HUMAN MUTATION 29(4), 532^536, 2008 RESEARCH ARTICLE Sedoheptulokinase Deficiency Due to a 57-kb Deletion in Cystinosis Patients Causes Urinary Accumulation of Sedoheptulose: Elucidation of the CARKL Gene Mirjam M.C. Wamelink, 1Ã Eduard A. Struys, 1 Erwin E.W. Jansen, 1 Elena N. Levtchenko, 2 Fokje S.M. Zijlstra, 3 Udo Engelke, 3 Henk J. Blom, 1 Cornelis Jakobs, 1 and Ron A. Wevers 3 1 Department of Clinical Chemistry, Metabolic Unit, VU University Medical Center, Amsterdam, The Netherlands; 2 Department of Pediatric Nephrology, Radboud University Nijmegen Medical Center, Nijmegen, The Netherlands; 3 Laboratory of Pediatrics and Neurology, Radboud University Nijmegen Medical Center, Nijmegen, The Netherlands Communicated by Johannes Zschocke The most common mutation in the nephropathic cystinosis (CTNS) gene is a homozygous 57-kb deletion that also includes an adjacent gene carbohydrate kinase-like (CARKL). The latter gene encodes a protein that is predicted to function as a carbohydrate kinase. Cystinosis patients with the common 57-kb deletion had strongly elevated urinary concentrations of sedoheptulose (28–451 mmol/mol creatinine; controls and other cystinosis patients o9) and erythritol (234–1110 mmol/mol creatinine; controls and other cystinosis patients o148). Enzyme studies performed on fibroblast homogenates derived from patients carrying the 57-kb deletion revealed 80% reduction in their sedoheptulose phosphorylating activity compared to cystinosis patients with other mutations and controls. This indicates that the CARKL-encoded protein, sedoheptulokinase (SHK), is responsible for the reaction: sedoheptulose 1 ATP - sedoheptulose-7-phosphate 1 ADP and that deletion of CARKL causes urinary accumulation of sedoheptulose and erythritol. Hum Mutat 29(4), 532–536, 2008. r r 2008 Wiley-Liss, Inc. KEY WORDS: sedoheptulokinase; sedoheptulose; CARKL; cystinosis INTRODUCTION Recently we described two new defects in the pentose phosphate pathway (PPP): Transaldolase deficiency (MIM] 606003) and ribose-5-phosphate isomerase deficiency (MIM] 608611) [Verhoeven et al., 2001; Huck et al., 2004]. Here, we report further elucidation of another gene and protein involved in the PPP. Carbohydrate kinase-like (CARKL; MIM] 605060) encodes a deduced 478–amino acid protein [Touchman et al., 2000]. The CARKL protein contains motifs showing weak similarity to two domains of the FGGY family of carbohydrate kinases and appears to be localized in the cytoplasm. With northern blot analysis, two CARKL transcripts were detected: one of 3.9kb expressed predominantly in liver, kidney, and pancreas, with weaker expression in heart, placenta, brain, and lung, and one 2.7-kb transcript that was detected in liver and, to a lesser extent, in heart [Touchman et al., 2000]. Carbohydrate kinases are a class of proteins involved in the phosphorylation of sugars as they enter a cell, inhibiting their transport across the cell membrane. Nephropathic cystinosis (MIM]s 219800; 219900; 219750) is an autosomal recessive lysosomal storage disease caused by mutations in the nephropathic cystinosis (CTNS) gene [Town et al., 1998] (MIM] 606272). The most common mutation in cystinosis is the 57,257-bp deletion [Touchman et al., 2000], which is found in a homozygous state in approximately 50% of patients from northern European descent. Within this common deletion the adjacent CARKL gene is included (Fig. 1). In urine of one patient with cystinosis (whose clinical details have been published previously) [Levtchenko and Monnens, 2006] caused by the 57-kb deletion, we retrospectively found elevated concentra- tions of sedoheptulose and erythritol. We subsequently analyzed urinary concentrations of sedoheptulose and erythritol and sedohep- tulokinase (SHK) activity in fibroblasts from nephropathic cystinosis patients with and without the homozygous 57-kb deletion. Here, we report the function of the CARKL gene and show that cystinosis patients caused by the 57-kb deletion have elevated concentrations of sedoheptulose and erythritol and decreased SHK activity in fibroblasts due to deficiency of the CARKL gene. MATERIALS AND METHODS Patients Urine samples from 10 nephropathic cystinosis patients were analyzed. The diagnoses of cystinosis was made biochemically (by the Published online 10 January 2008 in Wiley InterScience (www. interscience.wiley.com). DOI 10.1002/humu.20685 Received 8 June 2007; accepted revised manuscript 12 October 2007. Ã Correspondence to: Mirjam M.C. Wamelink, Metabolic Unit, Department of Clinical Chemistry, VU University Medical Center, De Boelelaan1117,1081HVAmsterdam,The Netherlands. E-mail: M.Wamelink@vumc.nl r r 2008 WILEY-LISS, INC.