Human papillomavirus genomics: past, present and future

Abstract

Human papillomaviruses (HPV) are a group of divergent DNA viruses, of which a select few evolutionarily related HPVs have emerged to be highly oncogenic and of significant medical importance. Essentially all cases of cervical cancer, as well as a subset of other anogenital and oral cancers are caused by this limited set of HPV types. At present, over 150 HPV types have been identified and may be classified into genera, species and types based upon comparison of the viral genome. Established nucleotide phylogenies sort the highly pathogenic HPV types to the genus Alphapapillomavirus (α-PV). A species group includes viral types with 60-70% genomic nucleotide similarity that share a most-recent common ancestor; for example the species group's alpha-9 (HPV16-related) and alpha-7 (HPV18-related), contain the majority of known oncogenic HPV types. Genomes from the same HPV type with 1-10% nucleotide differences designate HPV variant lineages. The established nucleotide variations observed in extant HPV genomes have been fixed through evolutionary processes prior to human population expansion and global dissemination. To characterize viral types and variants associated with pathology for clinical applications (e.g. screening), molecular epidemiological studies have proven essential for identifying links between HPV natural history and carcinogenicity. This chapter presents a historical account of HPV genomics in the context of major discoveries and advances over the past 2 thousand years.

Figure 1. History of HPV, Cervix Cancer and Technological Advances

The timeline displays the landmarks in Papillomavirus (PV) biology and clinical discovery. The color of the boxes serves to distinguish events and/or discoveries relating to basic PV scientific advancements in red, biological discoveries/technology innovations in blue, or epidemiology/public health in green.

Figure 2. Schematic of HPV16R Genome

The HPV16 genome displayed shows the general organization of the HPV open reading frames (ORFs) and regulatory regions. Early (E) genes and related transcriptional regulatory motifs are shown in green; late (L) genes and related transcriptional regulatory are shown in blue; and the 2 noncoding regions are shown in grey, the smaller region between E5 and L2 is termed the NCR (non-coding region), and the larger region between L1 and E6 is known as the URR (Upstream Regulatory Region). The URR contains regulatory elements including the viral DNA origin of replication (Ori) depicted by a yellow oval and the early gene promoter, p97 depicted by a green arrow. The blue arrow in the E7 ORF depicts late gene expression regulated by the late promoter, p670. Polyadenylation sites termed early (pAE) or late (pAL) are shown as green or blue triangles, respectively. Arrowheads at the tip of each ORF depict their overlap. The thick solid line represents genomic regions commonly overexpressed in cancer, the thin black line represents genomic regions frequently disrupted by integration, and the dashed line represents the genomic region rarely expressed in cancer.

Figure 3. Human papillomavirus phylogenetic tree clustering the majority of HPVs into three genera, Alphapapillomavirus, Betapapillomavirus and Gammapapillomavirus

A maximum likelihood (ML) tree was constructed using RAxML v7.2.8.27 [77] and an alignment of the nucleotide sequences of the L1 open reading frame (ORF) of 120 published HPV types. HPV species groups were generally classified according to the classification system for PVs by Bernard and colleagues [18]. All high-risk mucosal HPV types cluster within the genus Alphapapillomavirus, highlighted in red. The clades in blue represent mucosal HPV types with low risk or no risk to cervical cancer. The scale bar represents nucleotide change of 0.2 per site.

Figure 4. HPV16 variant tree topology and pairwise comparisons of individual complete genomes

A maximum likelihood (ML) tree was inferred from a global alignment of 13 complete nucleotide sequence genomes of HPV16 using RAxML v7.2.8 [77]. Distinct variant lineages (i.e., termed A/B/C/D) and sublineages (e.g., termed A1/A2/A3/A4) were classified according to the topology and nucleotide sequence differences from > 1% to < 10%, and > 0.5% to < 1%, respectively [18,26]. The percent nucleotide sequence differences were calculated for each isolate compared to all other isolates based on the nucleotide sequences (complete genome) and are shown in the panel to the right of the phylogeny. Values for comparison from an isolate are connected by lines and the comparison to self is indicated by the 0% difference point. Symbols and colored lines are used to distinguish each isolate. The scale bar at the bottom of the tree represents nucleotide change of 0.002 per site. Multiple isolates for the HPV16 A4 sublineage are shown to highlight the intra-sublineage relationship, note the clustering in the right hand graph that depicts nucleotide sequence differences.