, Switzerland. September 2009.



BIOGRAPHICAL NOTES
Academic Studies: B.Sc. Simon Fraser University, 2005. Molecular Biology and Biochemistry


GRADUATE STUDIES
Field of Study: Molecular genetic characterization of intracellular transport
Courses:
MEDG 419: Human Cytogenetics. Dr. Wendy Robinson
MEDG 505: Genome Analysis. Dr. Phil Hieter
MEDG 520: Advanced Human Molecular Genetics. Dr. Lorne Clarke
MEDG 530: Human Genetics. Dr. Jan Friedman
MEDG 545: Current Topics in Medical Genetics Research. Dr. Blair Leavitt
MEDG 548: Directed Studies. Dr. Elizabeth Conibear


AWARDS
Postgraduate Scholarship (PGS-D) Natural Sciences and Engineering Research Council of Canada (NSERC)
Research Trainee Award Michael Smith Foundation for Health Research (MSFHR)
Canada Graduate Scholarship (CGS-M) Natural Sciences and Engineering Research Council of Canada (NSERC)