Regulation of Notch1 Gene Expression by p53 in Epithelial Cells

Abstract

The E6 protein of cervical cancer-associated human papillomaviruses (HPVs) is known to suppress keratinocyte differentiation through unidentified mechanisms. Notch1 is a determinant of keratinocyte differentiation and functions as a tumor suppressor in mammalian epidermis. Here, we report that the Notch1 gene is a novel target of p53 and can be down-regulated by E6 through p53 degradation in normal human epithelial cells. Thus, inactivation of p53 by E6 or short-hairpin RNA (shRNA) resulted in reduced Notch1 expression at the transcription level, and a p53-responsive element could be identified in the Notch1 promoter. The expression of E6, p53 shRNA, or Notch1 shRNA suppressed both spontaneous keratinocyte differentiation in culture and its induction upon DNA damage. Furthermore, the induction of Notch1 and differentiation makers as well as thickening of the epidermal layer upon UV irradiation was observed in wild-type but not in p53-deficient mouse skin. Together, our findings not only demonstrate a novel link between p53 and Notch1 in keratinocyte differentiation upon genotoxic stress but also suggest a novel tumor suppressor mechanism of p53 in the development of squamous cell carcinomas, including HPV-induced tumors.A specific group of so-called high-risk human papillomaviruses (HPVs), such as HPV16 and HPV18, is associated with more than 90% of cervical cancers (60). Infection with these HPVs causes cervical dysplasia or low-grade cervical intraepithelial neoplasia (CIN), and cervical cancers are thought to arise from these lesions after long periods of time (32, 70). The E6 and E7 proteins of HPVs are expressed at relatively low levels in the basal cells of low-grade CIN lesions, where the viral genomes replicate episomally. When high-level expression of E6 and E7 occurs, in most cases with integration of viral genomes into the host genome, neoplastic development is believed to be initiated (59). In fact, E6 and E7 proteins are invariably expressed in HPV-positive cervical cancer cells and inactivate the major tumor suppressors p53 and Rb, respectively, thus contributing to HPV-induced oncogenesis.

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

“…In keratinocytes, p53 was proposed as a positive regulator of Notch signaling. Specifically, p53‐responsive element has been identified in the Notch1 promoter 26, 45, 46. Consequently, NOTCH1 expression is upregulated by increased p53 in human primary keratinocytes and CSCC cell lines 26, 45.…”

p53 Regulates Toll-Like Receptor 3 Expression and Function in Human Epithelial Cell Lines

“…Samples were analyzed by PCR using LA Taq polymerase (TaKaRa) according to the recommended protocol. The primers used (Table 6) recognize a fragment of the human TLR3 promoter, p21 promoter, or GAPDH promoter (45).…”

Developmental SHP2 dysfunction underlies cardiac hypertrophy in Noonan syndrome with multiple lentigines

“…Previously, it was demonstrated that NOTCH1 is a direct target of p53 in human keratinocytes (54) and epithelial cells (55). Increasing AKT signaling, which drives MDM2-mediated p53 degradation (56), may therefore cause decreased NOTCH1 expression in NSML.…”