Genetic analyses in neurons and neural crest-derived post mitotic cells.
Cationic lipids are shown to be efficient DNA carriers for gene transfer into neurons and chromaffin cells. Chimeric genes containing the coding sequence for the bacterial gene chloramphenicol acetyl transferase (CAT), under various promoter sequences were used to estimate transfection efficiency and to analyse transcriptional mechanisms. Using a ubiquitously expressed construct (RSV-CAT), and an anti-CAT antibody to identify CAT positive cells transfection efficiency was found to be approximately 70% in cultures of hypothalamic neurons. Proenkephalin gene expression was studied in chromaffin cells by employing a chimeric gene containing the rat pro-enkephalin promoter fused to the CAT reporter gene. Using this model we show that the transfected gene is inducible by neurotransmitters and second messengers. The contribution of a specific kinase (protein kinase A, PKA) to proenkephalin gene regulation is analysed using this model system, and expression vectors coding for the catalytic subunit of PKA.
Boutillier
A L
AL
Laboratoire de Physiologie Générale, URA 1446 CNRS, Université Louis Pasteur, Strasbourg, France.
Barthel
F
F
Loeffler
J P
JP
Hassan
A
A
Demeneix
B A
BA
D000818:Animals; D002417:Cattle; D002642:Chick Embryo; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D004745:Enkephalins / Q000096:biosynthesis / Q000235:genetics; D004759:Enterochromaffin Cells / Q000502:physiology; D005786:Gene Expression Regulation; D006579:Heterozygote; D007150:Immunohistochemistry; D008938:Mitosis; D009432:Neural Crest / Q000166:cytology*; D009474:Neurons / Q000502:physiology*; D011494:Protein Kinases / Q000378:metabolism; D011498:Protein Precursors / Q000096:biosynthesis / Q000235:genetics; D014158:Transcription, Genetic; D014162:Transfection
D004745:Enkephalins; D011498:Protein Precursors; C029992:proenkephalin; D015500:Chloramphenicol O-Acetyltransferase; D011494:Protein Kinases