Conditional gene expression systems to study herpesvirus biology in vivo.
Cytomegalovirus (CMV), a prototypic beta-herpesvirus, is an important human pathogen causing protean clinical manifestations in immature and immunocompromised patients. Mechanisms of infection can be studied in a mouse model. Mouse cytomegalovirus (MCMV) resembles in pathogenesis its human counterpart in many ways. Although MCMV infection is studied extensively on the level of organs, the contribution of specific cell types to viral replication in vivo is still elusive. Here we describe our approach based on the the Cre/loxP-system to investigate MCMV infection at the level of cell types in vivo. Using bacterial artificial chromosome (BAC)-technology, we created an MCMV virus containing an enhanced green fluorescent protein (egfp) reporter-gene which is not expressed due to a 'Stop' cassette flanked by two loxP-sites between promoter and coding sequence. Infection of cre-transgenic mice with this reporter virus results in the deletion of the 'Stop' cassette and expression of EGFP in a cell type-specific manner. Using this conditional gene expression system we are able to quantify viral productivity in specific cell types and to determine their contribution to viral dissemination in vivo. Furthermore, the deletion of viral genes can be used to screen for cell type-specificity of viral gene functions. Hence, conditional MCMV mutants allow the study of herpesvirus biology on the level of cell types in vivo.
Sacher
Torsten
T
Max von Pettenkofer-Institute, Ludwig Maximilians-University, Pettenkoferstrasse 9a, 80336 Munich, Germany.
Jordan
Stefan
S
Mohr
Christian A
CA
Vidy
Aurore
A
Weyn
Annelies M G
AM
Ruszics
Zsolt
Z
Koszinowski
Ulrich H
UH
D000818:Animals; D022202:Chromosomes, Artificial, Bacterial; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006566:Herpesviridae Infections / Q000821:virology*; D051379:Mice; D008822:Mice, Transgenic; D008967:Molecular Biology / Q000379:methods*; D018146:Muromegalovirus / Q000235:genetics / Q000254:growth & development*; D011995:Recombination, Genetic
C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins