SOX2 regulates self-renewal and tumorigenicity of human melanoma-initiating cells

Abstract

Melanoma is one of the most aggressive types of human cancer, characterized by enhanced heterogeneity and resistance to conventional therapy at advanced stages. We and others have previously shown that HEDGEHOG-GLI (HH-GLI) signaling is required for melanoma growth and for survival and expansion of melanoma-initiating cells (MICs). Recent reports indicate that HH-GLI signaling regulates a set of genes typically expressed in embryonic stem cells, including SOX2 (sex-determining region Y (SRY)-Box2). Here we address the function of SOX2 in human melanomas and MICs and its interaction with HH-GLI signaling. We find that SOX2 is highly expressed in melanoma stem cells. Knockdown of SOX2 sharply decreases self-renewal in melanoma spheres and in putative melanoma stem cells with high aldehyde dehydrogenase activity (ALDHhigh). Conversely, ectopic expression of SOX2 in melanoma cells enhances their self-renewal in vitro. SOX2 silencing also inhibits cell growth and induces apoptosis in melanoma cells. In addition, depletion of SOX2 progressively abrogates tumor growth and leads to a significant decrease in tumor-initiating capability of ALDHhigh MICs upon xenotransplantation, suggesting that SOX2 is required for tumor initiation and for continuous tumor growth. We show that SOX2 is regulated by HH signaling and that the transcription factors GLI1 and GLI2, the downstream effectors of HH-GLI signaling, bind to the proximal promoter region of SOX2 in primary melanoma cells. In functional studies, we find that SOX2 function is required for HH-induced melanoma cell growth and MIC self-renewal in vitro. Thus SOX2 is a critical factor for self-renewal and tumorigenicity of MICs and an important mediator of HH-GLI signaling in melanoma. These findings could provide the basis for novel therapeutic strategies based on the inhibition of SOX2 for the treatment of a subset of human melanomas.

Sox2 is dispensable for primary melanoma and metastasis formation

“…Nonetheless, our findings do not rule out that SOX2 can exert certain activities in melanoma cells, for instance in isolated cells exposed to varying culture conditions or in the non-physiological environment encountered by human cells upon xenotransplantation into immunocompromised mouse model systems 22 . Of note, studies performed solely in vitro or in combination with xenotransplantation assays provided controversial results, either implicating SOX2 in melanoma cell proliferation or invasion, or also failing to reveal a crucial role in melanoma as in our experiments performed with CRISPR-Cas-mediated knock out cells [20][21][22][23][24] .…”

“…Tissue microarray (TMA) expression analysis revealed that SOX2 protein is expressed in around 50% of all human melanoma 20,21 . In a recent study, RNAi-mediated silencing of SOX2 led to reduced melanoma sphere self-renewal and tumor growth of xenotransplanted cells, supporting an important role of Sox2 in tumor initiation and tumor maintenance 22 . In other reports, however, knockdown of SOX2 did not affect proliferation of human melanoma cells 20,21,23 .…”

“…Of note, studies performed solely in vitro or in combination with xenotransplantation assays provided controversial results, either implicating SOX2 in melanoma cell proliferation or invasion, or also failing to reveal a crucial role in melanoma as in our experiments performed with CRISPR-Cas-mediated knock out cells [20][21][22][23][24] . While the discrepancies between our work and previously published data could potentially be explained by different experimental settings 37 , they could also reflect the heterogeneity of melanoma and, thus, leave open whether SOX2 might fulfill a role in some cases of human melanoma.…”

Physcion inhibits the metastatic potential of human colorectal cancer SW620 cells in vitro by suppressing the transcription factor SOX2

“…The shRNA oligonucleotides for SOX2 gene knockdown were designed and synthesized as previously described [23] . Two different shRNA sequences and one scramble sequence as a control were subcloned into plasmid vector pGCsi-H1 following manufacturer's instructions.…”

ALDH1 expression and activity increase during tumor evolution in sarcoma cancer stem cell populations

“…Consequently, the ALDEFLUOR assay has been widely used to isolate CSC subpopulations in various tumors including different types of sarcomas 23,[27][28][29][30][31] . These CSC subpopulations commonly overexpress pluripotency factors such as Sex-determining region Y-Box2 (SOX2), which plays important roles in the regulation of self-renewal and tumorigenicity in CSC subpopulations of several types of cancer [32][33][34][35][36][37] . The process of emergence/evolution of CSC subpopulations from the cell-of-origin in sarcomagenesis has barely been addressed.…”