GAP-43 mRNA suppression by the ribozyme in PC12 cells and inhibition of evoked dopamine release.
We constructed a ribozyme designed to cleave the GAP-43 mRNA and which contained a bacteriophage T7 transcription terminator at its 3' site to maintain stability. The ribozyme was overexpressed in PC12 cells by pEF-BOS, a powerful mammalian expression vector. Consequently, PC12 cells overexpressing the GAP-43 ribozyme revealed a drastic decrease in the levels of GAP-43 mRNA expression, and the evoked dopamine release was significantly suppressed in these cell lines. These results support the previous observations that GAP-43 is associated with Ca-dependent dopamine release in PC12 cells, and the ribozyme expression system used in the present study was demonstrated to be useful for suppression of the functions of specific mRNAs and exploration of specific gene products.
Imaizumi
K
K
Department of Molecular Neurobiology, TANABE, Osaka University Medical School, Japan.
Katoh
T
T
Tsuda
M
M
Takagi
T
T
Kiyama
H
H
D000818:Animals; D015152:Blotting, Northern; D004298:Dopamine / Q000378:metabolism*; D019922:GAP-43 Protein; D015870:Gene Expression; D008562:Membrane Glycoproteins / Q000235:genetics*; D009419:Nerve Tissue Proteins / Q000235:genetics*; D016900:Neurofilament Proteins / Q000235:genetics*; D016716:PC12 Cells / Q000187:drug effects*; D011188:Potassium / Q000494:pharmacology; D016337:RNA, Catalytic / Q000494:pharmacology*; D012333:RNA, Messenger / Q000096:biosynthesis*; D051381:Rats; D014158:Transcription, Genetic
D019922:GAP-43 Protein; D008562:Membrane Glycoproteins; D009419:Nerve Tissue Proteins; D016900:Neurofilament Proteins; D016337:RNA, Catalytic; D012333:RNA, Messenger; D011188:Potassium; D004298:Dopamine