A mouse with an N-Ethyl-N-nitrosourea (ENU) Induced Trp589Arg Galnt3 mutation represents a model for hyperphosphataemic familial tumoural calcinosis

Journal article

Mutations of UDP-N-acetyl-alpha-D-galactosamine polypeptide N-acetyl galactosaminyl transferase 3 (GALNT3) result in familial tumoural calcinosis (FTC) and the hyperostosis-hyperphosphataemia syndrome (HHS), which are autosomal recessive disorders characterised by soft-tissue calcification and hyperphosphataemia. To facilitate in vivo studies of these heritable disorders of phosphate homeostasis, we embarked on establishing a mouse model by assessing progeny of mice treated with the chemical mutagen N-ethyl-N-nitrosourea (ENU), and identified a mutant mouse, TCAL, with autosomal recessive inheritance of ectopic calcification, which involved multiple tissues, and hyperphosphataemia; the phenotype was designated TCAL and the locus, Tcal. TCAL males were infertile with loss of Sertoli cells and spermatozoa, and increased testicular apoptosis. Genetic mapping localized Tcal to chromosome 2 (62.64-71.11 Mb) which contained the Galnt3. DNA sequence analysis identified a Galnt3 missense mutation (Trp589Arg) in TCAL mice. Transient transfection of wild-type and mutant Galnt3-enhanced green fluorescent protein (EGFP) constructs in COS-7 cells revealed endoplasmic reticulum retention of the Trp589Arg mutant and Western blot analysis of kidney homogenates demonstrated defective glycosylation of Galnt3 in Tcal/Tcal mice. Tcal/Tcal mice had normal plasma calcium and parathyroid hormone concentrations; decreased alkaline phosphatase activity and intact Fgf23 concentrations; and elevation of circulating 1,25-dihydroxyvitamin D. Quantitative reverse transcriptase-PCR (qRT-PCR) revealed that Tcal/Tcal mice had increased expression of Galnt3 and Fgf23 in bone, but that renal expression of Klotho, 25-hydroxyvitamin D-1α-hydroxylase (Cyp27b1), and the sodium-phosphate co-transporters type-IIa and -IIc was similar to that in wild-type mice. Thus, TCAL mice have the phenotypic features of FTC and HHS, and provide a model for these disorders of phosphate metabolism.

Authors: Esapa, C; Head, R; Jeyabalan, J; Evans, H; Hough, T

• Publication status: Published
• Peer review status: Peer reviewed
• Publisher: Public Library of Science
• Journal: PLoS ONE
• Volume: 7
• Issue: 8
• Pages: e43205
• Publication date: 2012-08-13
• Acceptance date: 2012-07-18
• DOI: 10.1371/journal.pone.0043205
• EISSN: 1932-6203
• ISSN: 1932-6203
• Language: English
• Keywords: Animals; Calcinosis; N-Acetylgalactosaminyltransferases; Reverse Transcriptase Polymerase Chain Reaction; Fibroblast Growth Factors; Sertoli Cells; Hyperphosphatemia; Hyperostosis, Cortical, Congenital; Mice; Mutation, Missense; SBTMR; Disease Models, Animal; Testis; Blotting, Western; Spermatozoa; Green Fluorescent Proteins; Apoptosis; Male; COS Cells; Ethylnitrosourea; Cercopithecus aethiops; Bone and Bones; Genes, Recessive