Journal
ELIFE
Volume 11, Issue -, Pages -Publisher
eLIFE SCIENCES PUBL LTD
DOI: 10.7554/eLife.74602
Keywords
contact sites; YPR097W; vps13 family; lipid droplets; lec1; S; cerevisiae
Categories
Funding
- European Molecular Biology Organization [ALTF-580-2017]
- Volkswagen Foundation [93092]
- Deutsche Forschungsgemeinschaft [SFB1190, MC_UU_00012/6]
- Biotechnology and Biological Sciences Research Council [BB/M011801/1]
- Canada Foundation for Innovation [30636]
- Canadian Institutes of Health Research [148756]
- Canadian Institutes of Health Research
- Natural Sciences and Engineering Research Council of Canada
- BC Children's Hospital
- University of British Columbia
- Natural Sciences and Engineering Research Council of Canada [PIN303585-2016]
- European Commission [ERC-CoG 866011]
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This study systematically characterized the proteomes of contact sites in Saccharomyces cerevisiae using co-localization imaging, identifying over 100 new potential residents and effectors. Several newly identified residents were found to play a role in the regulation of ergosterol distribution.
Actively maintained close appositions between organelle membranes, also known as contact sites, enable the efficient transfer of biomolecules between cellular compartments. Several such sites have been described as well as their tethering machineries. Despite these advances we are still far from a comprehensive understanding of the function and regulation of most contact sites. To systematically characterize contact site proteomes, we established a high-throughput screening approach in Saccharomyces cerevisiae based on co-localization imaging. We imaged split fluorescence reporters for six different contact sites, several of which are poorly characterized, on the background of 1165 strains expressing a mCherry-tagged yeast protein that has a cellular punctate distribution (a hallmark of contact sites), under regulation of the strong TEF2 promoter. By scoring both co-localization events and effects on reporter size and abundance, we discovered over 100 new potential contact site residents and effectors in yeast. Focusing on several of the newly identified residents, we identified three homologs of Vps13 and Atg2 that are residents of multiple contact sites. These proteins share their lipid transport domain, thus expanding this family of lipid transporters. Analysis of another candidate, Ypr097w, which we now call Lec1 (Lipid-droplet Ergosterol Cortex 1), revealed that this previously uncharacterized protein dynamically shifts between lipid droplets and the cell cortex, and plays a role in regulation of ergosterol distribution in the cell. Overall, our analysis expands the universe of contact site residents and effectors and creates a rich database to mine for new functions, tethers, and regulators.
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