A conserved megaprotein-based molecular bridge critical for lipid trafficking and cold resilience

Abstract

Cells adapt to cold by increasing levels of unsaturated phospholipids and membrane fluidity through conserved homeostatic mechanisms. Here we report an exceptionally large and evolutionarily conserved protein LPD-3 in C. elegans that mediates lipid trafficking to confer cold resilience. We identify lpd-3 mutants in a mutagenesis screen for genetic suppressors of the lipid desaturase FAT-7. LPD-3 bridges the endoplasmic reticulum (ER) and plasma membranes (PM), forming a structurally predicted hydrophobic tunnel for lipid trafficking. lpd-3 mutants exhibit abnormal phospholipid distribution, diminished FAT-7 abundance, organismic vulnerability to cold, and are rescued by Lecithin comprising unsaturated phospholipids. Deficient lpd-3 homologues in Zebrafish and mammalian cells cause defects similar to those observed in C. elegans. As mutations in BLTP1, the human orthologue of lpd-3, cause Alkuraya-Kucinskas syndrome, LPD-3 family proteins may serve as evolutionarily conserved highway bridges critical for ER-associated non-vesicular lipid trafficking and resilience to cold stress in eukaryotic cells.

LPD-3 as a megaprotein brake for aging and insulin-mTOR signaling inC. elegans

“…The drastically shortened lifespan of lpd-3 mutants prompted us to seek to identify the underlying mechanisms. By analyzing RNA-seq datasets for lpd-3 mutants compared to wild type 14 , we discovered that LPD-3 deficiency led to nearly 40-fold increase in the expression level of ins-7, which represents one of the most highly lpd-3 up-regulated genes and encodes an agonist insulin with previously reported effects on aging and lifespan in C. elegans [18][19][20] (Fig. 1f).…”

“… a , RNA-seq reveals drastic and specific ins-7 up-regulation among all ins genes examined. Results were derived and analyzed from RNA-seq datasets we published earlier 14 . FPKM, Fragments Per Kilobase of transcript per Million mapped reads.…”

“…By contrast, ins-7p:: ins-7 ::GFP in lpd-3 mutants showed drastically increased GFP abundance throughout the intestine, and particularly at higher cultivation temperatures (25 °C). We showed previously that lpd-3 mutants are defective in ER-to-PM phospholipid trafficking, exhibit sensitivity to thermal stress that can be normalized by Lecithin 14 . We confirmed in this study that both shortened lifespan and ins-7 up-regulation phenotypes were also rescued by Lecithin (Supplementary Fig.…”

“…2e). As reduced LPD-3 activity results in impaired ER-to-PM phospholipid trafficking, a compensatory lipid homeostatic mechanism is likely triggered to up-regulate sphingolipids in lpd-3 mutants 14 . Given reported sphingolipid modulation of mTOR signaling and longevity [27][28][29] , we next explored specifically the link of hexaceramide biosynthetic enzyme HYL-1 to ins-7 and found that hyl-1 RNAi led to strong suppression of ins-7p::ins-7::GFP in lpd-3 mutants.…”

“…In recent studies, we identified lpd-3 in a mutagenesis screen and discovered that the 452 kDa megaprotein LPD-3 is a bridge-like tunnel protein with evolutionarily conserved roles in lipid trafficking from endoplasmic reticulum to plasma membranes (Fig. 1a) 14 . Mutations in BLTP1 , the human orthologue of lpd-3 , cause an autosomal recessive Alkuraya-Kucinskas syndrome (AKS) 15–17 Since most AKS patients die prematurely, we sought to determine the lifespan of lpd-3 mutants in C. elegans .…”