Application of a rapid method (targeted display) for the identification of differentially expressed mRNAs following NGF-induced neuronal differentiation in PC12 cells.
Nerve growth factor (NGF)-induced differentiation of the rat pheochromocytoma, PC12, cell line presents a model system for the study of early gene expression changes involved in neuronal differentiation. Rapid alterations in mRNA expression patterns were investigated in PC12 cells following exposure to NGF using a set of statistically designed primers that exhibit coding-strand bias, and the products were analyzed on agarose gels. This simple and rapid method (targeted display) generated reproducible expression profiles, indicating a complex pattern of gene regulation, and resulted in the identification of a number of NGF-regulated transcripts. Thirty-two of these were selected at random and sequenced, revealing 19 known and 13 novel genes (or ESTs). Northern blot analysis and RT-PCR confirmed the differential regulation of 22 genes (16 known, 6 novel) and demonstrated 1 false positive result. Antisense application of one isolated gene product, the serine/threonine kinase MARK1, prevented neuronal differentiation in transiently transfected PC12 cells.
Brown
A J
AJ
Department of Biochemistry, University of Sussex, Falmer, Brighton, BN1 9RY, United Kingdom.
Hutchings
C
C
Burke
J F
JF
Mayne
L V
LV
D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects; D018076:DNA, Complementary / Q000235:genetics; D004587:Electrophoresis, Agar Gel / Q000379:methods; D005346:Fibroblast Growth Factors / Q000494:pharmacology; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects*; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D051379:Mice; D009414:Nerve Growth Factors / Q000494:pharmacology*; D016716:PC12 Cells / Q000187:drug effects* / Q000378:metabolism; D016133:Polymerase Chain Reaction / Q000379:methods*; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000502:physiology; D012333:RNA, Messenger / Q000096:biosynthesis*; D012334:RNA, Neoplasm / Q000096:biosynthesis; D051381:Rats; D012689:Sequence Homology, Nucleic Acid; D014158:Transcription, Genetic; D014162:Transfection
D018076:DNA, Complementary; D009414:Nerve Growth Factors; D012333:RNA, Messenger; D012334:RNA, Neoplasm; D005346:Fibroblast Growth Factors; C484552:Mark1 protein, rat; D017346:Protein Serine-Threonine Kinases