The Hydrophobic Domain of ENV9 is a Lipid Droplet Localization Signal in Saccharomyces cerevisiae

Abstract

Lipid droplets (LDs) are dynamic organelles involved in the storage of neutral lipids for membrane biogenesis and metabolism. Malfunction of LDs has been associated with diabetes, obesity, and cancer, whereas proper organelle and cell function is mediated through cellular trafficking. Therefore, uncovering the mechanism by which proteins that are necessary for LD function are transported is essential for understanding the development of these diseases. We have found Saccharomyces cerevisiae Env9 to be a novel oxidoreductase that is conserved and involved in LD morphology. It contains a hydrophobic domain (HD) sequence (aa241‐265) that we have previously reported as necessary for Env9 localization to the LD membrane. For a sequence to be categorized to a specific pathway it needs to be demonstrated as both necessary and sufficient. We hypothesize that the HD is a LD localization signal and is therefore sufficient to direct proteins to the LD. Here, we are testing if the HD sequence is sufficient to direct a reporter protein to the LD membrane that typically would not localize to the LD. The protein we chose is Env7, a protein kinase that normally anchors to the lysosomal vacuole by palmitoylation of a triple cysteine stretch at its N‐terminus. A new plasmid construct was engineered where the Env9 HD sequence was integrated into GFP tagged Env7 (C13‐15S) plasmid. Cells expressing ENV7 (C13‐15S)‐HD‐GFP were investigated using confocal microscopy to confirm localization of Env7. The HD of Env9 was seen to mediate the localization of Env7 (C13‐15S) to the LD membrane. Thus, the HD is sufficient for localization and is therefore a LD targeting signal. Biochemical analysis is being done to further confirm membrane association of Env7 (C13‐15S) to the LD. Thus far, this research could provide a better insight into LD protein trafficking mechanisms.Support or Funding InformationThis work was supported by NIH AREA #R15 GM85794‐02 to EG and by NSF‐MRI grant DBI0722757 for confocal microscopy. As well as, BUILD grants given by National Institute of General Medical Sciences of the National Institutes of Health under Award Numbers; 8UL1GM118979‐02; 8TL4GM118980‐02; 8RL5GM118978‐02This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.