Molecular Characterisation of High Risk Human Papillomavirus (Hpv): Genotyping, Genome Analysis and Functional Interaction Between Hpv 16 E2 and L1

Abstract

High risk human papillomavirus (HPV) is the etiological agent of cervical cancer, which is the third most important cancer mortality in women worldwide. The incidence of cervical cancer in Pakistan and its relevance to HPV is very little documented and exact figures are not known. Timely detection of HPV plays significant role to decrease the disease progression, therefore in this study the incidence of high risk HPV is investigated. 80 cervical lesion specimens were screened for HPV 16 and 18 infection and 92 % samples were found positive for HPV 16 and 18 along with high rate of co-infection. Based on these findings, genome analysis of HPV 16 from Pakistan was first time carried out using cervical cancer specimen. Sequence analysis has shown 11 changes both at amino acid and nucleotide level. Out of this, 7 novel and 2 sequence conflicts were observed. Such changes indicate the genetic basis of viral evolution. Moreover, the L1 protein based phylogenetic study suggests that HPV 16 strain from Pakistan groups with European taxa. The PV’s E2 and L1 protein are known to play key role in virus life cycle by interacting with different cellular and viral proteins. The association of E2 with L1 has been characterised for the first time in this study. The C terminus region from amino acid 335-365 of E2 binds with L1. Interestingly, this association modulates E2 dependent transcription and replication in a dose dependent manner. However, L1 was not forming the replication foci along with E1 and E2, when fixed with formaldehyde. Cellular localisation studies shows that both E2 and L1 co-localised in nucleus. E2 alone is expressed in nucleolus however, presence of L1 excludes nucleolar E2 and redistributes it into nucleus. It is hypothesised that re- localisation of E2 by L1 protein is responsible for E2 dependent functions. The co- localisation of both proteins was also observed in intermediate epithelium layer of HPV 16 positive organotypic raft sections, where the vegetative virus replication takes place. The functional interaction of HPV 16 E2-L1 shown in this study provides novel insight towards virus life cycle.