Abstract
Whereas molecular cloning experiments have provided evidence for the presence of two closely related genes for the catalytic subunit of the cAMP-dependent protein kinase, the possible interchangeability of these isoforms in initiating biological processes has not been examined. To test the role of the two forms of the kinase in regulating transcription, expression vectors containing the coding sequence of either kinase have been cotransfected with a fusion gene containing the prolactin promoter coupled to an appropriate marker gene. The results demonstrate that expression vectors for both isoforms of the catalytic subunit are able to increase prolactin promotor activity in a dose-dependent manner. The effects can be observed by measuring either marker gene activity or RNA levels. Transfection of an expression vector encoding an inactive catalytic subunit did not stimulate prolactin promoter activity. The results provide additional evidence for the role of the catalytic subunit of the cAMP-dependent kinase in mediating regulation of specific gene transcription and demonstrate that both forms of the catalytic subunit are capable of participating in the regulation of transcription.
Highlights
Prolactin promoter coupled to an appropriate marker gene
The results provide additional evidence for the proteins encoded by the mouse Cat-a [8]and Cat-@ [4]cDNAs, the role of the catalytic subunit of the CAMP-dependent respectively.The Cat-n and Catc-poding sequenceplus the glycoprokinase in mediating regulation of specific gene transcription and demonstrate that both forms of the catalytic subunit are capable of participating in the regulation of transcription
To test thaebility of the two different catalytic subunitsto participate in the regulation of transcription, gene transfer treatment with cAMP ( B ) .GHs cells were transfected with 5 pg of the prolactin-luciferase reporter gene plus the indicated concentration of either Cat-a or Cat-pexpression vector and sufficient pUC18 DNA to make a total of 20 pg of DNA
Summary
Prolactin promoter coupled to an appropriate marker gene. The results demonstrate that expression vectors for both isoforms of the catalytic subunit are able to increase prolactin promoter activity ina dose-dependent manner. Nuclease Protection Assay of Fusion Gene Transcripts-RNA was prepared from cells 24 h aftertransfection with prolactin-globin of the known effects of cAMP in eucaryotes are mediated by marker constructs.
Published Version
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