Identification of Mutations Underlying 20 Inborn Errors of Metabolism in the United Arab Emirates Population

Abstract

Inborn errors of metabolism (IEM) are frequently encountered by physicians in the United Arab Emirates (UAE). However, the mutations underlying a large number of these disorders have not yet been determined. Therefore, the objective of this study was to identify the mutations underlying a number of IEM disorders among UAE residents from both national and expatriate families. A case series of patients from 34 families attending the metabolic clinic at Tawam Hospital were clinically evaluated, and molecular testing was carried out to determine their causative mutations. The mutation analysis was carried out at molecular genetics diagnostic laboratories. Thirty-eight mutations have been identified as responsible for twenty IEM disorders, including in the metabolism of amino acids, lipids, steroids, metal transport and mitochondrial energy metabolism, and lysosomal storage disorders. Nine of the identified mutations are novel, including two missense mutations, three premature stop codons and four splice site mutations. Mutation analysis of IEM disorders in the UAE population has an important impact on molecular diagnosis and genetic counseling for families affected by these disorders.

Introduction

Inborn errors of metabolism (IEM) are heritable metabolic disorders that result from biochemical alterations caused by mutations in genes encoding proteins involved in the various metabolic pathways in human. Depending on the nature of the biological function of the defective protein, mutations could result in loss of the enzymatic, hormonal, or transport function of the protein. In addition, mutations in certain genes could result in defects in the proper functioning of whole organelles such as the lysosomes, mitochondria, or peroxisomes. This explains the most striking characteristic of IEM disorders, which is their clinical and molecular heterogeneity (Sanjurjo et al., 2008). The majority of IEM disorders are autosomal recessive, but some are autosomal dominant or X-linked. Many IEM disorders carry serious clinical consequences for the affected neonates or young children, including mild to severe irreversible mental retardation, physical handicaps, or even early death (Sanjurjo et al., 2008). In most cases, early detection and appropriate management reduces or even eliminates the complications associated with many of these disorders and, therefore, many countries have developed national neonatal screening programs (Marsden et al., 2006). These screening programs allow for early diagnosis and intervention to prevent the associated complications. Due to their mode of inheritance, IEM disorders are generally more common in highly consanguineous populations such as those found in most Arab countries, including the United Arab Emirates (UAE) (Al-Gazali and Ali, 2010). It has been observed that the prevalence of this group of genetic disorders in the UAE is relatively high and heterogeneous in nature due to the ethnic diversity of the current UAE inhabitants (Ali et al., 2011). More than 40 metabolic disorders have been identified so far in the UAE population (Tadmouri et al., 2006; Abdulrazzaq et al., 2009; Hertecant et al., 2009; Ali et al., 2011). In addition, it was evident from several studies that the most common metabolic disorder in the UAE is phenylketonuria (PKU). Indeed, since its inclusion in the United Arab Emirates National Newborn Screening Program in 1995, it was found to have an incidence of ∼1 in 14,000 (Al-Hosani et al., 2008; Ali et al., 2011). Here, we report the mutation analysis of patients residing in the UAE who are affected by a spectrum of IEM disorders including PKU, lysinuric protein intolerance, maple syrup urine disease, carbamoyl phosphate synthase I deficiency, biotinidase deficiency, glutaric acidemia type I, medium chain acyl CoA dehydrogenase deficiency, multiple sulfatase deficiency, Wilson disease, cystinosis, Niemann-Pick disease type B, alpha-mannosidosis, GM2-gangliosidosis, and GM1-gangliosidosis. Several of the identified mutations are novel, while others have previously been documented in other parts of the world. The importance of these mutations is in their application as diagnostic tools for IEM disorders and for developing and implementing prevention strategies. In addition, reporting mutations, especially novel ones, is important for demonstrating the variations among different populations and ethnic groups.

Materials and Methods

Families and patients

The subjects described in this study were diagnosed and managed by clinicians at Tawam Hospital, Al-Ain, UAE. Blood samples were collected from each subject, and molecular testing was carried out at the specified diagnostic laboratory indicated in the next section. The medical records were examined, and the molecular genetics diagnostic and medical reports were used to compile the data presented in this article. A total of 34 patients from 34 different families have been studied from 2010 to 2011 Informed consent for the genetic testing was obtained from the guardians of all the patients. This project was approved by the Al-Ain Medical Human Research Ethics Committee-protocol number 10/09. Control samples were tested for the absence of the novel mutation in the populations.

Mutation analysis

After PCR amplification, direct genomic sequencing of the known genes responsible for the suspected IEM disorder was performed on samples from the affected individuals by accredited genetic diagnostic laboratories as follows: testing of mutations in the SLC7A7, BCKDHA, CPS1, GCDH, STS, SMPD1, MAN2B1, and GLB1 genes was carried out by members of the Genetic Diagnostic Network. Testing for mutations in the ASS gene was carried out by Universitäts Klinikum Münster, for the BTD gene by Mayo Clinic Medical Laboratories, for the PAH gene by Kinderspital Zürich, for the ACADM and ATP7B genes by the Sheffield Molecular Genetics Services, for the MPV17 gene by Stichting Klinisch-Genetisch Centrum Nijmegen, for the SUMF1 gene by Prevention Genetics, for the CTNS gene by the Arabian Diagnostic Laboratory, for the HEXB gene by Emory Genetics Laboratory, for the SLC26A3 gene by University of Helsinki Medical Genetics Department, and for the MC4R gene by the Department of Medicine and Clinical Biochemistry at the University of Cambridge.

Results

Mutation screening was performed on patients from 34 UAE national and expatriate families. Thirty-eight mutations causing various IEM disorders were identified as summarized in Tables 1–4. Nine of those mutations are novel, including two missense mutations (c.1034C>G in the SLC7A7 gene and c.278A>C in the MPV17 gene), three premature stop codon mutations (c.1590dupT in the CPS1 gene, c.2368C>T and c.2119C>T in the MAN2B1 gene), and four putative splice site mutations (c.499+1G>C in the SLC7A7 gene, c.603-2A>G in the SUMF1 gene, c.681+1G>A in the CTNS gene, and c.970-2A>G in the PAH gene).

Table 1.

Mutations Identified in Amino Acid Metabolic Disorders in the United Arab Emirates Population

Disorder (OMIM)	Gene access number	Nucleotide change	Protein change	National origin	Novelty
Lysinuric protein intolerance MIM 222700	SLC7A7 NM_003982	c.499+1G>C c.1034C>G	Splice p.P345R	UAE Jordan	Novel Novel
Citrullinemia MIM 215700	ASS NM_000050.4	c.349G>A	p.G117S	Pakistan	Reported Berning et al., 2008
Maple Syrup urine disease MIM 248600	BCKDHA NM_000709.3	Deletion of exons 2–4	-	Egypt	Reported Quental et al., 2008
Carbamoyl phosphate synthase I deficiency MIM 238970	CPS1 NM_001875.2	c.1590dupT	p.Val531CysfsX9	Oman	Novel
Biotinidase deficiency MIM 266150	BTD NM_000060	c.1330G>C/c.557G>A	p.D444H/p.C186Y	UAE	Reported Pomponio et al., 2000
Glutaric Acidemia I MIM 231670	GCDH NM_013976.2	c.505+1G>A	Splice	Palestine	Reported Christensen et al., 2004
		c.1169G>C	p.G390A	Iran	Reported Mushimoto et al., 2011

SLC7A7, solute carrier family 7 member 7; ASS, arginino succinate synthetase; BCKDHA, branched chain ketoacid dehydrogenase E1 alpha; CPS1, carbamoyl-phosphate synthase 1; BTD, biotidinase; GCDH, glutaryl-coadehydrgenase; UAE, United Arab Emirates; OMIM, Online Mendelian Inheritance in Man.

Table 4.

Mutations Identified in Lysosomal Storage Disorder and Miscellaneous Metabolic Disorder in the United Arab Emirates Population

Disorder (OMIM)	Gene access number	Nucleotide change	Protein change	National origin	Novelty
Lysosomal storage disorder
Multiple sulfatase deficiency MIM 272200	SUMF1 NM_001164674.1	c.603-2A>G	Splice	Iran	Novel
Cystinosis MIM 219800	CTNS NM_001031681.2	c.681+1G>A (IVS9+1G>A)	Splice	UAE	Novel
Niemann-Pick type B MIM 257220	SMPD1 NM_000543.4	c.1244C>T	p.A415V	UAE	Reported Lan et al., 2009
Alpha–mannosidosis MIM 248500	MAN2B1 NM_000528	c.2368C>T c.2119C>T	p.Q790X p.Q707X	UAE UAE	Novel Novel
Sandhoff GM2-gangliosidosis MIM 268800	HEXB NM_000521.3	16 kb deletion		Unknown	Reported Zhang et al., 1994
GM1-gangliosidosis MIM 230500	GLB1 NM_000404.2	c.914+4A>G	Splice	Palestine	Reported Georgiou et al., 2004
Miscellaneous disorder
Congenital chloride diarrhea 1 MIM 214700	SLC26A3 NM_000111.2	c.559G>T	p.G187X	UAE	Reported Höglund et al., 1998
Melanocortin-4 Receptor deficiency MIM 601665	MC4R NM_005912.2	c.485C>T	p.T187I	UAE	Reported Tan et al., 2009

SUMF1, sulfatase modifying factor 1; CTNS, cystinosis nephropathic; SMPD1, sphingomyelin phosphodiesterase; MAN2B1, mannosidase alpha class 2B member 1; HEXB, hexosaminidase B; GLB1, galactosidase beta 1; SLC26A3, solute carrier family 26 member 3; MC4R, melanocortin 4 receptor.

None of the novel variations were found in Arab controls. In addition, all the novel mutations were predicted by SIFT and Polyphen-2 softwares to be pathogenic. The change of the G on position +1 in the SLC7A7 and CTNS genes, and the change of the A on position −2 in the SUMF1 and PAH genes were predicted to disrupt the normal splice donor site, leading to alterations and premature degradation of the proteins. Concerning the three premature stop codons c.1590dupT (p.V531CysfsX9) in the CPS1 gene, c.2368C>T (p.Q790×) and c.2119 C>T (p.Q707×) in the MAN2B1 gene, these mutations are predicted to cause frame shifts resulting in premature stop codons. Therefore, they are likely to result in either truncated proteins or diminished quantity of mRNA due to mRNA decay by the nonsense mediated decay mechanism. The new missense variation (c.1034C>G) in the SLC7A7 gene was predicted to lead to the substitution of the proline residue by an arginine residue at position 345 of the resulting SLC7A7 protein (p.P345R). In silico prediction using Polyphen-2 and mutation tester programmes suggested that this mutation is pathogenic. In addition to the novel mutations, we identified 29 previously reported mutations. The majority of the mutations reported in our study were missense mutations. Only 7 of the 38 mutations were compound heterozygote, and the rest were homozygous. In the MAN2B1 gene responsible for alpha-mannosidosis, we report two novel premature stop codon mutations (Table 1). Compound heterozygous mutations were detected in the BTD and PAH genes, which are responsible for biotinidase deficiency and PKU, respectively (Tables 1 and 2). Among all mutations reported here, 15 were found in the PAH gene encoding the phenylalanine hydroxylase enzyme; 6 of them were compound heterozygous, and only one was novel (Table 2). Of the mutations identified, two were found in mitochondrial energy disorders, one in lipid disorders, one in steroid disorders, and one in metal disorders (Table 3). Moreover, seven mutations were found as the cause of lysosomal storage disorders, four of which are novel (Table 4).

Table 2.

Mutations Identified in Phenylketonuria (MIM 261600) Disorder in the United Arab Emirates Population

Nucleotide change	Protein change	National origin	Novelty
c.970-2A>G	Splice	Palestine	Novel
c.842+1G>A (IVS7+1G>A)	Splice	UAE	Reported Dianzani et al., 1991
c.592-613del22/C.581T>G	p.Y198Sfs136/p.L194R	Egypt	Reported Aurora et al., 2009
c.168+5G>C (IVS2+5G>C)	Splice	UAE	Reported Santos et al., 2008
c.592-613del22/c.721C>T	p.Y198Sfs136/p.R241C	Egypt	Reported Aurora et al., 2009
c.143T>C/c.842C>T	p.L48S/p.P281L	Egypt	Reported Karacic et al., 2009
c.143T>C/c.727C>T	p.L48S/p.R243×	Palestine	Reported Karacic et al., 2009
c.169-171delGAG/c.1066-11G>A	p.E57del/Splice	Palestine	Reported Dobrowolski et al., 2011
c.691>C	p.S231P	Yemen	Reported Dianzani et al., 1992
c.441+5G>T (IVS4+5G>T)	Splice	Iran	Reported Zekanowski et al., 1996
c.472C>T	p.R231W	India	Reported Takarada et al., 1993
c.1055del G/c.1066 11G>A (IVS10-11G>A)	p.G352Vfs/Splice	Morocco	Reported Benit et al., 1994

Numbering is based on PAH NM_000277.1 where the A of the start codon ATG is number 1.

Table 3.

Mutations Identified in Lipid, Steroid, Metal, and Mitochondrial Energy Metabolic Disorders in the United Arab Emirates Population

Disorder (OMIM)	Gene access number	Nucleotide change	Protein change	National origin	Novelty
Lipid disorder
Chylomicronemia familial MIM 118830	GPIHBP1 NM_178172.3	c.149G>A	p.G56Y	Oman	Reported Franssen et al., 2010
Steroid disorder
X-linked Ichthyosis MIM 308100	STS NM_000351.3	Deletion of the entire gene	—	Iraq	Reported Aviram-Goldring et al., 2000
Mitochondrial energy disorders
Medium chain acyl CoA dehydrogenase deficiency MIM 201450	ACADM NM_000016.4	c.985A>G	p.K985E	Palestine	Reported Matsubara and Kure, 2003
Mitochondrial DNA depletion syndrome MIM 256810	MPV17 NM_002437.4	c.278A>C	p.G93P	Syria	Novel
Metal disorder
Wilson disease MIM 277900	ATP7B NM 000053.2	c.122A>G	p.N41S	Palestine	Reported Deguti et al., 2004

GPIHBP1, glycosylphosphatidylinositol-anchored high density lipoprotein-binding protein 1; STS, steroidsulfatase (microsomal) isozyme S; ACA, acyl-CoAdehydrogenase; ATP7B, ATPase Cu (2+)-transporting beta polypeptide.

Discussion

The national Emirati inhabitants are ethnically diverse with ancestries from the north and south of the Arabian Peninsula, Iran, Baluchistan, and East Africa. However, the majority of the current eight million inhabitants are expatriates from the Asian subcontinent, the Middle East, Africa, and Europe. In spite of this mixture of populations, intermarriages between the different groups are rare, while consanguineous marriages within the local Emirati and within the other Arab populations are still the norm. This has led to the formation of population isolates and the appearance of cases of recessive conditions such as IEM disorders (Al-Gazali et al., 2005; Al-Gazali et al., 2006). Furthermore, hospital-based studies in the Sultanate of Oman, a country that is ethnically and geographically close to the UAE, has indicated a high prevalence of IEM disorders (Joshi et al., 2002; Joshi and Venugopalan, 2007).

Of the mutations reported in this study, only seven were compound heterozygote, and the rest were homozygous; this high percentage of homozygous mutations is most probably due to the high rates of consanguinity in these populations. The most commonly found IEM disorder in the UAE is PKU (Al-Hosani et al., 2008). Here, we report 15 mutations responsible for PKU in the UAE (Table 2), 1 of which is novel. The remaining mutations are compound heterozygous and have been previously reported in other populations (Al-Hosani et al., 2008). The large number of possible allelic combinations for this gene makes predicting the phenotype difficult (Dipple and Mccabe, 2000; Kasnauskiene et al., 2003; Kim et al., 2006; Bercovich et al., 2008; Daniele et al., 2009). Previously performed in vitro expression analysis studies have demonstrated a large range of residual activities among different mutations, of which ∼75% are null mutations (Waters et al., 1998, Jennings et al., 2000, Waters, 2003). It has been proposed that the PAH genotype is not a rigorous predictor for clinical progression of the condition in PKU patients. Many factors can influence the phenotypic variation in PKU, such as inter-individual variations in intestinal absorption, hepatic uptake of dietary phenylalanine, rate of incorporation of phenylalanine into proteins, rates of influx of phenylalanine across the blood brain barrier, and interactions of the PAH gene with other loci (Kayaalp et al., 1997, Scriver, 2007). Correlation studies are difficult to perform because of the high allelic heterogeneity and broad phenotypic variability. In any case, our findings are important for carrier screening and for implementing prevention strategies.

Our study supports that PKU is indeed the most frequent metabolic disorder in the UAE population. More than 500 mutations in the PAH gene have been described worldwide, and most PKU patients are compound heterozygote (Santos et al., 2010). In addition, a study by Ali et al. (2011) reported seven mutations in PAH in the UAE population, six of which had previously been reported in European populations and one in a Chinese patient. In fact, this variation is mainly due to the considerable allelic heterogeneity in the PAH gene locus and to the heterogeneity of the UAE population. In conclusion, our results demonstrate that there is a high diversity of mutations responsible for PKU and that there is no predominant mutation in the UAE population.

In addition to the novel mutation in PKU, we identified two novel premature stop codon mutations in the MAN2B1 gene responsible for alpha-mannosidosis (Table 1). Alpha-mannosidosis is one of more than 40 distinct lysosomal storage diseases. These mutations are likely to result in either truncated proteins or diminished quantity of mRNA due to mRNA decay by the nonsense mediated decay mechanism. We also report several mutations responsible for other lysosomal storage disorders and disorders of lipid, steroid, metal, and mitochondrial energy as well as miscellaneous disorders (Tables 3 and 4). The p.D444H mutation identified in the BTD gene in the UAE population has been previously reported in Turkish patients (Pomponio et al., 2000). Moreover, we report a novel duplication mutation in the CPS1 gene (Table 1). The majority of the previously reported mutations in this gene are missense, and only 7% are duplications and small insertions. Therefore, our report adds a new duplication, bringing the total number of mutations found in the CPS1 gene to 223; thereby confirming the high molecular heterogeneity in this disorder.

With reports of new mutations constantly appearing, the field of metabolic disorders continues to expand. Early diagnosis remains essential to prevent organ damage or death, and newborn screening programmes provide the opportunity for universal identification of these conditions. Testing for at-risk relatives is possible and will be facilitated if the disease-causing mutations in the family are known. Thus, practicing physicians need to be aware of new metabolic disorders that could be amenable to screening because of emerging therapeutic options. In addition, documenting mutations will help in setting up molecular screening programmes in the short run, and providing opportunities for additional diagnostic and preventive services.

Acknowledgments

The authors are indebted to the families for their invaluable cooperation and to the molecular geneticists and clinicians at the various diagnostic laboratories for carrying out the testing. The laboratories of B.R.A. and S.A.A. are funded by the UAE and Sultan Qaboos Universities GCC grant number CL/SQU/UAEU/10/02.

Disclosure Statement

No competing financial interests exist.