The effect of cycloheximide on the regulation of proenkephalin and prodynorphin gene expressions induced by kainic acid in rat hippocampus.
The effect of cycloheximide (CHX), a protein synthesis inhibitor, on the regulation of proenkephalin (proENK) and prodynorphin (proDYN) mRNA levels, proto-oncogenes, such as c-fos, 35-kDa fra and c-jun mRNA, and the levels of their products induced by kainic acid (KA) in rat hippocampus was studied. The proENK and proDYN mRNA levels were markedly increased 4 and 8 h after KA (10 mg/kg i.p.) administration. However, the intracellular proENK protein level was not affected by KA. The elevations of both proENK and proDYN mRNA levels induced by KA were inhibited by pre-administration of CHX (15 mg/kg i.p.). The increases of proENK and proDYN mRNA levels induced by KA were well-correlated with the increases of c-Fos, 35-kDa Fra and c-Jun protein levels. KA administration increased the hippocampal levels of c-Fos, 35-kDa Fra and c-Jun proteins with the time. The increases of c-Fos, 35-kDa Fra and c-Jun protein levels induced by KA administration were also inhibited by CHX pre-administration. KA administration markedly increased both c-fos and c-jun mRNA levels during 1 and 4 h and the increased levels of these proto-oncogene mRNA were further prolonged by the treatment with CHX. In addition, CHX alone increased both c-fos and c-jun mRNA levels although the onset times of induction were different. In electrophoretic mobility shift-assay, both AP-1 and ENKCRE-2 DNA-binding activities were increased by KA. KA-induced increases of AP-1 and ENKCRE-2 DNA-binding activities were also attenuated by CHX. In addition, KA-induced AP-1 and ENKCRE-2 DNA-binding activities were diminished by the antibodies against Fos and Jun family proteins. Furthermore, the cross-competition studies revealed that AP-1 proteins actively participated in ENKCRE-2 DNA domain. The results suggest that KA-induced proENK and proDYN mRNA expressions may require on-going synthesis of proteins, such as c-Fos, c-Jun and 35-kDa Fra, which may have a possible role in the up-regulation of proENK and proDYN gene expression through the binding with AP-1 and ENKCRE-2 DNA-binding motifs.
Won
J S
JS
Department of Pharmacology, Institute of Natural Medicine, College of Medicine, Hallym University, Chunchon, Kangwon-Do, South Korea.
Kim
Y H
YH
Song
D K
DK
Suh
H W
HW
D000818:Animals; D015153:Blotting, Western; D003513:Cycloheximide / Q000494:pharmacology*; D004745:Enkephalins / Q000187:drug effects* / Q000378:metabolism; D015870:Gene Expression / Q000235:genetics*; D006624:Hippocampus / Q000187:drug effects* / Q000378:metabolism; D007608:Kainic Acid / Q000494:pharmacology*; D008297:Male; D011498:Protein Precursors / Q000187:drug effects* / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley
D004745:Enkephalins; D011498:Protein Precursors; C029992:proenkephalin; C033660:preproenkephalin; D003513:Cycloheximide; D007608:Kainic Acid