SPAG6 silencing induces autophagic cell death in SKM-1 cells via the AMPK/mTOR/ULK1 signaling pathway

Abstract

As a member of the cancer-testis antigen family, sperm-associated antigen 6 (SPAG6) has been reported to be associated with the pathogenesis of myelodysplastic syndromes (MDS). Previous studies have demonstrated that SPAG6 is upregulated in bone marrow from patients with MDS and MDS-transformed acute myeloid leukemia and that knockdown of SPAG6 expression levels suppressed proliferation and promote apoptosis and differentiation in SKM-1 cells. However, the association between SPAG6 and autophagy in SKM-1 cells remains unclear. Hence, the aim of the present study was to investigate this association and its underlying mechanism. The present study used a short hairpin RNA (shRNA) lentivirus to silence SPAG6 expression levels in SKM-1 cells and demonstrated that SPAG6 knockdown increased autophagy and apoptosis. Furthermore, pharmacologically inhibiting autophagy with chloroquine and 3-methyladenine decreased SPAG6 knockdown-mediated apoptosis, indicating that SPAG6 knockdown-mediated autophagy promoted apoptosis in SKM-1 cells. Additionally, comparedwiththeexpressionlevelsinnegativecontrol-shRNA lentivirus-transfected SKM-1 cells, the protein expression levels of phosphorylated AMP-activated protein kinase (p-AMPK) and phosphorylated unc-51-like autophagy activating kinase 1 (p-ULK1) were upregulated, while phosphorylated mammalian target of rapamycin (p-mTOR) protein expression was downregulated in SPAG6-shRNA lentivirus-transfected cells. Moreover, inhibiting AMPK expression levels with Compound C, a specific inhibitor of AMPK, attenuated SPAG6 knockdown-induced autophagy and apoptosis, suggesting that AMPK-mediated autophagy enhanced the pro-apoptotic effect of SPAG6 knockdown in SKM-1 cells. Taken together, the results of the present study demonstrated that SPAG6 silencing triggered autophagy via regulation of the AMPK/mTOR/ULK1 signaling pathway, which further contributed to the apoptosis of SKM-1 cells induced by SPAG6 knockdown. Thus, the current results indicate that SPAG6 may be a potential therapeutic target against MDS, and that autophagy may represent a potential mechanism for the treatment of MDS.