Transient expression of virus-specific promoters in murine resident peritoneal macrophages.
Monocyte-macrophages (MO), being non-permissive for most viruses, play an important role in resistance to virus infection. In order to establish the mechanism of abortive infection of murine resident peritoneal MO (ResPMO) by herpes simplex virus type 1 (HSV-1), it is desirable to transfect these cells with viral promoters linked to an assayable gene, for example, the bacterial chloramphenicol acetyl transferase (CAT) gene. This will facilitate studies designed to measure levels of promoter activation or repression in these specialized cells. Transient expression of CAT in ResPMO was achieved with DEAE-dextran, but not using either calcium phosphate precipitate or lipofectin. CAT expression driven by various virus-specific promoters was less efficient in ResPMO compared with Vero cells and approximately 50% of input plasmid DNA remained in Vero cells at 48 h post transfection, but only 9% was detectable in ResPMO. However, approximately 6% of ResPMO and 9% of Vero cells contained CAT-specific DNA at 24 h post transfection. In addition, 2% of cells of either cell type contained CAT-specific polypeptide at 48 h. This is therefore the first report that the non-replicating murine ResPMO can be transfected in vitro and more importantly, that these cells express the transfected gene products.
Department of Microbiology and Immunology, Medical College of Pennsylvania, Philadelphia 19129.
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Wu
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Anaraki
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Morahan
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Leary
D000818:Animals; D002130:Calcium Phosphates; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics / Q000378:metabolism; D003637:DEAE-Dextran; D004247:DNA / Q000032:analysis / Q000235:genetics; D005260:Female; D005455:Fluorescent Antibody Technique; D015870:Gene Expression / Q000502:physiology*; D005814:Genes, Viral / Q000235:genetics; D006561:Herpes Simplex / Q000276:immunology; D007113:Immunity, Innate / Q000235:genetics*; D008264:Macrophages / Q000201:enzymology / Q000378:metabolism / Q000502:physiology*; D051379:Mice; D009000:Monocytes / Q000201:enzymology / Q000378:metabolism / Q000502:physiology; D010529:Peritoneal Cavity / Q000166:cytology*; D011401:Promoter Regions, Genetic / Q000502:physiology*; D014162:Transfection
D002130:Calcium Phosphates; C485828:alpha-tricalcium phosphate; C485830:tetracalcium phosphate; C485838:calcium phosphate, monobasic, anhydrous; D004247:DNA; D003637:DEAE-Dextran; C020243:calcium phosphate; D015500:Chloramphenicol O-Acetyltransferase; C485829:calcium phosphate, dibasic, anhydrous