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Fiber swap between adenovirus subgroups B and C alters intracellular trafficking of adenovirus gene transfer vectors.

TitleFiber swap between adenovirus subgroups B and C alters intracellular trafficking of adenovirus gene transfer vectors.
Publication TypeJournal Article
Year of Publication1999
AuthorsMiyazawa N, Leopold PL, Hackett NR, Ferris B, Worgall S, Falck-Pedersen E, Crystal RG
JournalJ Virol
Volume73
Issue7
Pagination6056-65
Date Published1999 Jul
ISSN0022-538X
KeywordsAdenoviruses, Human, Biological Transport, Capsid, Capsid Proteins, Carbocyanines, Cell Nucleus, DNA, Viral, Fluorescent Dyes, Genes, Viral, Genetic Vectors, Genome, Viral, Humans, Intracellular Fluid, Kinetics, Serotyping, Tumor Cells, Cultured
Abstract

Following receptor binding and internalization, intracellular trafficking of adenovirus (Ad) among subgroups B and C is different, with significant amounts of Ad serotype 7 (Ad7) (subgroup B) virions found in cytoplasm during the initial hours of infection while Ad5 (subgroup C) virions rapidly translocate to the nucleus. To evaluate the role of the fiber in these differences, we examined intracellular trafficking of Ad5, Ad7, and Ad5f7 (a chimeric vector composed of the Ad5 capsid with the fiber replaced by the Ad7 fiber) by conjugating Ad capsids directly with Cy3 fluorescent dye, permitting the trafficking of the capsids to be examined by fluorescence microscopy. The human lung carcinoma cell line A549 was infected with Cy3-conjugated viruses for 10 min followed by a 1-h incubation. Ad5 virions rapidly translocated to the nucleus (within 1 h of infection), while Ad7 virions were widely distributed in the cytoplasm at the same time point. Interestingly, chimeric Ad5f7 virions behaved similarly to Ad7 but not Ad5. In this regard, the percentages of nuclear localization of Ad5, Ad7, and Ad5f7 at 1 h following infection were 72% +/- 4%, 32% +/- 6%, and 38% +/- 2%, respectively. Consistent with these observations, fluorescence in situ hybridization demonstrated that most of the Ad5 DNA was detected at the nucleus after 1 h, but at the same time point, DNA of Ad7 and Ad5f7 was distributed in both the nucleus and cytoplasm. Quantification of the kinetics of Ad genomic DNA delivery to the nucleus using a fluorogenic probe-based PCR assay (TaqMan PCR) demonstrated that the percentages of nuclear association of Ad5 DNA and Ad5f7 DNA at 1 h postinfection were 80% +/- 13% and 43% +/- 1%, respectively. Although it has been generally accepted that Ad fiber protein mediates attachment of virions to cells and that fibers dissociate during endocytic uptake, these data suggest that in addition to mediating binding to the cell surface, fiber likely modulates intracellular trafficking as well.

DOI10.1128/JVI.73.7.6056-6065.1999
Alternate JournalJ Virol
PubMed ID10364358
PubMed Central IDPMC112667
Grant ListP01 HL051746 / HL / NHLBI NIH HHS / United States
R29 AI-42250 / AI / NIAID NIH HHS / United States
P01 HL059312 / HL / NHLBI NIH HHS / United States
P01 HL59312 / HL / NHLBI NIH HHS / United States
P01 HL51746 / HL / NHLBI NIH HHS / United States

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