MicroRNA profiling in E6/E7 HPV-transformed human keratinocytes.

MicroRNAs (miRNAs) are a class of naturally occurring noncoding RNAs that play a key role in gene regulation. These transcripts are highly conserved single-stranded RNAs (~22 nucleotides) that are cleaved from larger hairpin precursor transcripts. MiRNAs exert their regulatory effects through the RNA interference machinery by cleaving the mRNA targets or repressing their translation, and are predicted to regulate at least one third of all human genes. This newly discovered form of posttranscriptional gene regulation has been demonstrated to play important roles in a variety of fundamental cellular processes, including cell proliferation, differentiation, and death. As a result, miRNAs may be directly involved in the development and progression of human diseases, such as cancer. In fact, accumulating evidence indicates a role of miRNAs in tumorigenesis as it is well known that tumor cells bear a specific and altered pattern of miRNA expression. We perfomed TaqMan® MicroRNA Assays (Applied Biosystems, USA) for the quantitation of 384 mature miRNA expression in normal primary keratinocytes (HFK) compared to primary keratinocytes transformed by both E6 and E7 proteins derived from mucosal HPV-16 or cutaneous HPV-38 (named K16 and K38 cells, respectively). Preliminary results identified 53 miRNAs deregulated in K16 and K38 cells with respect to HFK. Interestingly, it is detectable a strong increase (at least 25-fold) in the expression of 12 miRNAs in K16 and 7 miRNAs in K38, as well as a strong decrease in the expression of 14 miRNAs in K16 and 35 miRNAs in K38 cells. Moreover, 14 miRNAs are up-regulated in K16 but down-regulated in K38 whereas only two miRNAs are down-regulated in K16 but up-regulated in K38 cells. These data suggest that different clusters of miRNA targets may be correlated with different HPV genotypes-induced tumorigenesis. In this respect, our results have been compared to miRNA expression profiles of HPV-positive cervical cancer cell lines and tissues, obtained by other authors. As expected, the miRNA expression profile of K16 cells appears to be similar. Conversely, major differences in K38 cells miRNA profile have been observed. In addition, studies of miRNA expression in K16 and K38 cells treated with IFN-β are in progress, since preliminary results have indicated the ability of the antitumoral agent IFN-β to interfere with E6 and/or E7-induced miRNA deregulation.