title: Differential expression of HPV16 L2 gene in cervical cancers harboring episomal HPV16 genomes: influence of synonymous and non-coding region variations.
abstract: We tested the hypothesis that (i) synonymous variations within the coding regions, and (ii) variations within the non-coding regions of HPV, influence cervical cancer (CaCx) pathogenesis under the impact of intact HPV16 genomes. Whole genome sequence analysis of HPV16 isolates within 70 CaCx cases and 25 non-malignant samples revealed that synonymous variations were significantly higher within the E6 (p = 0.014), E5 (p = 0.001) and L2 (p = 0.0002) genes of HPV16 isolates within cases, compared to isolates within non-malignant samples. All of the 25 (100%) humanized codons identified within L2 ORF of the samples analyzed, were harbored by CaCx cases, while 8 out of 25 (32%) were harbored by HPV16 positive non-malignant samples (p = 3.87105E-07). L2 (mRNA and protein) expression was evident only among cases with episomal viral genomes and L2 mRNA expression correlated significantly with E2 gene copy numbers suggesting expression from all episomal genomes. Among such cases, Asian American (AA) isolates portrayed all of the humanized codons (100%; 4-6/sample) recorded within L2, which was significantly higher (p = 2.02E-7) compared to the European (E) isolates (22.8%; none or 1-2/sample). Additionally, majority of E variant isolates within cases (54/57; 94.7%) portrayed a variation (T4228C) within the short non-coding region (NCR2) between E5 and L2 genes, which portrays a weak promoter activity specific for L2 mRNA expression. This resulted in loss of 9 out of 14 miRNA binding sites (hsa-miR-548 family), despite the significant overexpression of miR548a-5p and miR548d-5p among such cases (28.64 and 36.25 folds, respectively), in comparison to HPV negative control samples. The findings exemplify the biological relevance of sequence variations in HPV16 genomes and highlight that episomal HPV16 in CaCx cases employ multiple mechanisms to sustain L2 expression, thereby justifying the potential role of L2 in such cancers, as opposed to those harboring viral integration.
authors.affiliation: National Institute of Biomedical Genomics, Kalyani, West Bengal, India.
authors.forename: Paramita
authors.initials: P
authors.lastname: Mandal
authors.forename: Bornali
authors.initials: B
authors.lastname: Bhattacharjee
authors.forename: Damayanti
authors.initials: D
authors.lastname: Das Ghosh
authors.forename: Nidhu Ranjan
authors.initials: NR
authors.lastname: Mondal
authors.forename: Rahul
authors.initials: R
authors.lastname: Roy Chowdhury
authors.forename: Sudipta
authors.initials: S
authors.lastname: Roy
authors.forename: Sharmila
authors.initials: S
authors.lastname: Sengupta
mesh_terms: D000328:Adult; D000368:Aged; D036022:Capsid Proteins / Q000235:genetics* / Q000378:metabolism; D002294:Carcinoma, Squamous Cell / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D005260:Female; D018628:Gene Dosage; D015967:Gene Expression Regulation, Viral; D016679:Genome, Viral; D054884:Host-Pathogen Interactions; D052162:Human papillomavirus 16 / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism; D008875:Middle Aged; D009856:Oncogene Proteins, Viral / Q000235:genetics* / Q000378:metabolism; D016366:Open Reading Frames; D030361:Papillomavirus Infections / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D010957:Plasmids; D011110:Polymorphism, Genetic; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D017422:Sequence Analysis, DNA; D002583:Uterine Cervical Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology
chemical_list: D036022:Capsid Proteins; C063316:L2 protein, Human papillomavirus type 16; C561887:MIRN548 microRNA, human; D035683:MicroRNAs; D009856:Oncogene Proteins, Viral; D012333:RNA, Messenger