Identification of G-quadruplex forming sequences in three manatee papillomaviruses

Abstract

The Florida manatee (Trichechus manatus latirotris) is a threatened aquatic mammal in United States coastal waters. Over the past decade, the appearance of papillomavirus-induced lesions and viral papillomatosis in manatees has been a concern for those involved in the management and rehabilitation of this species. To date, three manatee papillomaviruses (TmPVs) have been identified in Florida manatees, one forming cutaneous lesions (TmPV1) and two forming genital lesions (TmPV3 and TmPV4). We identified DNA sequences with the potential to form G-quadruplex structures (G4) across the three genomes. G4 were located on both DNA strands and across coding and non-coding regions on all TmPVs, offering multiple targets for viral control. Although G4 have been identified in several viral genomes, including human PVs, most research has focused on canonical structures comprised of three G-tetrads. In contrast, the vast majority of sequences we identified would allow the formation of non-canonical structures with only two G-tetrads. Our biophysical analysis confirmed the formation of G4 with parallel topology in three such sequences from the E2 region. Two of the structures appear comprised of multiple stacked two G-tetrad structures, perhaps serving to increase structural stability. Computational analysis demonstrated enrichment of G4 sequences on all TmPVs on the reverse strand in the E2/E4 region and on both strands in the L2 region. Several G4 sequences occurred at similar regional locations on all PVs, most notably on the reverse strand in the E2 region. In other cases, G4 were identified at similar regional locations only on PVs forming genital lesions. On all TmPVs, G4 sequences were located in the non-coding region near putative E2 binding sites. Together, these findings suggest that G4 are possible regulatory elements in TmPVs.

Emerging viruses in marine mammals.

“…In all TmPVs, G4 sequences were located in the non-coding region near putative E2 binding sites. Based on the role of G4 in PVs of humans, these findings suggest that G4 in TmPVs are possible regulatory elements of protein synthesis [32].…”

“…TmPV1 is widely dispersed among manatees and genetically similar to close-to-root forming verruciform to papular cutaneous papillomas [29,31] while not much is known about TmPV2 [30]. The two mucosotropic PVs form sessile mucosal genital papillomas (TmPV3 and TmPV4) [21,30,32]. Both TmPV3 and TmPV4 are similar in size and in genomic characterization of all PVs, with one non-coding region and seven open reading frames (ORFs) including the E7 ORF which is absent in cetacean PVs [32].…”

“…The two mucosotropic PVs form sessile mucosal genital papillomas (TmPV3 and TmPV4) [21,30,32]. Both TmPV3 and TmPV4 are similar in size and in genomic characterization of all PVs, with one non-coding region and seven open reading frames (ORFs) including the E7 ORF which is absent in cetacean PVs [32]. Alternatively, TmPV3 and TmPV4 are the first known genital mucosotropic PVs in manatees presenting a suspected novel sexually transmitted viral disease in this species similar to the emerging PV-associated genital lesions in bottlenose dolphins (see below).…”

G-Quadruplexes: More Than Just a Kink in Microbial Genomes

“…Given the functional significance of repeat sequences in microbial genomes it may be interesting to investigate the link between the tandem array of G4s and molecular processes related to microbial pathogenesis. Recent reports on G4 motifs in viruses infecting nonhuman hosts shed light on how G4s have been exploited by viruses for virulence and genome regulation throughout evolution [52,98].…”

“…Tluckova et al identified the presence of three-tetrad G4 motifs in the long control region (LCR) and in the coding regions of E1, E2/E4, and L2 proteins of eight HPV types [51]. Interestingly, two-tetrad quadruplexes were identified in the same genomic regions (i.e., E1, E2/E4 and LCR) of manatee papilloma viruses [52]. In papilloma viruses, the LCR contains a number of cis-acting regulatory elements for virus replication and transcription that play a role in determining tissue tropism [53].…”