A trypsin-sensitive protein is required for utilization of exogenous cholesterol for pregnenolone synthesis by placental mitochondria.
The utilization of cholesterol for steroid hormone synthesis by human placental mitochondria is poorly understood. The human placenta does not express the steroidogenic acute regulator protein, which is critical for cholesterol delivery to the cholesterol side chain cleavage system in adrenal and gonadal mitochondria. We explored the mechanism underlying cholesterol transport in human placental mitochondria by measuring its transformation into pregnenolone. Mitochondria of syncytiotrophoblast from human term placenta were isolated by centrifugation through a sucrose gradient. The synthesis of pregnenolone in the presence of exogenous cholesterol was increased two-fold in syncytiotrophoblast mitochondria. Treatment of mitochondria with trypsin prevented the increase in the synthesis of pregnenolone in the presence of exogenous cholesterol. However, when 22-OH cholesterol, a substrate that readily crosses membranes, was added, the trypsin-treated mitochondria synthesized increased amounts of pregnenolone. The trypsin-treated mitochondria were intact, since oxygen consumption, succinate dehydrogenase and the adenine nucleotide translocase activities were not significantly different from in untreated mitochondria. However, activity of NADH cytochrome c oxidoreductase, an outer mitochondrial membrane enzyme, was reduced in the trypsin-treated mitochondria, reflecting the selective degradation of proteins. In addition, SDS-PAGE analysis revealed the loss of a prominent 34 kDa band which proved to be a novel porin-like protein that binds to cholesterol. These results support our previous assumption that human placental mitochondria employ a novel protein(s)-mediated the mechanism to take up cholesterol for steroidogenesis.
Espinosa-García
M T
MT
Departamento de Bioquímica, Facultad de Medicina, Universidad Nacional Autónoma de México, México, D.F., Coyoacán 04510, Mexico.
Strauss
J F
JF
Martínez
F
F
D000595:Amino Acid Sequence; D002784:Cholesterol / Q000378:metabolism*; D005260:Female; D006801:Humans; D008928:Mitochondria / Q000378:metabolism*; D009245:NADH Dehydrogenase / Q000378:metabolism; D010920:Placenta / Q000648:ultrastructure*; D018272:Porins / Q000737:chemistry / Q000378:metabolism; D011247:Pregnancy; D011284:Pregnenolone / Q000096:biosynthesis*; D014357:Trypsin / Q000494:pharmacology*
D018272:Porins; D011284:Pregnenolone; D002784:Cholesterol; D009245:NADH Dehydrogenase; D014357:Trypsin