A Rare Missense Mutation and a Polymorphism with High Frequency in LDLR Gene among Iranian Patients with Familial Hypercholesterolemia

Tajamolian, Masoud; Kolahdouz, Parisa; Nikpour, Parvaneh; Forouzannia, Seyed Khalil; Sheikhha, Mohammad Hasan; Yazd, Ehsan Farashahi

A Rare Missense Mutation and a Polymorphism with High Frequency in LDLR Gene among Iranian Patients with Familial Hypercholesterolemia

Document Type : Original Article

Authors

¹ Department of Genetics, Faculty of Medicine; Medical Genetics Research Center, Yazd, Iran

² Yazd Clinical and Research Center for Infertility, Yazd, Iran

³ Department of Genetics and Molecular Biology, Faculty of Medicine, Isfahan University of Medical Sciences, Isfahan, Iran

⁴ Yazd Cardiovascular Research Center, Shahid Sadoughi University of Medical Sciences, Yazd, Iran

⁵ Department of Genetics, Faculty of Medicine; Yazd Clinical and Research Center for Infertility, Yazd, Iran

Abstract

Background: Familial hypercholesterolemia (FH) is a disorder that is inherited by autosomal dominant pattern. The main cause of FH disease is the occurrence of mutations in low-density lipoprotein receptor (LDLR) gene sequence, as well as apolipoprotein B and proprotein convertase subtilisin/kexin type 9 genes, located in the next ranks, respectively. Materials and Methods: Forty-five unrelated Iranian patients with FH were screened using a high-resolution melting (HRM) method for exon 9 along with intron/exon boundaries of LDLR gene. Samples with shift in resultant HRM curves were compared to normal ones, sequenced, and analyzed. Results: Our findings revealed a missense mutation c. 1246C>T and a known variant IVS9-30C>T (rs1003723) that was recognized in 71% of the patients (22%: homozygous and 49%: heterozygous genotypes). In silico analysis, predicted the pathological effect of the c. 1246C>T mutation in LDLR protein structure, but IVS9-30C>T variant had no predicted effect on splice site and branch point function. Conclusion: FH is a hereditary type of hypercholesterolemia that leads to premature cardiovascular disease and atherosclerosis, and early diagnosis is needed. We detected a rare missense mutation (1246C>T) and a common single nucleotide polymorphism (SNP) in the Iranian population. These reports could help in the genetic diagnosis and counseling of FH patients.

Keywords

References

1.	Chahil JK, Lye SH, Bagali PG, Alex L. A novel pathogenic variant of the LDLR gene in the Asian population and its clinical correlation with familial hypercholesterolemia. Mol Biol Rep 2012;39:7831-8.
2.	Goldstein JL, Hobbs HH, Brown MS. Familial hypercholesterolemia. In: The Metabolic and Molecular Bases of Inherited Disease. 8^th ed. New York: McGraw-Hill Book Co.; 2001. p. 2863-913.
3.	Goldstein JL, Hobbs HH, Brown MS. Familial hypercholester-olemia. In: Scriver CR, Beaudet AL, Sly WS, Valle D, editors. The Metabolic Basis of Inherited Disease. New York, NY: McGraw-Hill Book Co; 1995. p. 1981-2030.
4.	Varret M, Abifadel M, Rabès JP, Boileau C. Genetic heterogeneity of autosomal dominant hypercholesterolemia. Clin Genet 2008;73:1-13.
5.	Brown MS, Goldstein JL. A receptor-mediated pathway for cholesterol homeostasis. Science 1986;232:34-47.
6.	Usifo E, Leigh SE, Whittall RA, Lench N, Taylor A, Yeats C, et al. Low-density lipoprotein receptor gene familial hypercholesterolemia variant database: Update and pathological assessment. Ann Hum Genet 2012;76:387-401.
7.	Palacios L, Grandoso L, Cuevas N, Olano-Martín E, Martinez A, Tejedor D, et al. Molecular characterization of familial hypercholesterolemia in Spain. Atherosclerosis 2012;221:137-42.
8.	Austin MA, Hutter CM, Zimmern RL, Humphries SE. Genetic causes of monogenic heterozygous familial hypercholesterolemia: A HuGE prevalence review. Am J Epidemiol 2004;160:407-20.
9.	Anderson RG. Joe Goldstein and Mike Brown: From cholesterol homeostasis to new paradigms in membrane biology. Trends Cell Biol 2003;13:534-9.
10.	Lye SH, Chahil JK, Bagali P, Alex L, Vadivelu J, Ahmad WA, et al. Genetic polymorphisms in LDLR, APOB, PCSK9 and other lipid related genes associated with familial hypercholesterolemia in Malaysia. PLoS One 2013;8:e60729.
11.	Ahmed W, Ajmal M, Sadeque A, Whittall RA, Rafiq S, Putt W, et al. Novel and recurrent LDLR gene mutations in Pakistani hypercholesterolemia patients. Mol Biol Rep 2012;39:7365-72.
12.	Al-Allaf FA, Coutelle C, Waddington SN, David AL, Harbottle R, Themis M. LDLR-Gene therapy for familial hypercholesterolaemia: Problems, progress, and perspectives. Int Arch Med 2010;3:36.
13.	Jarvik GP, Brunzell JD, Motulsky AG. Frequent detection of familial hypercholesterolemia mutations in familial combined hyperlipidemia. J Am Coll Cardiol 2008;52:1554-6.
14.	Farrokhi E, Shayesteh F, Asadi Mobarakeh S, Roghani Dehkordi F, Ghatreh Samani K, Hashemzadeh Chaleshtori M. Molecular characterization of Iranian patients with possible familial hypercholesterolemia. Indian J Clin Biochem 2011;26:244-8.
15.	Fard-Esfahani P, Khatami S. Familial hypercholesterolemia in Iran: A novel frameshift mutation in low density lipoprotein receptor (LDLR) gene. Iran J Pathol 2010;5:22-6.
16.	Fard-Esfahani P, Zeinali C, Rouhi-Dehboneh S, Taghikhani M, Khatami S.A novel mutation in exon 4 of the low density lipoprotein (LDL) receptor gene in an Iranian familial hypercholesterolemia patient. Iran Biomed J 2005;9:139-42.
17.	Shayesteh F, Farrokhi E, Shirani M, Modarresi M, Roghani F, Hashemzadeh M. The study of mutations of the 9 exons of LDLR gene in patients with familial hypercholesterolemia in Cheharmahal Bakhtiari province. Arak Univ Med Sci J 2011;13:30-7.
18.	Yuan G, Wang J, Hegele RA. Heterozygous familial hypercholesterolemia: An underrecognized cause of early cardiovascular disease. CMAJ 2006;174:1124-9.
19.	Ye J, Coulouris G, Zaretskaya I, Cutcutache I, Rozen S, Madden TL. Primer-BLAST: A tool to design target-specific primers for polymerase chain reaction. BMC Bioinformatics 2012;13:134.
20.	Whittall RA, Scartezini M, Li K, Hubbart C, Reiner Z, Abraha A, et al. Development of a high-resolution melting method for mutation detection in familial hypercholesterolaemia patients. Ann Clin Biochem 2010;47(Pt 1):44-55.
21.	Adzhubei IA, Schmidt S, Peshkin L, Ramensky VE, Gerasimova A, Bork P, et al. Amethod and server for predicting damaging missense mutations. Nat Methods 2010;7:248-9.
22.	Schwarz JM, Cooper DN, Schuelke M, Seelow D. MutationTaster2: Mutation prediction for the deep-sequencing age. Nat Methods 2014;11:361-2.
23.	Kumar P, Henikoff S, Ng PC. Predicting the effects of coding non-synonymous variants on protein function using the SIFT algorithm. Nat Protoc 2009;4:1073-81.
24.	Venselaar H, Te Beek TA, Kuipers RK, Hekkelman ML, Vriend G. Protein structure analysis of mutations causing inheritable diseases. An e-Science approach with life scientist friendly interfaces. BMC Bioinformatics 2010;11:548.
25.	Di Tommaso P, Moretti S, Xenarios I, Orobitg M, Montanyola A, Chang JM, et al. T-Coffee: A web server for the multiple sequence alignment of protein and RNA sequences using structural information and homology extension. Nucleic Acids Res 2011;39:W13-7.
26.	Hebsgaard SM, Korning PG, Tolstrup N, Engelbrecht J, Rouzé P, Brunak S. Splice site prediction in Arabidopsis thaliana pre-mRNA by combining local and global sequence information. Nucleic Acids Res 1996;24:3439-52.
27.	Desmet FO, Hamroun D, Lalande M, Collod-Béroud G, Claustres M, Béroud C. Human splicing finder: An online bioinformatics tool to predict splicing signals. Nucleic Acids Res 2009;37:e67.
28.	Dogan RI, Getoor L, Wilbur WJ, Mount SM. SplicePort – An interactive splice-site analysis tool. Nucleic Acids Res 2007;35:W285-91.
29.	Jeon H, Meng W, Takagi J, Eck MJ, Springer TA, Blacklow SC. Implications for familial hypercholesterolemia from the structure of the LDL receptor YWTD-EGF domain pair. Nat Struct Biol 2001;8:499-504.
30.	Springer TA. An extracellular beta-propeller module predicted in lipoprotein and scavenger receptors, tyrosine kinases, epidermal growth factor precursor, and extracellular matrix components. J Mol Biol 1998;283:837-62.
31.	Chiou KR, Charng MJ. Detection of mutations and large rearrangements of the low-density lipoprotein receptor gene in Taiwanese patients with familial hypercholesterolemia. Am J Cardiol 2010;105:1752-8.
32.	Etxebarria A, Benito-Vicente A, Palacios L, Stef M, Cenarro A, Civeira F, et al. Functional characterization and classification of frequent low-density lipoprotein receptor variants. Hum Mutat 2015;36:129-41.
33.	Andreotti G, Menashe I, Chen J, Chang SC, Rashid A, Gao YT, et al. Genetic determinants of serum lipid levels in Chinese subjects: A population-based study in Shanghai, China. Eur J Epidemiol 2009;24:763-74.
34.	Andreotti G, Chen J, Gao YT, Rashid A, Chen BE, Rosenberg P, et al. Polymorphisms of genes in the lipid metabolism pathway and risk of biliary tract cancers and stones: A population-based case-control study in Shanghai, China. Cancer Epidemiol Biomarkers Prev 2008;17:525-34.
35.	Ladewig E, Okamura K, Flynt AS, Westholm JO, Lai EC. Discovery of hundreds of mirtrons in mouse and human small RNA data. Genome Res 2012;22:1634-45.

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