Rat brain creatine kinase messenger RNA levels are high in primary cultures of brain astrocytes and oligodendrocytes and low in neurons.
Rat brain creatine kinase (CKB) gene expression is highest in the brain but is also detectable at lower levels in some other tissues. In the brain, the CKB enzyme is thought to be involved in the regeneration of ATP necessary for transport of ions and neurotransmitters. To understand the molecular events that lead to high CKB expression in the brain, we have determined the steady-state levels of CKB mRNA in homogeneous cultures of primary rat brain astrocytes, oligodendrocytes, and neurons. Northern blot analysis showed that whereas the 1.4-kb CKB mRNA was detectable in neurons, the level was about 17-fold higher in oligodendrocytes and 15-fold higher in astrocytes. The blots were hybridized with a CKB-specific 32P-antisense RNA probe, complementary to the 3' untranslated sequence of CKB, which hybridizes to CKB mRNA but not CKM mRNA. Also, the 5' and 3' ends of CKB mRNA from the glial cells were mapped, using exon-specific antisense probes in the RNase-protection assay, and were found to be the same in astrocytes and oligodendrocytes. This indicated that (a) the site of in vivo transcription initiation in astrocytes and oligodendrocytes was directed exclusively by the downstream, nonconcensus TTAA sequence at -25 bp in the CKB promoter that is also utilized by all other cell types that express CKB and (b) the 3' end of mature CKB mRNA was the same in astrocytes and oligodendrocytes. In addition, there was no detectable alternate splicing in exon 1, 2, or 8 of CKB mRNA in rat astrocytes and oligodendrocytes.(ABSTRACT TRUNCATED AT 250 WORDS)
Molloy
G R
GR
School of Life and Health Sciences, University of Delaware, Newark 19716.
Wilson
C D
CD
Benfield
P
P
de Vellis
J
J
Kumar
S
S
D000818:Animals; D001253:Astrocytes / Q000201:enzymology* / Q000378:metabolism; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000201:enzymology* / Q000378:metabolism; D003402:Creatine Kinase / Q000235:genetics*; D005786:Gene Expression Regulation; D009474:Neurons / Q000201:enzymology*; D009836:Oligodendroglia / Q000201:enzymology* / Q000378:metabolism; D012333:RNA, Messenger / Q000096:biosynthesis / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar
D012333:RNA, Messenger; D003402:Creatine Kinase