Identification and Characterization of a Gene with Base Substitutions Associated with the Absorptive Hypercalciuria Phenotype and Low Spinal Bone Density

Abstract

Absorptive hypercalciuria (AH) is a kidney stone-forming condition frequently complicated by bone loss. Previously, we mapped the locus for an inherited form of AH to chromosome 1q23.3-q24. We have sequenced a putative gene (subsequently shown by others to be homologous with the rat soluble adenylate cyclase gene) in this region in 12 unrelated Caucasian AH patients. Eighteen base substitutions were identified in the soluble adenylate cyclase human homolog gene. All sequence variations were further evaluated in 3-68 additional unrelated AH patients and 19-132 normal subjects, and 1 additional base substitution was identified. Six of the identified sequence variations occurred with increased frequency in the AH population and tracked with the AH phenotype in AH families. Calculated odds ratios showed that the occurrence of any 4 of these individual base substitutions was associated with a 2.2- to 3.5-fold increase in estimated risk for AH (P < 0.02). In addition, 1 or more base changes was associated with a lower L2-L4 vertebral bone density. Sequence analysis of 3 other genes within the AH linkage interval showed no difference in the distribution of sequence variations between AH and normal populations. This is the first description of a specific gene defect associated with AH.

Soluble Adenylyl Cyclase Mediates Nerve Growth Factor-induced Activation of Rap1

“…Presence and Localization of sAC in PC12 Cells-Similar to tmACs, sAC is widely expressed (17,18,35). We confirmed sAC mRNA expression in PC12 cells by reverse transcription-PCR (data not shown) and protein expression by immunoblotting, using a sAC-specific monoclonal antibody.…”

“…We confirmed sAC mRNA expression in PC12 cells by reverse transcription-PCR (data not shown) and protein expression by immunoblotting, using a sAC-specific monoclonal antibody. Recent studies in our (data not shown) and others' laboratories (17,35) reveal complex patterns of alternative splicing leading to multiple isoforms of sAC protein in somatic tissues. Western blotting identified a single protein band of ϳ70 kDa (Fig.…”

“…Similar to NGF, PACAP activates Rap1 and elicits neurite outgrowth (13)(14)(15), but all attempts to link NGF or its receptors to tmAC activation have been unsuccessful. We have identified an additional, ubiquitously expressed (17,18) source of cAMP in mammalian cells, "soluble" adenylyl cyclase (sAC) (16). In contrast to tmACs, sAC is not regulated by heterotrimeric G proteins but is, instead, regulated by bicarbonate (19) and calcium (20,21) ions.…”

New Insights Into the Pathogenesis of Idiopathic Hypercalciuria

“…A genome-wide linkage study of three kindreds with IH and low bone density identified a locus that encodes a gene for the human soluble adenylyl cyclase [44]. Several sequence variations in the gene tracked with IH in an unrelated group of 80 hypercalciuric patients with kidney stones, compared with a group of non-stone formers [45]. The protein is found in kidney and in bone, and may function as a bicarbonate sensor [46].…”

Molecular Mechanisms of Primary Hypercalciuria

“…Reed et al (58) have mapped a defect in three kindreds with increased intestinal absorption of calcium to 1q23.3-q24, and they have identified a group of base substitutions in a putative gene within that locus, four of which increased the relative risk of disease 2.2-fold to 3.5-fold (59). The sequence has pronounced homology to the rat soluble adenylate cyclase gene and transcripts were detected, although the functionality of this gene has yet to be established (59). It may be of interest that rat CaT1 has a potential protein kinase A phosphorylation site near the amino terminus that is absent from the corresponding region of ECaC (38).…”