Sequence and functional characterisation of the marmoset monkey (Callithrix jacchus) prolactin receptor: comparative homology with the human long-form prolactin receptor

This study demonstrates the cloning and in-vitro characterisation of the ma rmoset monkey (Callithrix jacchus) prolactin receptor cDNA. The marmoset pr olactin receptor cDNA was generated by reverse transcription-polymerase cha in reaction using adrenal RNA and primers designed from prolactin receptor conserved regions. Sequence analysis predicts a mature protein of 598 amino acids exclusive of the 24 amino acid signal peptide. The marmoset prolacti n receptor cDNA shares 93 and 61% base pair, and 89 and 61% amino acid sequ ence homologies with the long form human and rat prolactin receptor cDNA, r espectively. The marmoset prolactin receptor cDNA sequence retains all the receptor sequences that have been shown previously to be essential for liga nd binding, structural integrity and signal transduction. Transfection of h uman 293 fibroblast cells with the marmoset prolactin receptor cDNA (three independent experiments) confirmed the expression of a receptor that has hi gh, binding affinity to human growth hormone (K-a = 3.6 +/- 0.07 nM(-1) and B-max = 7.55 +/- 2.06 x 10(-11) M) and human prolactin (K-a = 3.1 +/- 0.12 nM(-1) and B-max = 2.87 +/- 0.66 x 10(-11) M). Functionality of the recept or was assessed by co-transfection of 293 fibroblast cells with marmoset pr olactin receptor cDNA and the Jak2 cDNA, or marmoset prolactin receptor and a Stat5 responsive element linked to the luciferase coding sequence. Incub ation of the cells with 18 nM ovine prolactin resulted in rapid phosphoryla tion of Jak2 as ascertained by Western blotting. In addition, the marmoset prolactin receptor cDNA led to 9.06 +/- 0.47-fold induction of luciferase g ene activity: This was comparable with the induction observed following tra nsfection with the human prolactin receptor cDNA (8.55 +/- 0.5-fold). In-vi vo prolactin receptor expression in the marmoset monkey was assessed by rib onuclease protection assay and detected in a number of tissues including fe male reproductive organs. These data confirm the cloning and functionality of the marmoset prolactin receptor cDNA. The marmoset prolactin receptor sh ares a high sequence homology with the long-form human prolactin receptor, and both receptors bind hormones with comparable affinity and confer a simi lar intracellular response. The marmoset monkey may provide a useful tool t o investigate the role of prolactin in primate reproduction. (C) 2000 Elsev ier Science Ireland Ltd. All rights reserved.