Carboxy-fluorescein diacetate, succinimidyl ester labeled papillomavirus virus-like particles fluoresce after internalization and interact with heparan sulfate for binding and entry.
Human papillomaviruses (HPVs) infect epithelial cells and are associated with genital carcinoma. Most epithelial cell lines express cell-surface glycosaminoglycans (GAGs) usually found attached to the protein core of proteoglycans. Our aim was to study how GAGs influenced HPV entry. Using a human keratinocyte cell line (HaCaT), preincubation of HPV virus-like particles (VLPs) with GAGs showed a dose-dependent inhibition of binding. The IC(50) (50% inhibition) was only 0.5 microg/ml for heparin, 1 microg/ml for dextran sulfate, and 5-10 microg/ml for heparan sulfate from mucosal origin. Mutated chinese hamster ovary (CHO) cell lines lacking heparan sulfate or all GAGs were unable to bind HPV VLPs. Here we also report a method to study internalization by using VLPs labeled with carboxy-fluorescein diacetate, succinimidyl ester, a fluorochrome that is only activated after cell entry. Pretreatment of labeled HPV VLPs with heparin inhibited uptake, suggesting a primary interaction between HPV and cell-surface heparan sulfate.
Drobni
Peter
P
Department of Virology, Umeå University, SE-901 85 Umeå, Sweden.
Mistry
Nitesh
N
McMillan
Nigel
N
Evander
Magnus
M
D000818:Animals; D016466:CHO Cells; D002460:Cell Line; D006224:Cricetinae; D005452:Fluoresceins; D005453:Fluorescence; D006025:Glycosaminoglycans / Q000502:physiology; D006493:Heparin / Q000494:pharmacology; D019763:Heparin Lyase / Q000494:pharmacology; D006497:Heparan Sulfate / Q000502:physiology*; D006801:Humans; D015603:Keratinocytes / Q000821:virology; D027383:Papillomaviridae / Q000502:physiology*; D014771:Virion / Q000502:physiology*
D005452:Fluoresceins; D006025:Glycosaminoglycans; D006493:Heparin; D019763:Heparin Lyase; D006497:Heparan Sulfate; C018506:diacetylfluorescein