Open AccessCell cycle analysis of retroviral vector gene expression during early infection
Author(s) -
Sladek Todd L.,
Jacobberger James W.
Publication year - 1998
Publication title -
cytometry
Language(s) - English
Resource type - Journals
eISSN - 1097-0320
pISSN - 0196-4763
DOI - 10.1002/(sici)1097-0320(19980401)31:4<235::aid-cyto2>3.0.co;2-i
Subject(s) - biology , cell cycle , provirus , population , propidium iodide , microbiology and biotechnology , flow cytometry , virology , viral vector , long terminal repeat , murine leukemia virus , immunofluorescence , gene expression , virus , vector (molecular biology) , gene , genetics , apoptosis , antibody , recombinant dna , programmed cell death , genome , demography , sociology
To examine the pattern of retroviral vector gene expression during early stages of infection, a Moloney murine leukemia virus (M‐MuLV)‐based vector that transcribes the simian virus 40 (SV40) large T antigen (Tag) gene from the viral long terminal repeat (LTR) was used to infect proliferating rodent fibroblasts. At various times after infection, cells were fixed and stained for Tag by indirect immunofluorescence, and for DNA using propidium iodide. Tag immunofluorescence and DNA content were both quantified by flow cytometry. The results showed that Tag expression was first detected exclusively in the late G 1 and early S phases of the cell cycle, approximately 12 h postinfection. The infection was synchronous in that Tag‐expressing cells detected at 12 h, in late G 1 and early S, moved as a discrete population through S phase, into the G 2 + M phases of the cell cycle and then back into G 1 during the next 8–10 h. The presence of a synchronous Tag‐expressing cell population suggests that, at time of infection, cells in certain phases of the cell cycle were more susceptible to infection than cells in other phases. This may be related to synchronizing events that must occur before viral genes are expressed in infected cells; one such event may be integration of viral DNA into cellular chromosomes (i.e., provirus formation) that requires cells to pass through an M phase. Cytometry 31:235–241, 1998. © 1998 Wiley‐Liss, Inc.