Endogenous HS resists mitochondria-mediated apoptosis in the adrenal glands via ATP5A1 S-sulfhydration in male mice.

In a previous study, we showed that endogenous hydrogen sulfide (HS) plays a key role in the maintenance of intact adrenal cortex function via the protection of mitochondrial function during endoxemia. We further investigated whether mitochondria-mediated apoptosis is involved in HS protection of adrenal function. LPS treatment resulted in mitochondria-mediated apoptosis in the adrenal glands of male mice, and these effects were prevented by the HS donor GYY4137. In the model of Y1 cells, the LPS-induced mitochondria-mediated apoptosis and blunt response to ACTH were rescued by GYY4137. The HS-generating enzyme cystathionine-β-synthase (CBS) knockout heterozygous (CBS) mice showed mitochondria-mediated apoptosis in the adrenal gland and adrenal insufficiency. GYY4137 treatment restored adrenal function and eliminated mitochondria-mediated apoptosis. Maleimide assay combined with mass spectrometry analysis showed that a number of proteins in mitochondria were S-sulfhydrated in the adrenal gland. ATP5A1 was further confirmed as S-sulfhydrated using a modified biotin switch assay. The level of S-sulfhydrated ATP5A1 was decreased in the adrenal gland of endotoxemic and CBS mice, which was restored by GYY4137. ATP5A1 was identified as sulfhydrated at cysteine 244 by HS. Overexpression of the cysteine 244 mutant ATP5A1 in Y1 cells resulted in a loss of LPS-induced mitochondria-mediated apoptosis and GYY4137 restoration of LPS-induced hyporesponsiveness to ACTH. Collectively, the present study revealed that decreased HS generation leads to mitochondrial-mediated apoptosis in the adrenal cortex and a blunt response to ACTH. S-sulfhydration of ATP5A1 at cysteine 244 is an important molecular mechanism by which HS maintains mitochondrial function and steroidogenesis in the adrenal glands.