Asymptomatic Isolated Human Glycerol Kinase Deficiency Associated with Splice-Site Mutations and Nonsense-Mediated Decay of Mutant RNA

Abstract

Isolated glycerol kinase deficiency (GKD) is an Xlinked inborn error of metabolism that is either symptomatic or asymptomatic. GKD is due to deletions of, or mutations within, the GK gene, and there is no genotype-phenotype correlation. We identified three patients with asymptomatic GKD, determined that they had GK splice-site mutations, and studied the stability of their GK mRNA to understand the molecular mechanism of the GKD. All three patient mutations caused a frameshift and introduction of a premature stop codon. A fourth patient had an Alu insertion in intron 4 that led to alternative splicing. To study the effect of splice-site mutations on RNA species, we performed reverse transcriptase PCR and found only normal-sized products for all patients. Incubation with anisomycin to block nonsense-mediated decay (NMD), revealed two RNA species for each individual. Sequence analysis revealed that the larger bands represented the wild-type GK RNA and smaller bands represented mutant misspliced RNA, suggesting that the abnormal RNA species were targeted by NMD. Normal RNA species observed in each patient are likely responsible for their mild phenotypes. We speculate that influences on RNA processing and protein stability represent modifiers of the GKD phenotype. (Pediatr Res 59: 590-592, 2006) I solated GKD is an X-linked inborn error of metabolism (1). We have previously shown that GKD is due to deletions of, or mutations within, the glycerol kinase gene (GK) located on Xp21 that encodes a 524 aa protein (1). Isolated GKD is either asymptomatic or symptomatic with episodes of vomiting, acidosis (metabolic crises), and lethargy that can progress to coma or CNS crises. GKD is caused by large deletions, insertions, missense mutations, nonsense mutations, and splice-site mutations (1-3). Our previous work has shown that there is no genotype-phenotype correlation in GKD (2). We hypothesize that this lack of genotype-phenotype correlation in patients with GKD is due to the role of modifier genes in the GK phenotype (2,4).NMD is a quality-control mechanism naturally occurring in cells to maintain homeostasis and destroy mRNA that contain premature stop codons (5,6). This process is evolutionarily conserved and is found in species from yeast (Saccharomyces cerevisiae) to worms (Caenorhabditis elegans) to humans (5). NMD occurs in the nucleus and cytoplasm (5,6), and evidence suggests that NMD and alternative splicing are coupled (7). Better understanding of this natural process to minimize the effect of DNA mutations in the genome will be important in understanding human disease.The purpose of this investigation was to identify patients with GKD due to splice-site mutations and to determine the effect of the splice-site mutation on the GK mRNA stability. Our results indicate that GKD is one of the many disorders with splice-site mutations and that the aberrant transcripts are targeted for NMD. METHODSIndividuals with isolated GKD. Patient 020518-1 presented with pseudohypertriglyceridemia and asymptomatic GKD. H...

A Novel Splice-Site Mutation of the Gk Gene in a Chinese Patient with Isolate Glycerol Kinase Deficiency

“…Third, it may be related to Nonsense Mediated Decay (NMD). Zhang et al, found that aberrant mRNA generated by splice site mutations of the GK gene can be eliminated by NMD in the asymptomatic form of GKD [9]. In contrary, there was no NMD in our study, and reviewed the other split-site mutations previously reported, no NMD was found too [4,5].…”

“…Interestingly, most of the patients with split-site mutation reported previously presented with asymptomatic form of GKD [4,5,[7][8][9]. Only one patient from the literature (IVS9-1G>T) who had a history of metabolic and CNS crises at the age 6 years and 3 months, indicating the symptomatic form of GKD, but he was uneventful in the neonatal period and early childhood [10].…”

Glycerol kinase deficiency in adults: Description of 4 novel cases, systematic review and development of a clinical diagnostic score

“…The 15 studies included a total of 39 adult participants (aged 23-70 years). All articles reported severe HTG: thirteen articles showed a range of TG from 484 to 1992 mg/dL [5,7,8,[14][15][16][17][18][19][20][21][22][23], however two of them did not report concrete values of TG [24,25]. Twelve articles reported high levels of plasma glycerol [5,7,8,14,15,[17][18][19][20][21][22][23][24], with a range from 0.195 to 210 mmol/L, and eight of them showed the presence of glycerol in urine.…”

“…p. (Asn288Asp), p.(Ala305Val), p.(Gln438Arg), p.(Thr96Ile) p.(Cys358Tyr), p.(Gly280Ala) [5,20,21,24]; c) three articles [20,22,25] reported four causative splice site mutation:…”

Closing the Gap: Inverting the Genetics Curriculum to Ensure an Informed Public

“…In GK deficiency, variation in protein stability and RNA processing appear to modify phenotype. 10 The genetic underpinnings of Mendelian disease will continue to integrate with theoretical and experimental work in biochemistry to explicate phenotypic complexity, for example, by providing a better understanding of the kinetic behavior of enzyme reactions and metabolic flux, which behave as nonlinear systems. 11 However, the precise phenotypic outcomes will probably remain predictable only at low resolution because ''for many diseases, only a subset of all mutations reliably predicts phenotypes.''…”