A positive feedback loop between the oncogenic WIP1 phosphatase and the transcription factor GLI1 regulates the Hedgehog signaling

The Hedgehog-GLI (HH-GLI) signaling plays a critical role in controlling growth and tissue patterning during embryogenesis and is implicated in a variety of human malignancies, including those of the skin. Phosphorylation events have been shown to regulate the activity of the GLI transcription factors, the final effectors of the HH-GLI signaling pathway. Here, we show that WIP1 (or PPM1D), an oncogenic phosphatase amplified/overexpressed in several types of human cancer, is a positive modulator of the HH signaling. Mechanistically, WIP1 enhances the function of GLI1 by increasing its transcriptional activity, nuclear localization and protein stability, but not of GLI2 nor GLI3. We also find that WIP1 and GLI1 are in a complex. Modulation of the transcriptional activity of GLI1 by WIP1 depends on the latter’s phosphatase activity and, remarkably, does not require p53, a known WIP1 target. Functionally, we find that WIP1 is required for melanoma and breast cancer cell proliferation and self-renewal in vitro and melanoma xenograft growth induced by activation of the HH signaling. Pharmacological blockade of the HH pathway with the SMOOTHENED antagonist cyclopamine acts synergistically with inhibition of WIP1 in reducing growth of melanoma and breast cancer cells in vitro. We also show that HH-GLI signaling directly regulates the expression of the transcription factor E2F1 through the direct binding of GLI1 and GLI2 to a region of E2F1 promoter between -132bp and -269bp from the transcription start site. Functionally, we find that E2F1 is required for melanoma cell proliferation induced by activation of the HH signaling and that E2F1 silencing reduces self-renewal of melanoma cells. Because E2F1 directly controls WIP1 expression, our data indicate the presence of a positive feedback loop that fuels HH pathway activation in melanoma, thus sustaining cancer cell growth and cancer stem cell self-renewal. We also suggest a possible novel therapeutic approach for human melanomas and, possibly, other cancer types expressing WIP1 and with activated HH pathway, by combining WIP1 and HH signaling inhibition.