Conditional targeting of phosphatidylserine decarboxylase to lipid droplets

Abstract

Phosphatidylethanolamine is an abundant component of most cellular membranes whose physical and chemical properties modulate multiple aspects of organelle membrane dynamics. An evolutionarily ancient mechanism for producing phosphatidylethanolamine is to decarboxylate phosphatidylserine and the enzyme catalyzing this reaction, phosphatidylserine decarboxylase, localizes to the inner membrane of the mitochondrion. We characterize a second form of phosphatidylserine decarboxylase, termed PISD-LD, that is generated by alternative splicing of PISD pre-mRNA and localizes to lipid droplets and to mitochondria. Sub-cellular targeting is controlled by a common segment of PISD-LD that is distinct from the catalytic domain and is regulated by nutritional state. Growth conditions that promote neutral lipid storage in lipid droplets favors targeting to lipid droplets, while targeting to mitochondria is favored by conditions that promote consumption of lipid droplets. Depletion of both forms of phosphatidylserine decarboxylase impairs triacylglycerol synthesis when cells are challenged with free fatty acid, indicating a crucial role phosphatidylserine decarboxylase in neutral lipid storage. The results reveal a previously unappreciated role for phosphatidylserine decarboxylase in lipid droplet biogenesis.

ER-localized phosphatidylethanolamine synthase plays a conserved role in lipid droplet formation

“…When considering whether this was true of Psd1 orthologs in other organisms, we noted that our previous work identified a glycosylation site on the N-terminus of native Psd1, indicating the ER-localized form of the enzyme must contain a TMD and be integral to the ER membrane (Friedman et al, 2018). In contrast, human PISD is alternatively spliced to produce two isoforms; the canonical long isoform (isoform 1) localizes to the IMM and a shorter isoform (isoform 2) exhibits dual localization (when tagged and overexpressed) at mitochondria and LDs (Kumar et al, 2021). Importantly, the LD-localized isoform in human cells does not contain a predicted TMD and likely targets to LDs via an amphipathic -helix (Kumar et al, 2021).…”

“…Tagging with mNG did not interfere with Psd1 mitochondrial function, as assessed by growth on carbon sources that require respiration (Fig. 1 Recently, work in human cells by Burd and colleagues revealed that the single human PSD, PISD, has multiple spliced isoforms that encode for differentially localized proteins (Kumar et al, 2021). When overexpressed and GFP-tagged, a long isoform of PISD exclusively localized to mitochondria, while a shorter isoform localized to both mitochondria and lipid droplets (LDs).…”

“…In contrast, human PISD is alternatively spliced to produce two isoforms; the canonical long isoform (isoform 1) localizes to the IMM and a shorter isoform (isoform 2) exhibits dual localization (when tagged and overexpressed) at mitochondria and LDs (Kumar et al, 2021). Importantly, the LD-localized isoform in human cells does not contain a predicted TMD and likely targets to LDs via an amphipathic -helix (Kumar et al, 2021). Additionally, while PISD-depleted cells exhibit a defect in incorporation of radiolabeled oleate into TAG, it is unclear whether the LD-targeted form of PISD plays a direct role in LD biogenesis as, to date, no experiments have selectively depleted the LD form of the enzyme (Kumar et al, 2021).…”

“…Importantly, the LD-localized isoform in human cells does not contain a predicted TMD and likely targets to LDs via an amphipathic -helix (Kumar et al, 2021). Additionally, while PISD-depleted cells exhibit a defect in incorporation of radiolabeled oleate into TAG, it is unclear whether the LD-targeted form of PISD plays a direct role in LD biogenesis as, to date, no experiments have selectively depleted the LD form of the enzyme (Kumar et al, 2021). Thus, while yeast Psd1 and human PISD likely target to LDs via distinct mechanisms, whether PSD plays a conserved role in contributing to LD formation is an open question.…”

“…Humans have a PSD encoded by the PISD gene, which is critical for mitochondrial morphology and function (Steenbergen et al, 2005;Tasseva et al, 2013). Recently, however, human PISD was demonstrated to express an alternate isoform (isoform 2) that encodes for an enzyme with a distinct N-terminus (Kumar et al, 2021). When GFP-tagged and overexpressed, this isoform exhibited localization to not only mitochondria, but also to an ER-derived organelle, the lipid droplet (LD).…”