Permanent URL to this publication: http://dx.doi.org/10.5167/uzh-39494
Püntener, D; Engelke, M F; Ruzsics, Z; Strunze, S; Wilhelm, C; Greber, U F (2011). Stepwise loss of fluorescent core protein V from human adenovirus during entry into cells. Journal of Virology, 85(1):481-496.
View at publisher
Human adenoviruses (HAdVs, short Ad) replicate and assemble particles in the nucleus. They organise a linear double-strand DNA-genome into a condensed core with about 180 nucleosomes by the viral protein VII (pVII), pX, and pV attaching the DNA to the capsid. Using reverse genetics we generated a novel, non-conditionally replicating Ad reporter by inserting green fluorescent protein (GFP) at the amino-terminus of pV. Purified Ad2-GFP-pV virions had an oversized complete genome, and incorporated about 38 GFP-pV molecules per virion, about 25% of Ad2 pV. GFP-pV cofractionated with the DNA-core like pV, and newly synthesized GFP-pV had a subcellular localization indistinguishable from pV, indicating that GFP-pV is a valid reporter for pV. Ad2-GFP-pV completed the replication cycle, although at lower yields than Ad2. Incoming GFP-pV (or pV) was not imported into the nucleus. Virions lost GFP-pV at two points during the infection process, entry into the cytosol, and at the nuclear pore complex, where capsids disassemble. Disassembled capsids, positive for the conformation specific anti-hexon antibody R70, were devoid of GFP-pV. The loss of GFP-pV was reduced by the macrolide antibiotic leptomycin B (LMB), which blocks nuclear export and adenovirus attachment to the nuclear pore complex. LMB inhibited the appearance of R70 epitopes on Ad2 and Ad2-GFP-pV, indicating that the loss of GFP-pV from Ad2-GFP-pV is an authentic step in the adenovirus uncoating program. Ad2-GFP-pV is genetically complete, and hence enables detailed analyses of infection and spreading dynamics in cells and model organisms, or assessment of oncolytic adenoviral potential.
139 downloads since deposited on 15 Jan 2011
20 downloads since 12 months
|Item Type:||Journal Article, refereed, original work|
|Communities & Collections:||07 Faculty of Science > Institute of Molecular Life Sciences|
|Dewey Decimal Classification:||570 Life sciences; biology|
|Deposited On:||15 Jan 2011 16:01|
|Last Modified:||27 Nov 2013 16:19|
|Publisher:||American Society for Microbiology|
|Free access at:||PubMed ID. An embargo period may apply.|
Users (please log in): suggest update or correction for this item
Repository Staff Only: item control page