The Nb2 form of prolactin receptor is able to activate a milk protein gene promoter.
We have recently cloned a cDNA encoding a mutant form of PRL receptor (PRL-R) from Nb2 cells, a PRL-dependent T lymphocyte-derived cell line. This cDNA is identical to the long form of the rat PRL-R, except for a deletion of 594 base pairs in the cytoplasmic domain, resulting in a mature receptor protein of 393 amino acids. Although a segment containing three cytoplasmic regions of moderate to high amino acid sequence identity with members of the PRL/GH receptor family is missing in this receptor form, the region of highest (70%) identity is retained. In the following studies, a homologous functional assay was developed to test the activity of three forms of receptor with respect to their ability to transmit a lactogenic signal. In this system, CHO cells were transiently transfected with a construct containing 2300 base pairs of the 5'-flanking sequence of the rat beta-casein gene fused to the chloramphenicol acetyltransferase (CAT) gene and an expression vector containing the various forms of rat PRL-R cDNA. The transfected cells were grown in serum-free medium in the absence or presence of PRL. In cells transfected with the long form of the PRL-R and beta-casein/CAT construct, a 7.2- +/- 0.9-fold induction (n = 3) of CAT activity was seen when cells were cultured in the presence of 400 ng/ml PRL and 1 micrograms/ml hydrocortisone. This level of stimulation was similar to that observed for the ovine beta-lactoglobulin/CAT construct in which a 5.7- +/- 1.2-fold (n = 3) effect was found.(ABSTRACT TRUNCATED AT 250 WORDS)
Ali
S
S
Laboratory of Molecular Endocrinology, McGill University, Royal Victoria Hospital, Montréal, Québec, Canada.
Edery
M
M
Pellegrini
I
I
Lesueur
L
L
Paly
J
J
Djiane
J
J
Kelly
P A
PA
D000818:Animals; D016466:CHO Cells; D002364:Caseins / Q000235:genetics*; D006224:Cricetinae; D004247:DNA / Q000235:genetics; D017353:Gene Deletion; D005786:Gene Expression Regulation / Q000502:physiology*; D009154:Mutation / Q000235:genetics; D011388:Prolactin / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000502:physiology; D051381:Rats; D011981:Receptors, Prolactin / Q000235:genetics / Q000502:physiology*; D015398:Signal Transduction / Q000235:genetics; D014158:Transcription, Genetic / Q000235:genetics; D014162:Transfection / Q000235:genetics
D002364:Caseins; D011981:Receptors, Prolactin; D011388:Prolactin; D004247:DNA