Cystine-dependent antiporters prevent sulfur stress by excreting surplus supersulfide from cells

<p>Supersulfides play a role in redox homeostasis, but adaptive cell responses to excess sulfur stress are not well understood. To address this issue, we generated transgenic (Tg) mice overexpressing cystathionine gamma-lyase (CSE) to catalyze transformations of cystathionine to cysteine (CysSH) and of cystine (CysSSCys) to CysSH persulfide (CysSSH). CSE Tg mice treated with excess cystine exhibited clear sulfur stress, as manifested by atrophy of the organs. Contrary to expectations, CysSSH concentrations were comparable in a variety of tissues of wild-type (WT) and CSE Tg mice; however, plasma levels of CysSSH in CSE Tg mice were significantly higher than those in WT mice. This export of surplus intracellular CysSSH was also characterized in primary hepatocytes from CSE Tg mice. Exposure of primary hepatocytes to sodium tetrasulfide (Na₂S₄) as a sulfur stress generator resulted in enhancement and rapid decline of intracellular CysSSH levels, accompanied by a concentration-dependent release of CysSSH into the extracellular space. Interestingly, among all amino acids, CysSSCys was found to be essential for CysSSH export from primary mouse hepatocytes, HepG2 cells, and HEK293 cells during Na₂S₄ exposure, suggesting that cystine/glutamate transporter (SLC7A11) contributes, at least partially, to CysSSH export. We established HepG2 cell lines with knockout and overexpression of SLC7A11 and used them to confirm that SLC7A11 is a predominant antiporter of CysSSCys and CysSSH. Furthermore, we found that cellular proteins undergo polysulfidation in response to Na₂S₄-mediated sulfur stress, which results in a lower mitochondrial membrane potential and concentration-dependent Na₂S₄ cytotoxicity. These results suggest that there are CysSSCys-dependent antiporters that pump excess CysSSH from cells during sulfur stress.</p>