Nervous and nonnervous cell transduction by recombinant adenoviruses that inducibly express the human PrP.
The study of the prion protein (PrP) physiological functions or its specific role in transmissible spongiform encephalopathies (TSE) requires new tools, particularly those able to induce PrP overexpression in a large range of cells, in vivo as well as in vitro. Here we describe the construction of two recombinant adenoviruses encoding the human PrP either with a valine at position 129 (AdTRVal) or a methionine (AdTRMet). Both genes were put under the control of the tetracycline-responsive promoter, allowing tight regulation of PrP expression. AdTRVal and AdTRMet induced high expression of the human PrP in CHO-KI cells and in organotypic brain slices in culture. The proteins expressed from these viruses exhibited a glycosylphosphatidyl inositol (GPI) anchor, proper glycosylation and sensitivity to proteinase K digestion. AdTRVal and AdTRMet will allow future studies on the human PrP and on the role of the codon 129 polyphormism in human TSE.
Arrabal
S
S
CEA, Service de Neurovirologie, BP 6 92265 Fontenay-aux-Roses Cedex, France.
Touchard
M
M
Mouthon
F
F
Klonjkowski
B
B
Deslys
J P
JP
Dormont
D
D
Eloit
M
M
D000256:Adenoviridae / Q000235:genetics*; D019943:Amino Acid Substitution; D000818:Animals; D016466:CHO Cells / Q000166:cytology / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002531:Cerebellum / Q000378:metabolism; D006224:Cricetinae; D004318:Doxycycline / Q000494:pharmacology; D005434:Flow Cytometry; D005786:Gene Expression Regulation / Q000187:drug effects; D017930:Genes, Reporter; D005822:Genetic Vectors / Q000235:genetics / Q000378:metabolism; D006624:Hippocampus / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D051379:Mice; D018345:Mice, Knockout; D016297:Mutagenesis, Site-Directed; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D010957:Plasmids / Q000235:genetics; D011328:Prions / Q000096:biosynthesis* / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000187:drug effects / Q000235:genetics; D012333:RNA, Messenger / Q000096:biosynthesis; D013752:Tetracycline / Q000494:pharmacology; D014161:Transduction, Genetic / Q000379:methods*
D011328:Prions; D012333:RNA, Messenger; D013752:Tetracycline; D004318:Doxycycline