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Human papillomavirus tropism: determinants of viral tissue specificity
Umeå University, Faculty of Medicine, Clinical Microbiology.
2007 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Cervical cancer is the second most common cancer among women worldwide and human papillomavirus (HPV) is a prerequisit for the development of this cancer. HPV belongs to the Papillomaviridae family and infects the basal layer of epithelial cells where it generally progresses into warts or condylomas. HPV can only reproduce in differentiating epithelia and it is therefore difficult to study the natural infection of HPV. More than 100 HPV types exist and they are divided into different genera based on their L1 open reading frame sequence. Most of the HPV types in the alpha-papillomavirus genus infect the mucosal epithelium while HPVs from the beta-papillomavirus genus usually infect cutaneous epithelial cells. Presently, it is not known what decides the anatomical tropism and our aim was to study determinants of this tropism.

By using HPV virus like particles (VLP) and pseudovirus we found that VLPs from the two alpha-papillomaviruses HPV-6 and HPV-16 interacted with cell-surface heparan sulfate (HS) for initial attachment. When we labelled HPV VLPs with a fluorescent dye to study internalization HPV-6 was more strongly inhibited than HPV-16. Furthermore, a pseudovirus infection assay demonstrated that the beta-papillomavirus HPV-5 was less dependent on HS for infection than HPV-16. By analyzing the isoelectric point (p1) of the HPV L1 capsid protein we found that alpha HPV types were more positively charged than beta HPV types. Also, HPV-6 had a higher positive charge than HPV-16. Thus, the inhibition of the negatively charged heparin against HPV infection was clearly related to the charge of the HPV L1 capsid. This suggested that the initial interaction could be one of the determinants of tropism although not the sole factor.

Lactoferrin is a protein found in milk, saliva, semen, tear fluid and endocervical secretions that has antiviral activities. Both human and bovine lactoferrin inhibited HPV infection but we found no significant differences in inhibition of alpha- and beta-papillomavirus infection. We could however demonstrate that different lactoferricins, small peptide derivates from the N-terminal part of lactoferrin, were able to inhibit HPV infection. This antiviral activity depended on lactoferricin peptide, HPV type and cell origin.

The regulation of HPV gene expression in the host cell could also determine HPV tropism. The HPV long control region (LCR) contains cis-responsive elements that regulate HPV transcription and the epithelial tropism of HPV is determined by epithelial specific constitutive enhancers in the LCR. It has been hypothesized that the combination of transcription factors in the host cell determines the cell-type-specific expression. In cells with a skin origin the HPV-5 LCR was twice as efficient in transcriptional activation compared to HPV-16 LCR, while in cervical cells the HPV-16 LCR was almost twice as effective in activating transcription compared to HPV-5 LCR.

To conclude, alpha- and beta-papillomaviruses differed regarding their ability to infect cells and regulate viral gene expression. These abilities corresponded with their natural host cells and suggested that HPV anatomical tropism could be determined at several steps in the HPV life cycle.

Place, publisher, year, edition, pages
Umeå: Klinisk mikrobiologi , 2007. , 57 p.
Series
Umeå University medical dissertations, ISSN 0346-6612 ; 1105
Keyword [en]
papillomavirus, receptor, heparan sulfate, pI, lactoferrin, lactoferricin, transcription, long control region, tropism
National Category
Microbiology in the medical area
Identifiers
URN: urn:nbn:se:umu:diva-1149ISBN: 978-91-7264-332-1 (print)OAI: oai:DiVA.org:umu-1149DiVA: diva2:140375
Public defence
2007-06-05, Astrid Fagraeus Salen, 6A 103, Umeå Universitet, Umeå, 09:00 (English)
Opponent
Supervisors
Available from: 2007-05-15 Created: 2007-05-15 Last updated: 2009-06-15Bibliographically approved
List of papers
1. Carboxy-fluorescein diacetate, succinimidyl ester labeled papillomavirus virus-like particles fluoresce after internalization and interact with heparan sulfate for binding and entry
Open this publication in new window or tab >>Carboxy-fluorescein diacetate, succinimidyl ester labeled papillomavirus virus-like particles fluoresce after internalization and interact with heparan sulfate for binding and entry
2003 (English)In: Virology, ISSN 0042-6822, E-ISSN 1096-0341, Vol. 310, no 1, 163-172 p.Article in journal (Refereed) Published
Abstract [en]

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.

Keyword
Papillomavirus; CFDA SE; Heparan sulfate; Uptake, Receptor
Identifiers
urn:nbn:se:umu:diva-2394 (URN)10.1016/S0042-6822(03)00114-4 (DOI)12788640 (PubMedID)
Available from: 2007-05-15 Created: 2007-05-15 Last updated: 2017-12-14Bibliographically approved
2. Entry of human Papillomavirus type 5 is only partly dependent of a heparan sulfate receptor
Open this publication in new window or tab >>Entry of human Papillomavirus type 5 is only partly dependent of a heparan sulfate receptor
(English)Manuscript (Other academic)
Identifiers
urn:nbn:se:umu:diva-2395 (URN)
Available from: 2007-05-15 Created: 2007-05-15 Last updated: 2011-04-05Bibliographically approved
3. The anti-papillomavirus activity of human and bovine lactoferricin.
Open this publication in new window or tab >>The anti-papillomavirus activity of human and bovine lactoferricin.
Show others...
2007 (English)In: Antiviral Research, ISSN 0166-3542, E-ISSN 1872-9096, Vol. 75, no 3, 258-265 p.Article in journal (Refereed) Published
Abstract [en]

Human papillomavirus (HPV) cause common warts, laryngeal papilloma and genital condylomata and is necessary for the development of cervical cancer. We have previously found that lactoferrin has antiviral activity against HPV-16 and others have demonstrated that lactoferricin, an N-terminal fragment of lactoferrin, has inhibitory activities against several viruses. Two cell lines and two virus types, HPV-5 and HPV-16, were used to study if lactoferrin and lactoferricin could inhibit HPV pseudovirus (PsV) infection. We demonstrated that bovine lactoferrin (bLf) and human lactoferrin (hLf) were both potent inhibitors of HPV-5 and -16 PsV infections. Among the four lactoferricin derivatives we analyzed, a 15 amino acid peptide from bovine lactoferricin (bLfcin) 17-31 was the most potent inhibitor of both HPV-5 and HPV-16 PsV infection. Among the other derivatives, the human lactoferricin (hLfcin) 1-49 showed some antiviral activity against HPV PsV infection while bLfcin 17-42 inhibited only HPV-5 PsV infection in one of the cell lines. When we studied initial attachment of HPV-16, only bLfcin 17-42 and hLfcin 1-49 had an antiviral effect. This is the first time that lactoferricin was demonstrated to have an inhibitory effect on HPV infection and the antiviral activity differed depending on size, charge and structures of the lactoferricin.

Identifiers
urn:nbn:se:umu:diva-20708 (URN)10.1016/j.antiviral.2007.03.012 (DOI)17481742 (PubMedID)
Available from: 2009-03-24 Created: 2009-03-24 Last updated: 2017-12-13
4. Transcriptional activation of the human papillomavirus type 5 and 16 long control region in cells from cutaneous and mucosal origin
Open this publication in new window or tab >>Transcriptional activation of the human papillomavirus type 5 and 16 long control region in cells from cutaneous and mucosal origin
2007 (English)In: Virology journal, ISSN 1743-422X, Vol. 4, 4:27- p.Article in journal (Refereed) Published
Identifiers
urn:nbn:se:umu:diva-2397 (URN)10.1186/1743-422X-4-27 (DOI)17352804 (PubMedID)
Available from: 2007-05-15 Created: 2007-05-15 Last updated: 2011-04-05Bibliographically approved

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