Reporter gene transfer induces apoptosis in primary cortical neurons.
Modern cell biologists typically use reporter genes either alone or co-expressed with a protein of interest to facilitate the localization or quantification of protein expression. Our work demonstrates that reporter genes should be used cautiously, as several common reporter gene products are toxic to primary cortical neuronal cultures. We used the herpes simplex virus-based viral amplicon vector to transduce cortical neurons with three different reporter genes and assessed whether any reporter gene products were toxic over time, by monitoring neurite disintegration and apoptosis. Toxicity varied as a function of the reporter gene, the gene product localization, and the level of reporter gene expression. Transduction of enhanced green fluorescent protein or nuclear-localized beta-galactosidase was more toxic than non-nuclear localized beta-galactosidase. This work underscores the need for careful design of gene expression constructs. Moreover, in studies where cell injury or toxicity is being evaluated, their use should be carefully considered.
Detrait
Eric R
ER
Center for Aging and Developmental Biology, Department of Pathology and Laboratory Medicine, University of Rochester, Rochester, New York 14642, USA.
Bowers
William J
WJ
Halterman
Marc W
MW
Giuliano
Rita E
RE
Bennice
Lisa
L
Federoff
Howard J
HJ
Richfield
Eric K
EK
D000818:Animals; D017209:Apoptosis; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D018014:Gene Transfer Techniques; D017930:Genes, Reporter; D005822:Genetic Vectors / Q000378:metabolism; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000235:genetics / Q000633:toxicity*; D051379:Mice; D009474:Neurons / Q000166:cytology / Q000473:pathology*; D018139:Simplexvirus / Q000378:metabolism; D019076:Transgenes; D001616:beta-Galactosidase / Q000235:genetics / Q000633:toxicity*
D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins; D001616:beta-Galactosidase