The LIM homeobox protein mLIM3/Lhx3 induces expression of the prolactin gene by a Pit-1/GHF-1-independent pathway in corticotroph AtT20 cells.
mLIM3, a member of the LIM homeobox family, was recently demonstrated to be critical for proliferation and differentiation of the pituitary cell lineage. Using a pool of degenerate oligonucleotides we determined the DNA sequence ANNAGGAAA(T/C)GA(CIG)AA as the set preferentially recognized by mLIM3. A nearly identical sequence is found in the prolactin (PRL) promoter, within a 15-mer stretch from nucleotides (nts) -218 to -204 which is highly conserved between human, rat, and bovine. In order to test the hypothesis of a transcriptional effect of mLIM3 on the prolactin promoter, stable transfectants of mLIM3 cDNA in AtT20 tumor cells revealed that PRL mRNA expression was induced in 3 separate stable clones. Gel retardation experiments performed using nuclear extracts isolated from one of the AtT20/mLIM3 stable transfectants revealed affinity towards the 15-mer element of the PRL promoter. From these results, we propose that the PRL promoter element (nts -218 to -204) could be functional in vivo. Finally, we demonstrate that in AtT20 cells prolactin mRNA expression is not induced by the Pit-1/GHF-1 pathway and that growth hormone mRNA is not detected concomitantly with prolactin. We conclude that mLIM3 may play a key role in inducing PRL gene expression in lactotrophs by binding to a conserved motif close to a Pit-1/GHF-1 site within the proximal PRL promoter.
Laboratory of Biochemical Neuroendocrinology, Clinical Research Institute of Montreal, Qué., Canada. seidahn@ircm.umontreal.ca
S E
SE
Girardin
S
S
Benjannet
J C
JC
Barale
M
M
Chrétien
N G
NG
Seidah
D000818:Animals; D001483:Base Sequence; D002417:Cattle; D002454:Cell Differentiation; D002460:Cell Line; D016384:Consensus Sequence; D004247:DNA / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005786:Gene Expression Regulation; D018398:Homeodomain Proteins / Q000378:metabolism*; D006801:Humans; D060850:LIM-Homeodomain Proteins; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D010902:Pituitary Gland / Q000166:cytology / Q000378:metabolism*; D011388:Prolactin / Q000235:genetics*; D011401:Promoter Regions, Genetic; D051381:Rats; D050817:Transcription Factor Pit-1; D014157:Transcription Factors / Q000378:metabolism*
D004268:DNA-Binding Proteins; D018398:Homeodomain Proteins; D060850:LIM-Homeodomain Proteins; C094898:Lhx3 protein; D009419:Nerve Tissue Proteins; C494187:POU1F1 protein, human; C494186:Pou1f1 protein, rat; D050817:Transcription Factor Pit-1; D014157:Transcription Factors; D011388:Prolactin; D004247:DNA