Determination of the amino acid sequence of bovine parathyroid hormone has led to the synthesis of a tetratriacontapeptide corresponding to the aminoterminal 1-34 residues of the native molecule. The specific biological effects of this synthetic peptide on bone and kidney are qualitatively identical to those of the native hormone in classical bioassays in vivo and in several systems in vitro. Potency of the synthetic peptide equals or exceeds that of a biologically active fragment of comparable size isolated from the native hormone; the synthetic and natural peptides show complete immunological crossreactivity. Thus, essential requirements for the physiological actions of the peptide on both skeletal and renal tissue are contained within the 34 amino-terminal amino acids. The potency of the synthetic peptide, relative to that of the native (84-amino acid) polypeptide, is greater in vitro than in vivo; this suggests that the carboxyl terminal two-thirds of the native hormone may protect the circulating polypeptide from rapid metabolic degradation.

1 The binding characteristics of the relaxin receptor in rat atria, uterus and cortex were studied using a [ 33 P]-labelled human gene 2 relaxin (B33) and quantitative receptor autoradiography. 2 The binding kinetics of [ 33 P]-human gene 2 relaxin (B33) were investigated in slide-mounted rat atrial sections. The binding achieved equilibrium after 60 min incubation at room temperature (23+18C) and dissociated slowly. The association and dissociation rate constants were 4.31+0.34610 8 M 71 min 71 and 1.55+0.38610 73 min 71 respectively. Thus, the kinetic dissociation constant was 3.46+0.59 pM. 3 Binding was saturable to a single population of non-interacting sites throughout atria, in uterine myometrium and the 5th layer of cerebral cortex. The binding a nities (pK D ) of [ 33 P]-human gene 2 relaxin (B33) were 8.92+0.09 in atrial myocardium and 8.79+0.04 in cerebral cortex of male rats, and 8.79+0.10 in uterine myometrium. Receptor densities in the cerebral cortex and atria were higher than in uterine myometrium, indicating that relaxin also has important roles in nonreproductive tissues. 4 In male rats, treatment with 17b-oestradiol (20 mg in 0.1 ml sesame oil s.c., 18 ± 24 h) signi®cantly decreased the density of relaxin receptors in atria and cerebral cortex. Identical treatment in female rats had no signi®cant e ect in atria and cerebral cortex, but it signi®cantly increased the density of relaxin receptors in uterine myometrium. 5 Relaxin binding was competitively displaced by porcine and rat native relaxins. Porcine native relaxin binds to the relaxin receptor in male rat atria (8.90+0.02), and cerebral cortex (8.90+0.03) and uterine myometrium (8.89+0.03) with a nities not signi®cantly di erent from human gene 2 (B33) relaxin. Nevertheless, rat relaxin binds to the receptors with a nities (8.35+0.09 in atria, 8.22+0.07 in cerebral cortex and 8.48+0.06 in uterine myometrium) signi®cantly less than human gene 2 (B33) and porcine relaxins. 6 Quantitative receptor autoradiography is the method of choice for measurement of a nities and densities of relaxin receptor in atria, uterine myometrium and cerebral cortex. High densities were found in all these tissues. 17b-Oestradiol treatment produced complex e ects where it increased the densities of relaxin receptors in uterus but decreased those in atria and cerebral cortex of the male rats, and had no e ect on the atria and cerebral cortex of the female rats.

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The relaxin receptor (LGR7) and the insulin-like peptide 3 (INSL3) receptor (LGR8) are unique LGR family members in possessing a single, functionally important amino terminal LDL-A module.1 Mouse and rat cDNA was screened for LGR7 and LGR7 splice variant expression. A uterus-specific exon 4 deleted variant was identified and named LGR7-Truncate. Deletion of exon 4 results in a premature stop codon and a transcript that putatively encodes a secreted protein containing LGR7's LDL-A module. Expression of LGR7-Truncate with LGR7 in HEK-293T cells resulted in decreased relaxin-induced signaling of LGR7. LGR7-Truncate is potentially an endogenous regulator of LGR7 signaling.

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1. Relaxin is an extracellular matrix (ECM)-remodelling hormone that is functionally important in reproductive tissues, brain, lung and heart. 2. Recently, the human relaxin receptor was identified as leucine-rich repeat-containing G-protein-coupled receptor 7 (LGR7). 3. Using human LGR7 as a template, we identified mouse and rat LGR7 orthologues in the Celera and National Centre for Biotechnology Information databases. 4. At the protein level, mouse and rat LGR7 share 85.2 and 85.7% identity with human LGR7, respectively. 5. Mouse LGR7 mRNA was detected in all tissues where relaxin binding is observed. 6. Mouse and rat LGR7 bound [33P]-relaxin with high affinity and, upon relaxin treatment, both receptors stimulated cAMP production in transfected HEK 293T cells. 7. These results indicate that mouse and rat LGR7 are the relaxin receptors in these species. 8. The actions of relaxin in rodents are well characterized, providing an established platform for research into the molecular pharmacology of the highly conserved relaxin receptor.

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The gene encoding rhesus monkey relaxin has been investigated. A cDNA library was prepared using corpus luteal RNA from a pregnant rhesus monkey, cDNA clones encoding relaxin were isolated and the nucleotide sequence was determined. The amino acid sequence of rhesus monkey preprorelaxin, predicted from the cDNA, demonstrates that the sequence has not been strongly conserved when compared with that of man, although features characteristic of the relaxin molecule have been maintained. This structural information will allow production of rhesus monkey relaxin, leading to studies investigating the bioactivity of relaxin in a homologous primate system. Southern blot analysis indicated that there is only one relaxin gene in the rhesus monkey and baboon genomes. In this respect these primate genomes are different from the human genome which contains two relaxin genes.