Comparison of long and short forms of the prolactin receptor on prolactin-induced milk protein gene transcription.

Abstract

The biological activities of long and short forms of the prolactin receptor have been compared. These two receptors expressed in mammalian cells were shown to bind prolactin with equal high affinity. The ability of these different forms to transduce the hormonal message was estimated by their capacity to stimulate transcription by using the promoter of a milk protein gene fused to the chlormphenicol acetyltransferase (CAT) coding sequence. Experiments were performed in serum-free conditions to avoid the effect oflactogenic factors present in serum. An 417-fold induction of CAT activity was obtained in the presence of prolactin when the long form of the prolactin receptor was expressed, whereas no induction was observed when the short form was expressed. The present results clearly establish that only the long form of the prolactin receptor is involved in milk protein gene transcription.

A MGF/STAT5 binding site is necessary in the distal enhancer for high prolactin induction of transfected rabbit αs1‐casein‐CAT gene transcription

“…CHO K1 cells were grown and transfected as previously described [14] with modifications [11]. Transfection of plasmid DNA was performed using the calcium phosphate procedure, with 40 gg DNA in each precipitate.…”

“…Each precipitate was obtained with 6 gg of the CAT plasmid to be tested, 12 gg of the pCHll0 plasmid coding for 13-galactosidase (Pharmacia), 12 gg of the plasmid pER2-3 carrying the rabbit mammary prolactin receptor cDNA [15], and 10 gg of carrier DNA (genomic sonicated DNA from salmon testes). After ~5-h incubation in the presence of the precipitate, the cells were washed and cultured in serum-free medium [14] containing hormones. Two dishes were incubated with cortisol (60 nM).…”

Prolactin receptor signal transduction in cells of the immune system

“…In humans, two PRLr isoforms have been identified: the long (Boutin et al 1989) and a novel intermediate isoform, recently identified in our laboratory (Clevenger et al 1995a). All PRLr isoforms are homologous in their extracellular and transmembrane domain; alternative mRNA splicing accounts for the differing sizes of the cytoplasmic domain, with the exception of the Nb2 mutant form, which is believed to result from a mutation in the exon encoding for rat PRLr intracellular domain (Ali et al 1991). The extracellular domain of the PRLr demonstrates homology via a tryptophan-serine box motif with the gene superfamily of cytokine receptors that includes the receptors for IL-2 to IL-7, GM-CSF, GH, and erythropoietin (Bazan 1989).…”

“…Three isoforms of PRLr have been cloned in the rat (see Fig. 2): the 'short' (PRLr-S, approximately 291 amino acids, approximately 45 kDa), 'long' (PRLr-L, approximately 592-598 amino acids, 80-85 kDa), and intermediate (PRLr-I, a deletion mutant missing amino acids 323-520 of the PRLr-L isoform, found in the PRL-dependent rat T-cell lymphoma line Nb2; 393 amino acids, approximately 65 kDa) (Boutin et al 1988, Ali et al 1991. In humans, two PRLr isoforms have been identified: the long (Boutin et al 1989) and a novel intermediate isoform, recently identified in our laboratory (Clevenger et al 1995a).…”

Identification of Cytoplasmic Motifs Required for Short Prolactin Receptor Internalization

“…In addition to these PRLR isoforms found in vivo, an intermediate form, lacking 198 aa in the cytoplasmic domain with respect to the long form, has been identified in the rat pre-T lymphoma Nb2 cell line (20). Studies of the in vitro biological properties of PRLR isoforms have shown that the long and intermediate, but not the short form, are able to transactivate milk protein gene promoters (21,22). However, involvement of both long and short PRLR isoforms in cell proliferation has also been proposed (23).…”