Over the past several decades, many characterizations in vesicular trafficking system have led to great understanding of the endomembrane system, and several genetic screens have identified mutants defective in various protein sorting and trafficking functions within this system. Recently, a newly characterized group of mutants from our laboratory was reported to exhibit internal accumulation of the precursor carboxypeptidase (proCPY), at the post-endosome to vacuole trafficking stage, and named the late endosome and vacuole interface, or env mutants. The result from the screen has revealed a group of novel genes that have no previously established functions including ENV9. ENV9 is 993 base pairs in length, and encodes a protein of 330 amino acids (40 kDa) with homology to oxidoreductases. env9Δ mutant is caffeine sensitive, and exhibits an unusual vacuolar morphology, where accumulation of vesicles in the vacuole is observed. Env9 protein has also been reported to be localized to lipid droplets (LDs) from a systematic study. In this study, more research results were established on ENV9, its mutant, and its encoded protein. Further characterizations of the phenotypic traits of env9Δ mutant shows normal vacuolar acidification and autophagy function. The kinetics of CPY processing reveals a delay in maturation. Complementation of ENV9 in env9Δ mutant confirms that the deletion of the gene is responsible for the pleiotropic phenotype, while overexpression of the gene has no effects on vacuolar and LD morphologies, and caffeine sensitivity. Fluorescence microscopy on subcellular colocalization of GFP-tagged Env9p and Nile Red signal confirms that Env9p is localized to LDs. Biochemical assay using HA-tagged Env9p shows that it is membrane associated.
|Commitee:||Livingston, Brian, McAbee, Douglas|
|School:||California State University, Long Beach|
|School Location:||United States -- California|
|Source:||MAI 51/05M(E), Masters Abstracts International|
|Subjects:||Biology, Molecular biology, Cellular biology|
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