Systematic analysis of membrane contact sites in Saccharomyces cerevisiae uncovers modulators of cellular lipid 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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ety levels and cognitive defi cits) were most pronounced in elderly subjects. This study is the fi rst to show any dysfunction in healthy CJD mutation carriers. Scientifi c Background Creutzfeldt-Jacob Disease (CJD) is a fatal neurodegenerative disorder, the most prevalent of the human prion diseases. CJD often has a sudden onset and it is ultimately terminal, usually within a few months The pathogenesis of CJD is believed to relate to the accumulation of an abnormal prion protein isoform (PrP Sc ) of the normally occurring cellular protein (PrP c ) in the central nervous system Key Words Creutzfeldt-Jacob disease ؒ Anxiety ؒ Dementia ؒ Preclinical signs ؒ PRNP gene ؒ Neuropsychology ؒ Prion mutation Abstract Creutzfeldt-Jacob disease (CJD) is a rapidly progressing dementia with neurological, psychiatric and cognitive symptoms. We focused our study on the familial CJD form among Libyan Jews (the E200K mutation), trying to identify preclinical neuropsychological signs in mutation carriers to facilitate early diagnosis of the disease. A wide range of neuropsychological tests was administered to 27 healthy volunteers, all fi rst-degree relatives of genetic CJD patients. Thirteen of our participants were gene mutation carriers (E200K) and 14 controls. The healthy mutation carriers reported signifi cantly lower Trait and higher State anxiety scores. Repeated Measure analysis showed statistical signifi cance. The Anxiety Index (State-Trait Anxiety Score) progressed with age in the carriers' group but not in the controls. Since this was more pronounced in the older subjects, we suggest that abnormal stress mechanisms precede the clinical onset of CJD. Cognitive differences have also been found between carriers and controls, especially in visual recognition of pictured objects. Both kinds of differences (anxi