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Investigations of Functional Interactions Between Env7 and TOR Complex 1 in Saccharomyces cerevisae
Author(s) -
Shaoulian Jonathan,
Gharakhanian Editte
Publication year - 2016
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.30.1_supplement.860.2
Subject(s) - vacuole , autophagy , microbiology and biotechnology , endosome , biology , saccharomyces cerevisiae , fragmentation (computing) , kinase , fusion protein , protein kinase a , lipid bilayer fusion , chemistry , biochemistry , gene , cytoplasm , membrane , intracellular , apoptosis , recombinant dna , ecology
In eukaryotic cells, deficiencies in proteins involved in lysosomal trafficking and function result in serious human disorders like Tay‐Sach's Disease or Alzheimer's. Our lab has identified ENV7, a novel gene involved in the late‐endosome‐to vacuole interface in Saccharomyces cerevisae . ENV7, (a homolog of human STK16 ) encodes Env7, a vacuolar membrane kinase that negatively regulates vacuolar fusion. Tor1 is another vacuolar membrane kinase, and has been identified as a vacuolar fragmentation factor. Activated Tor Complex 1 (TORC1) stimulates vacuolar fragmentation, as well as cell growth and proliferation. Tor1 and TORC1 are also evolutionarily conserved, and in mammals, hyperactivity of TORC1 plays a major role in cancers. Furthermore, inhibition of TORC1 is associated with the initiation of autophagy, which itself involves a number of fusion events at the vacuole. Because of this, we hypothesized that there is a functional interface between the vacuolar membrane kinases Env7 and Tor1 in TORC1. Here, using confocal microscopy techniques, we report that while the absence of TOR1 is not sufficient to affect the localization of exogenously expressed GFP‐Env7, when TORC1 is inhibited by rapamycin in the same cells, there is a significant mislocalization of the GFP‐Env7 fusion protein to the plasma membrane. Cells of the tor1 Δ strain treated by rapamycin also display an enlarged vacuole morphology. Additionally, this enlarged vacuole morphology in tor1 Δ seems to be associated with slightly increased levels of autophagy. Therefore, the results thus far suggest a functional interaction between Env7 and TORC1. We are currently investigating the localization of native levels of Env7‐GFP when TORC1 is inhibited by rapamycin. We are also further exploring the TORC1 and Env7 interface by studying additional deletion mutants of TORC1 components. Support or Funding Information This project is supported by funding from the NIH (grant SC1 GM112560‐01 to Editte Gharakhanian)