Persulfidation of transcription factor FOXO1 at cysteine 457: A novel mechanism by which H2S inhibits vascular smooth muscle cell proliferation

Introduction: The proliferation of vascular smooth muscle cells (VSMCs) is an important physiological and pathological basis for many cardiovascular diseases. Endogenous hydrogen sulfide (H2S), the third gasotransmitter, is found to preserve vascular structure by inhibiting VSMC proliferation. However, the mechanism by which H2S suppresses VSMC proliferation has not been fully clear. Objectives: This study aimed to explore whether H2S persulfidates the transcription factor FOXO1 to inhibit VSMC proliferation. Methods: After the proliferation of VSMC A7r5 cells was induced by endothelin-1 (ET-1), FOXO1 phosphorylation and proliferating cell nuclear antigen (PCNA) expression were detected by Western blotting, the degree of FOXO1 nuclear exclusion and PCNA fluorescent signals in the nucleus were detected by immunofluorescence, and the persulfidation of FOXO1 was measured through a biotin switch assay. Results: The results showed that ET-1 stimulation increased cell proliferation, FOXO1 phosphorylation and FOXO1 nuclear exclusion to the cytoplasm in the cells. However, pretreatment with NaHS, an H2S donor, successfully abolished the ET-1-induced increases in the VSMC proliferation, FOXO1 phosphorylation, and FOXO1 nuclear exclusion to the cytoplasm. Mechanistically, H2S persulfidated the FOXO1 protein in A7r5 and 293T cells, and the thiol reductant DTT reversed this effect. Furthermore, the C457S mutation of FOXO1 abolished the H2S-induced persulfidation of FOXO1 in the cells and the subsequent inhibitory effects on FOXO1 phosphorylation at Ser256, FOXO1 nuclear exclusion to the cytoplasm and cell proliferation. Conclusion: Thus, our findings demonstrated that H2S might inhibit VSMC proliferation by persulfidating FOXO1 at Cys457 and subsequently preventing FOXO1 phosphorylation at Ser256. (C) 2020 The Authors. Published by Elsevier B.V. on behalf of Cairo University.