Deregulation of PiwiL1 and associated RNAs in cervical cancer: is HPV oncogene responsible?

Abstract

Although HPV infection is the major cause of tumorigenesis in cervical cancer; interestingly it has been noted that HPV infection does not always lead to cervical cancer. This suggests that there could be some unknown factor that directly or indirectly interacts with these HPV oncoproteins leading to tumorigenesis. Our study aims to understand whether any correlation exists between the Piwi homologs with HPV oncoproteins in cervical cancer. To begin with, the expression pattern of Piwi proteins were evaluated in cultured cervical cancer (C33A, SiHa, and CaSki) cell lines. We observed that all the Piwi variants were differentially expressed in all the selected cervical cancer cell lines with PiwiL1 showing the highest expression in CaSki cells, suggesting a positive correlation with HPV oncoproteins. On investigating this correlation further, we found that PiwiL1 was co‐expressed and physically interacted with HPV oncoproteins E6 and E7. Not only E6 and E7 physically interact with PiwiL1, but also accentuated its expression when over‐expressed in HaCaT cells. Since PiwiL1 protein is known for its overexpression in many cancers, we wanted to know how this molecule is regulated in cancer. For this, we have carried out an in silico analysis of the PiwiL1 promoter to find the possible binding TFs using multiple online tools such as Alibhabha, AllegenPromo, and TFsitescan. We further shortlisted the TFs based on their function in cancer, interestingly we found both p53 and E2F in the list. Further analysis of these selected factors suggested that PiwiL1 promoter activity was significantly upregulated in the presence of E2F, whereas it was downregulated with p53. The binding of these transcription factors and their differential regulation of PiwiL1 promoter could be one of the reasons for its over‐expressed status in cancer. In addition to this, it is also known that the Piwi‐piRNA complex act as guiding signals for factors that play a significant role in tumorigenesis. Therefore, we have constructed a pipeline to predict the piRNAs from a small RNA Sequencing Data (SRP119662) obtained from the NCBI database, we identified 2274 piRNAs from both control and tumor samples, out of this, only 9 piRNAs were found to be significantly differentially expressed compared to control. Even though in silico analysis predicted 6 piRNAs to be significantly upregulated and 3 piRNAs to be significantly downregulated, the qPCR analysis was not entirely in accordance to the prediction. Out of the 3 down‐regulated piRNAs, hsa_piR_007863 showed elevated levels in CaSki in comparison to HaCaT while hsa_piR_002320 which was predicted to be upregulated in tumor showed lower expression. Our preliminary data suggest that PiwiL1 protein and associated piRNAs may have an important role in HPV‐associated tumorigenesis, whose exact molecular mechanism needs to be further evaluated.