The renal urea transporter gene (UT-A) produces different transcripts in the inner medullary collecting ducts (UT-A1) and thin descending limbs of Henle's loop (UT-A2), coding for distinct proteins. Peptide-directed rabbit polyclonal antibodies were used to identify the UT-A2 protein in renal medulla of mouse and rat. In the inner stripe of outer medulla, an antibody directed to the COOH terminus of UT-A recognized a membrane protein of 55 kDa. The abundance of this 55-kDa protein was strongly increased in response to chronic infusion of the vasopressin analog 1-deamino-[8-D-arginine]vasopressin (DDAVP) in Brattleboro rats, consistent with previous evidence that UT-A2 mRNA abundance is markedly increased. Immunofluorescence labeling with the COOH-terminal antibody in Brattleboro rats revealed labeling in the lower portion of descending limbs from short-looped nephrons (in the aquaporin-1-negative portion of this segment). This UT-A labeling was increased in response to DDAVP. Increased labeling was also seen in descending limbs of long-looped nephrons in the base of the inner medulla. These results indicate that UT-A2 is expressed as a 55-kDa protein in portions of the thin descending limbs of Henle's loop and that the abundance of this protein is strongly upregulated by vasopressin.

The prohormone convertases (PCs) 1/3 and 2 accomplish the major proteolytic cleavage events in neuroendocrine tissues; each of these convertases has a small associated binding protein that inhibits convertase action in the secretory pathway. The proSAAS protein binds to PC1/3, whereas the 7B2 protein binds to PC2. However, both convertase-binding proteins are more widely expressed than their cognate enzymes, suggesting that they may perform other functions as well. All known mammalian proSAASs are over 85% conserved; thus, identifying functionally important segments has been impossible. Here, we report the first identification of nonmammalian proSAAS molecules, from Xenopus and zebrafish (Danio rerio). Although these two proteins show an overall amino acid sequence identity of only 29 and 30% with mouse proSAAS, two 14-16 residue hydrophobic segments (predicted to form alpha-helices) and two, nine through 11 residue sequences containing basic convertase cleavage sites are highly conserved; therefore, these sequences may be of functional importance. Confidence that these nonmammalian molecules represent authentic proSAAS is supported by the finding that both inhibit mouse PC1/3 with nanomolar inhibition constants; human furin was not inhibited. In vitro, the two proteins were cleaved by PC2 and furin to three or more peptide products. Both zebrafish and Xenopus proSAAS exhibited neural and endocrine distributions, as assessed by in situ and PCR experiments, respectively. In summary, the identification of proSAAS molecules in lower vertebrates provides clues as to functional regions within this widely expressed neuroendocrine protein.

Evidence suggested a role for genetic variation in IL6 and LRP5 in conferring risk for osteoporosis in Caucasian women, with the latter manifest only in smokers.