DNA sequences required for expression of the LH beta promoter in primary cultures of rat pituitary cells.
To begin analysis of the DNA sequences necessary for luteinizing hormone (LH) gene transcription, fusion genes containing the 5' flanking region of the rat LH beta or the human alpha-subunit gene linked to luciferase were transfected into primary cultures of rat pituitary cells. The LH beta-luciferase construct was expressed in the primary cultures at a level 50 times greater than a promoterless luciferase control plasmid. Little or no expression of the LH beta-luciferase construct was detected following transfection of MCF-7, JAR or GH3 tumor cell lines. Treatment of transfected cells with gonadotropin-releasing hormone resulted in a modest induction of LH beta-luciferase activity. Considerably higher levels of LH beta-luciferase activity were obtained with cultures from ovariectomized rats than were obtained with cultures from intact female rats. Analysis of 5' deletions of the LH beta-luciferase construct demonstrated that activity was well maintained even after substantial deletions. The shortest construct, which contained 75 base pairs of 5' flanking sequence had 38% of the activity of the longest which contained 1.7 kilobase pairs of flanking sequence. These findings demonstrate that transfection of primary cultures of rat pituitary cells may provide a useful system for analysis of the cis-acting sequences and trans-acting factors required for LH gene expression.
Kim
K E
KE
Department of Physiology and Biophysics, University of Iowa, Iowa City 52242.
Day
K H
KH
Howard
P
P
Salton
S R
SR
Roberts
J L
JL
Maurer
R A
RA
D000818:Animals; D002478:Cells, Cultured; D003001:Cloning, Molecular; D005260:Female; D005786:Gene Expression Regulation; D007987:Gonadotropin-Releasing Hormone / Q000494:pharmacology; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D007986:Luteinizing Hormone / Q000235:genetics* / Q000378:metabolism; D010902:Pituitary Gland / Q000166:cytology / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D051381:Rats; D012045:Regulatory Sequences, Nucleic Acid; D014162:Transfection
D007987:Gonadotropin-Releasing Hormone; D007986:Luteinizing Hormone; D008156:Luciferases