Abstract
ABSTRACTIt has now been clearly shown that lipid droplets (LDs) play a dynamic role in the cell. This was reinforced by LD proteomics which suggest that a significant number of trafficking proteins are associated with this organelle. Using microscopy, we showed that LDs partly co-localize with the vacuole in S. cerevisiae. Immunoblot experiments confirmed the association of the vacuolar Rab GTPase Rab7-like Ypt7p with LDs. We observed an increase in fatty acid content and LD number in ypt7Δ mutant and also changes in LD morphology and intra LD fusions, revealing a direct role for Ypt7p in LD dynamics. Using co-immunoprecipitation, we isolated potential Ypt7p partners including, Vma13p, the H subunit of the V1 part of the vacuolar (H+) ATPase (V-ATPase). Deletion of the VMA13 gene, as well as deletion of three other subunits of the V1 part of the V-ATPase, also increased the cell fatty acid content and LD number. Mutants of the Homotypic fusion and vacuole protein sorting (HOPS) complex showed similar phenotypes. Here, we demonstrated that LD dynamics and membrane trafficking between the vacuole and LDs are regulated by the Rab7-like Ypt7p and are impaired when the HOPS complex and the V1 domain of the V-ATPase are defective.
Highlights
In yeast, animals and other organisms, storage lipids are maintained in the cytoplasm in specialized organelles called lipid droplets (LDs) or lipid bodies (Brasaemle and Wolins, 2012; Chapman et al, 2012; Murphy, 2012; Walther and Farese, 2012)
LDs and their relationship with the vacuole/late endosome via Ypt7p transmission electron microscopy (TEM) and epifluorescence observations of S. cerevisiae wild-type (WT) cells (Fig. 1) showed that LDs are close to both the nucleus and vacuole (51% were in close proximity with nucleus alone, vacuole alone or both)
The Homotypic fusion and vacuole protein sorting (HOPS) pathway and LD dynamics We investigated whether the relationship between LDs and the vacuole could occur via the vacuolar HOPS tethering complex, which is recruited by Ypt7p
Summary
Animals and other organisms, storage lipids are maintained in the cytoplasm in specialized organelles called lipid droplets (LDs) or lipid bodies (Brasaemle and Wolins, 2012; Chapman et al, 2012; Murphy, 2012; Walther and Farese, 2012). These structures consist mainly of a core of neutral lipids (triacylglycerols and/or steryl esters) enclosed in a monolayer of phospholipids, and contain a number of proteins which vary considerably with the species (Beller et al, 2010; Hodges and Wu, 2010). It has been demonstrated that LDs were functionally connected to the ER in S. cerevisiae (Jacquier et al, 2011)
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