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TRANSLATIONAL PHYSIOLOGY

A novel missense mutation in SLC34A3 that causes hereditary hypophosphatemic rickets with hypercalciuria in humans identifies threonine 137 as an important determinant of sodium-phosphate cotransport in NaPi-IIc

¹Endocrine Unit and ²Department of Anesthesiology, Massachusetts General Hospital, Boston, Massachusetts; and ³Department of Pediatrics and Endocrine Unit, Yale School of Medicine, New Haven, Connecticut

Submitted 20 February 2008 ; accepted in final form 5 May 2008

The present study describes two novel compound heterozygous mutations, c.410C>T(p.T137M) (T137M) on the maternal and g.4225_50del on the paternal allele of SLC34A3, in a previously reported male with hereditary hypophosphatemic rickets with hypercalciuria (HHRH) and recurrent kidney stones (Chen C, Carpenter T, Steg N, Baron R, Anast C. Pediatrics 84: 276–280, 1989). For functional analysis in vitro, we generated expression plasmids encoding enhanced green fluorescence protein (EGFP) concatenated to the NH₂ terminus of wild-type or mutant human type IIc Na-P_i cotransporter (NaPi-IIc), i.e., EGFP-hNaPi-IIc, EGFP-[M137]hNaPi-IIc, or EGFP-[Stop446]hNaPi-IIc. The V446Stop mutant showed complete loss of expression and function when assayed for apical patch expression in opossum kidney (OK) cells and sodium-dependent ³³P uptake into Xenopus laevis oocytes. Conversely, EGFP-[M137]hNaPi-IIc was inserted into apical patches of OK cells and into oocyte membranes. However, when quantified by confocal microscopy, surface fluorescence was reduced to 40% compared with wild-type. After correction for surface expression, the rate of ³³P uptake by oocytes mediated by EGFP-[M137]hNaPi-IIc was decreased by an additional 60%. The resulting overall reduction of function of this NaPi-IIc mutant to 16%, taken together with complete loss of expression and function of g.4225_50del(V446Stop), thus appears to be sufficient to explain the phenotype in our patient. Furthermore, the stoichiometric ratio of ²²Na and ³³P uptake was increased to 7.1 ± 3.65 for EGFP-[M137]hNaPi-IIc compared with wild-type. Two-electrode studies indicate that EGFP-[M137]hNaPi-IIc is nonelectrogenic but displayed a significant phosphate-independent inward-rectified sodium current, which appears to be insensitive to phosphonoformic acid. M137 thus may uncouple sodium-phosphate cotransport, suggesting that this amino acid residue has an important functional role in human NaPi-IIc.

SLC34A3; rickets; hypercalciuria; nephrolithiasis

This article has been cited by other articles:

				M. Levi and S. Bruesegem Renal Phosphate-Transporter Regulatory Proteins and Nephrolithiasis N. Engl. J. Med., September 11, 2008; 359(11): 1171 - 1173. [Full Text] [PDF]

				M. Levi Novel NaPi-2c mutations that cause mistargeting of NaPi-2c protein and uncoupling of Na-Pi cotransport cause HHRH Am J Physiol Renal Physiol, August 1, 2008; 295(2): F369 - F370. [Full Text] [PDF]