MicroRNA-15b is induced with E2F-controlled genes in HPV-related cancer.

The expression of microRNAs (miRNAs) are altered in a number of human carcinomas and in many physiological processes in the organism (Calin et al, 2002; Michael et al, 2003; van Rooij et al, 2006; Volinia et al, 2006; Sonkoly et al, 2007; Wang et al, 2008). MicroRNAs are a class of conserved, short, non-coding RNAs that modulate gene expression through the RNAi pathway (Hutvagner and Zamore, 2002; Bartel, 2004). Mature miRNAs are single-stranded ∼22 nucleotides derived from hairpin-structured miRNA precursors, by the RNaseIII enzymes Drosha and Dicer (Lee et al, 2002; Bartel, 2004). MicroRNAs differ from siRNAs in that they generally require only partial complementarity to the target sequence, which is often in the 3′-untranslated region of the mRNAs. Binding of a miRNA to a target mRNA sequence leads to transcriptional or post-transcriptional changes of the expression of the corresponding protein (Bartel, 2004; Lim et al, 2005; Rajewsky, 2006). Cleavage of the target mRNA may occur when complementarity is perfect. This occurs mostly in plants, but can also be seen in animals (Hutvagner and Zamore, 2002; Zeng and Cullen, 2003). MicroRNAs have been postulated to regulate ∼30% of the human genome (Bartel, 2004). In cancers, miRNAs were proposed to work either as tumour suppressors or as oncogenes through regulation of the protein expression that contributes to oncogenesis (Esquela-Kerscher and Slack, 2006). Through genome-wide mapping of known miRNAs in the human genome, miRNA genes were located to fragile sites and to other genomic regions often involved in cancer development (Calin et al, 2004). For example, the relative incidence of miRNAs at human papilloma virus (HPV)16 integration sites was significantly higher than in the rest of the genome. This is interesting as HPV is accepted as an aetiological agent of some cancers, such as cervical cancer and anal squamous carcinoma (zur Hausen, 1989; Frisch et al, 1997; Daling et al, 2004; Madsen et al, 2008). Squamous cell carcinoma of the anal canal is a rare type of cancer with increasing incidence, probably related to changes in sexual behaviour (Daling et al, 2004). HIV-positive patients also have increased risk of developing anal cancer due to immune suppression secondary to HIV infection (Grulich et al, 2007). The transforming capability of HPV is assigned to E6 and E7 oncogenes (zur Hausen, 2000). These two proteins inactivate p53 and the retinoblastoma (RB1) protein, respectively (Dyson et al, 1989; Werness et al, 1990). Hence, two important pathways in cells that regulate cell growth and apoptosis are inhibited, leading to uncontrolled cell growth. Retinoblastoma protein 1 is a negative regulator of E2F transcription factors, which among others induce the transcription of many genes that control cell fate and proliferation, such as MCM7, MSH6, cyclin E, cyclin A and cyclin B (Iaquinta and Lees, 2007). Recently, HPV E7 was also reported to actually modulate the miR-203 level in keratinocytes to maintain cells in the proliferative state to produce virus particles (Melar-New and Laimins, 2010). In a previous study, we demonstrated that tumours from patients with anal carcinomas could be divided into two distinct groups based on global mRNA expression (Bruland et al, 2008). All tumours expressed HPV16 E7 mRNA. Cluster analysis on a number of E2F and/or high-risk HPV-regulated genes reproduced these two groups. Thus, HPV16 had major influences on the global gene expression in approximately half of the tumours studied. In this study, the aim was to study global miRNA expression in anal carcinomas. Therefore, the patient database and the biopsy material from the previous study have been supplemented with 11 additional tumours. The grouping of tumours into two groups based on E2F and/or high-risk HPV-regulated gene expression was verified in the larger number of HPV-positive anal carcinoma tumours. Here, we report that one miRNA was found to divide tumours into the same two groups, namely miR-15b.