Notch1 is a p53 target gene involved in human keratinocyte tumor suppression through negative regulation of ROCK1/2 and MRCKα kinases

Abstract

Little is known about the regulation and function of the Notch1 gene in negative control of human tumors.Here we show that Notch1 gene expression and activity are substantially down-modulated in keratinocyte cancer cell lines and tumors, with expression of this gene being under p53 control in these cells. Genetic suppression of Notch signaling in primary human keratinocytes is sufficient, together with activated ras, to cause aggressive squamous cell carcinoma formation. Similar tumor-promoting effects are also caused by in vivo treatment of mice, grafted with keratinocytes expressing oncogenic ras alone, with a pharmacological inhibitor of endogenous Notch signaling. These effects are linked with a lesser commitment of keratinocytes to differentiation, an expansion of stem cell populations, and a mechanism involving up-regulation of ROCK1/2 and MRCK␣ kinases, two key effectors of small Rho GTPases previously implicated in neoplastic progression. Thus, the Notch1 gene is a p53 target with a role in human tumor suppression through negative regulation of Rho effectors.[Keywords: Notch; p53; ROCK/MRCK; stem cells; squamous cell carcinoma; in vivo siRNA delivery] Supplemental material is available at http://www.genesdev.org.

Does Notch play a tumor suppressor role across diverse squamous cell carcinomas?

“…Inhibition of Notch activation in primary keratinocytes by the expression of a dominant negative peptide competing for MAML1 binding to Notch/CSL complex can cause aggressive SCC formation depending on activated ras 26. Consistently, conditional transgenic mice were generated by expressing a dominant negative MAML1 protein to inhibit Notch signaling in the epidermis.…”

“…Decreased levels of NOTCH transcript and protein together with a parallel reduction in HES1 gene (a common target of Notch pathway) were observed in CSCC cell lines 26. Furthermore, reduced levels of activated NOTCH1 were detected in CSCC, correlating with their NOTCH1 mutation status 24.…”

“…In vivo treatment of grafted mice with γ ‐secretase inhibitor DAPT (a pharmacological inhibitor of Notch activation) resulted in tumor‐promoting effects, depending on oncogenic ras 26. A recent clinical Phase III trial of Semagacestat, a γ ‐secretase/Notch inhibitor, was halted in part because of an increased risk of skin cancers compared with those in the placebo arm 31.…”

“…p21's role in tumor suppression was earlier investigated where p21 knockout keratinocytes exhibited significant downmodulation of differentiation markers and dramatic increase in proliferative potential 32. The inhibition of Notch signaling by genetic methods or by using pharmacological Notch inhibitors in human keratinocytes, together with activated ras, can cause aggressive SCC formation in grafted mice 26. This is similar to that previously revealed for ras‐transformed p21 knockout keratinocytes 32, suggesting that NOTCH1 and p21 may overlap to perform their tumor suppressing functions.…”

“…This is similar to that previously revealed for ras‐transformed p21 knockout keratinocytes 32, suggesting that NOTCH1 and p21 may overlap to perform their tumor suppressing functions. However, the general role of p21 remains to be investigated as p21 levels were selectively decreased in a subset of human skin SCCs tested 26, whereas in human keratinocytes, activation of NOTCH1 was found to lead to modest increase in p21 levels 33.…”