StarD7 Mediates the Intracellular Trafficking of Phosphatidylcholine to Mitochondria

Abstract

Steroidogenic acute regulatory protein-related lipid transfer (START) domains, found in 15 mammalian proteins termed StarD1-StarD15, are lipid-binding domains implicated in the intracellular lipid transport systems. In the present study, we analyzed the lipid ligand and function of StarD7. We found two variable forms of mammalian StarD7, termed StarD7-I and StarD7-II. Unlike StarD7-II, StarD7-I contained a mitochondrial-targeting sequence in its N terminus. Overexpressed StarD7-I tagged with V5/His in HEPA-1 cells was mainly observed in the mitochondria of cells prepared at low cellular density, but it was distributed in the cytoplasm of high density cells. StarD7-II was constantly distributed in the cytoplasm at any cellular density. Endogenous StarD7 in HEPA-1 cells and rat liver was also distributed in both the cytoplasm and the mitochondria. A protease K protection assay indicated that the mitochondrial StarD7 was associated with the outer mitochondrial membrane. The purified recombinant StarD7 specifically catalyzed the transfer of PC between lipid vesicles in vitro. Furthermore, the intracellular transport of fluorescent PC that was exogenously incorporated into the mitochondria was increased in cells that overexpressed StarD7-I. These results suggest that StarD7 facilitates the delivery of PC to mitochondria in non-vesicular system.

StarD7 Protein Deficiency Adversely Affects the Phosphatidylcholine Composition, Respiratory Activity, and Cristae Structure of Mitochondria

“…StarD7-I localizes in mitochondria and cytosol, whereas StarD7-II localizes in the cytosol. In HEPA-1 cells, the intracellular transport of exogenously incorporated fluorescent PC into mitochondria is increased when StarD7-I is overexpressed (9). Thus, we concluded that StarD7 is a soluble carrier protein involved in the intracellular transfer of PC to mitochondria.…”

“…In our previous study, we showed that the soluble carrier protein StarD7 selectively transfers PC to mitochondria via a mitochondrion-targeting signal at the N terminus and a lipid-transfer domain at the C terminus (9). Here, to further elucidate the role of StarD7 in mitochondrial phospholipid homeostasis, respiration, and morphology, the StarD7 gene was silenced and deleted.…”

“…Western Blotting Analysis-Anti-StarD7 antibody was prepared as reported previously (9). Other primary antibodies are as follows: total OXPHOS rodent WB antibody mixture (Abcam, Cambridge, UK) for complexes; rabbit anti-actin (Sigma); anti-optic atrophy 1 (Opa1) clone18/OPA1 (BD Transduction Laboratories); and anti-succinate dehydrogenase complex iron sulfur subunit A (SDHA) (Cell Signaling Technology).…”

“…StarD7 KO and KD Altered the Mitochondrial Phospholipid Content and Composition-We speculated that the amount of mitochondrial PC might be decreased in StarD7-KO cells because StarD7 functions in the transfer of PC to mitochondria (9). Thus, we analyzed the effect of deletion of StarD7 on the mitochondrial phospholipid content and constitution.…”

“…In a previous study, we found an alternative pathway for the transport of phospholipids to mitochondria involving StarD7 (START domain-containing protein) (9). StarD7 is a member of the steroidogenic acute regulatory protein-related lipid transfer (START) domain-containing family.…”