MicroRNA 203 Expression in Keratinocytes Is Dependent on Regulation of p53 Levels by E6

Abstract

A screen of microRNA (miRNA) expression following differentiation in human foreskin keratinocytes (HFKs) identified changes in several miRNAs, including miRNA 203 (miR-203), which has previously been shown to play an important role in epithelial cell biology by regulating p63 levels. We investigated how expression of human papillomavirus type 16 (HPV16) oncoproteins E6 and E7 affected miR-203 expression during proliferation and differentiation of HFKs. We demonstrated that miR-203 expression is reduced in HFKs where p53 function is compromised, either by the viral oncoprotein E6 or by knockout of p53 using short hairpin RNAs (p53i). We show that the induction of miR-203 observed during calcium-induced differentiation of HFKs is significantly reduced in HFKs expressing E6 and in p53i HFKs. Induction of miR-203 in response to DNA damage is also reduced in the absence of p53. We report that proliferation of HFKs is dependent on the level of miR-203 expression and that overexpression of miR-203 can reduce overproliferation in E6/E7-expressing and p53i HFKs. In summary, these results indicate that expression of miR-203 is dependent on p53, which may explain how expression of HPV16 E6 can disrupt the balance between proliferation and differentiation, as well as the response to DNA damage, in keratinocytes.MicroRNAs (miRNAs) are small, noncoding RNA molecules that can regulate protein expression at the posttranscriptional level by targeting mRNAs for degradation or translational repression (13). Over the past decade, a growing body of evidence has shown that they play a fundamental role in the development, function, and maintenance of tissues and cells in various organisms. Their importance is perhaps best demonstrated by studies on the RNase Dicer, which plays an integral role in the processing and generation of mature miRNA molecules, as mutations in Dicer cause severe developmental defects in Caenorhabditis elegans (22), while knockout of Dicer in mice is embryonically lethal (3). Moreover, it is now known that many miRNAs are implicated in several disease states, including heart disease (2, 23), viral infection (38), and many different cancers (7, 44), leading to increased interest in the biology and function of individual miRNAs in various cell processes.In skin physiology, miRNAs have been shown to be involved both in normal processes, including epidermal development and hair follicle morphogenesis (48), and in skin-specific pathologies, such as psoriasis (37), systemic lupus erythematous (SLE) (8), and skin carcinogenesis (14). The importance of miRNAs in skin epithelial development is particularly emphasized when their expression is repressed by epidermal cellspecific deletion of Dicer in mouse models, which results in several defects, such as epidermal evagination and abnormal hair follicle development, although it is worth noting that epidermal differentiation is apparently unaffected (1, 48). To date, several individual miRNAs that are highly expressed in the skin epithelium have been identified, including m...

Epstein-Barr Virus Downregulates MicroRNA 203 through the Oncoprotein Latent Membrane Protein 1: a Contribution to Increased Tumor Incidence in Epithelial Cells

“…Studies have shown that miR-203 decreases cell growth and inhibits tumor invasion by regulating the expression of its target genes (13,18,35). EBV and LMP1 facilitate the G 1 /S transition in cell cycle progression and transformation (32,33,51).…”

MicroRNA-203 induces apoptosis by upregulating Puma expression in colon and lung cancer cells

“…Likewise, it is well known that miR-203 promotes keratinocyte proliferation and differentiation by targeting p63, a member of the p53 family (29)(30)(31)(32)(33)(34). For instance, McKenna et al showed that p53 induces miR-203 in keratinocytes of human foreskin (12). Additionally, p53 has been shown to regulate miR-203 expression in colon cancer cells (35).…”

p63 maintains keratinocyte proliferative capacity through regulation of Skp2–p130 levels

“…Tissue embedding and H&E staining were carried out as previously described (McKenna et al, 2010). Images were taken on an Olympus BH-2 microscope with an Olympus D25 camera using Cell B software (Olympus).…”

“…The predominant isoform expressed in basal keratinocytes and stratifying epithelia (Yang et al, 1998) is DNp63a, which contains a sterile alpha motif (SAM domain) and transcriptional inhibitory domain in its extended Cterminus, which can facilitate transcriptional repression in cis or trans. Regulation of p63 levels is further complicated by posttranscriptional targeting by microRNAs such as mir-203 (McKenna et al, 2010;Yi et al, 2008) or by being targeted for degradation by E3 ligases such as ITCH and FBW7 (Galli et al, 2010;Rossi et al, 2006).…”