The vacuole in Saccharomyces cerevisiae (baker’s yeast) is functionally analogous to the human lysosome. Endosome and vacuolar interface 7 (Env7) has been recently identified in the Gharakhanian laboratory as a protein kinase that regulates vacuolar fusion and has a human ortholog, STK16. Previous research in our laboratory has shown that Env7 localizes to the vacuolar membrane in yeast. Proteins destined to function at the yeast vacuole are transported from the Golgi via two distinct pathways: the carboxypeptidase Y (CPY) pathway and the alkaline phosphatase (ALP) pathway. The CPY pathway requires an endosomal intermediate while the ALP pathway requires an adaptor protein complex (AP-3). The CPY pathway serves as the default pathway for the vast majority of proteins moving from the Golgi to the vacuole. Only 9 proteins are known to be transported via the ALP pathway in yeast, all of which are vacuolar membrane proteins and one of which is Yck3, a palmitoylated protein kinase with a similar function of Env7. Proteins normally transported to the yeast vacuole via the ALP pathway are able to partially revert to the CPY pathway when AP-3 function is disrupted. In the absence of a functional CPY pathway, however, the AP-3 pathway remains cargo-specific. Here we show that when the CPY pathway is disrupted in yeast cells, Env7 shows a significant increase in mislocalization. When the ALP pathway is disrupted, Env7 shows no significant mislocalization. Proteins transported via the ALP pathway interact with AP-3 via either an acidic-dileucine-like or a tyrosine-based adaptin sorting signal. Analysis of the amino acid sequence of Env7 showed one potential acidic-dileucine like signal and four potential tyrosine-based adaptin sorting signals. Mutagenesis of two of these sorting signals caused no significant mislocalization of Env7 in yeast. These results implicate the CPY pathway in the vacuolar trafficking of Env7 in yeast.
|Commitee:||Fraser, Deborah, Brusslan, Judith|
|School:||California State University, Long Beach|
|School Location:||United States -- California|
|Source:||MAI 81/1(E), Masters Abstracts International|
Copyright in each Dissertation and Thesis is retained by the author. All Rights Reserved
The supplemental file or files you are about to download were provided to ProQuest by the author as part of a
dissertation or thesis. The supplemental files are provided "AS IS" without warranty. ProQuest is not responsible for the
content, format or impact on the supplemental file(s) on our system. in some cases, the file type may be unknown or
may be a .exe file. We recommend caution as you open such files.
Copyright of the original materials contained in the supplemental file is retained by the author and your access to the
supplemental files is subject to the ProQuest Terms and Conditions of use.
Depending on the size of the file(s) you are downloading, the system may take some time to download them. Please be