title: A transmembrane domain and GxxxG motifs within L2 are essential for papillomavirus infection.
abstract: During cellular invasion, human papillomavirus type 16 (HPV16) must transfer its viral genome (vDNA) across the endosomal membrane prior to its accumulation at nuclear PML bodies for the establishment of infection. After cellular uptake, the capsid likely undergoes pH-dependent disassembly within the endo-/lysosomal compartment, thereby exposing hidden domains in L2 that facilitate membrane penetration of L2/vDNA complexes. In an effort to identify regions of L2 that might physically interact with membranes, we have subjected the L2 sequence to multiple transmembrane (TM) domain prediction algorithms. Here, we describe a conserved TM domain within L2 (residues 45 to 67) and investigate its role in HPV16 infection. In vitro, the predicted TM domain adopts an alpha-helical structure in lipid environments and can function as a real TM domain, although not as efficiently as the bona fide TM domain of PDGFR. An L2 double point mutant renders the TM domain nonfunctional and blocks HPV16 infection by preventing endosomal translocation of vDNA. The TM domain contains three highly conserved GxxxG motifs. These motifs can facilitate homotypic and heterotypic interactions between TM helices, activities that may be important for vDNA translocation. Disruption of some of these GxxxG motifs resulted in noninfectious viruses, indicating a critical role in infection. Using a ToxR-based homo-oligomerization assay, we show a propensity for this TM domain to self-associate in a GxxxG-dependent manner. These data suggest an important role for the self-associating L2 TM domain and the conserved GxxxG motifs in the transfer of vDNA across the endo-/lysosomal membrane.
authors.affiliation: Department of Immunobiology, The University of Arizona, Tucson, Arizona, USA.
authors.forename: Matthew P
authors.initials: MP
authors.lastname: Bronnimann
authors.forename: Janice A
authors.initials: JA
authors.lastname: Chapman
authors.forename: Chad K
authors.initials: CK
authors.lastname: Park
authors.forename: Samuel K
authors.initials: SK
authors.lastname: Campos
mesh_terms: D020816:Amino Acid Motifs; D001692:Biological Transport; D036022:Capsid Proteins / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D017124:Conserved Sequence; D004252:DNA Mutational Analysis; D004279:DNA, Viral / Q000378:metabolism*; D052162:Human papillomavirus 16 / Q000235:genetics / Q000502:physiology*; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D020125:Mutation, Missense; D009856:Oncogene Proteins, Viral / Q000235:genetics / Q000378:metabolism*; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D037521:Virulence Factors / Q000235:genetics / Q000378:metabolism*; D014779:Virus Replication; D057646:Virus Uncoating
chemical_list: D036022:Capsid Proteins; D004279:DNA, Viral; C063316:L2 protein, Human papillomavirus type 16; D008565:Membrane Proteins; D009856:Oncogene Proteins, Viral; D037521:Virulence Factors