title: A three-step programmed method for the identification of causative gene mutations of maturity onset diabetes of the young (MODY).
abstract: To establish a three-step programmed method to find gene mutations related to maturity onset diabetes of the young (MODY). Target region capture and next-generation sequencing (NGS) were performed using customized oligonucleotide probes designed to capture suspected genes for MODY in 11 probands with clinically diagnosed MODY. The suspected associations of certain genes with MODY were then confirmed by Sanger sequencing in the probands and their family members. Finally, to validate variants of one of the genes of interest (glucokinase, GCK) as pathogenic mutations, protein function editing by the variant genes was assessed. In the target region capture and NGS phase, a total of nine variants of seven genes (GCK, WFS1, SLC19A2, SH2B1, SERPINB4, RFX6, and GATA6) were identified in eight probands. Two heterozygous GCK mutations located on the same allele (p.Leu77Arg and p.Val101Met) were identified in a MODY family. Sanger sequencing was used to confirm the variants identified by NGS to be present in probands and their diabetic family members, but not in non-diabetic family members. Finally, enzyme kinetic and thermal stability analyses revealed that the p.Leu77Arg mutation or the p.Leu77Arg mutation in combination with the p.Val101Met mutation inactivates GCK function and stability, while mutation of p.Val101Met alone does not. The p.Leu77Arg but not p.Val101Met GCK mutation is therefore considered a pathogenic mutation associated with MODY. Genetic screening coupled with gene-editing protein function testing is an effective and reliable method by which causative gene mutations of MODY can be identified.
authors.affiliation: Department of Endocrinology, Beijing Tongren Hospital, Capital Medical University, Beijing 100730, China.
authors.forename: Qian
authors.initials: Q
authors.lastname: Li
authors.affiliation: Department of Endocrinology, Beijing Tongren Hospital, Capital Medical University, Beijing 100730, China; Beijing Key Laboratory of Diabetes Research and Care, Beijing 100730, China.
authors.forename: Xi
authors.initials: X
authors.lastname: Cao
authors.affiliation: Department of Endocrinology, Beijing Tongren Hospital, Capital Medical University, Beijing 100730, China.
authors.forename: Hai-Yan
authors.initials: HY
authors.lastname: Qiu
authors.affiliation: Department of Endocrinology, Beijing Tongren Hospital, Capital Medical University, Beijing 100730, China; Beijing Key Laboratory of Diabetes Research and Care, Beijing 100730, China.
authors.forename: Jing
authors.initials: J
authors.lastname: Lu
authors.affiliation: Beijing Genomic Institute (BGI) in Shenzhen, Shenzhen 518083, China.
authors.forename: Rui
authors.initials: R
authors.lastname: Gao
authors.affiliation: Beijing Genomic Institute (BGI) in Shenzhen, Shenzhen 518083, China.
authors.forename: Chao
authors.initials: C
authors.lastname: Liu
authors.affiliation: Department of Endocrinology, Beijing Tongren Hospital, Capital Medical University, Beijing 100730, China; Beijing Key Laboratory of Diabetes Research and Care, Beijing 100730, China.
authors.forename: Ming-Xia
authors.initials: MX
authors.lastname: Yuan
authors.affiliation: Department of Endocrinology, Beijing Tongren Hospital, Capital Medical University, Beijing 100730, China.
authors.forename: Guang-Ran
authors.initials: GR
authors.lastname: Yang
authors.affiliation: Department of Endocrinology, Beijing Tongren Hospital, Capital Medical University, Beijing 100730, China; Beijing Key Laboratory of Diabetes Research and Care, Beijing 100730, China. Electronic address: jkyang@ccmu.edu.cn.
authors.forename: Jin-Kui
authors.initials: JK
authors.lastname: Yang
mesh_terms: D000293:Adolescent; D000328:Adult; D017668:Age of Onset; D003924:Diabetes Mellitus, Type 2 / Q000175:diagnosis* / Q000235:genetics* / Q000503:physiopathology; D057075:Enzyme Assays; D005260:Female; D015870:Gene Expression; D005820:Genetic Testing; D005941:Glucokinase / Q000235:genetics*; D006579:Heterozygote; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D008297:Male; D008875:Middle Aged; D009154:Mutation; D015345:Oligonucleotide Probes / Q000737:chemistry; D010375:Pedigree; D010641:Phenotype
chemical_list: D015345:Oligonucleotide Probes; D005941:Glucokinase
keywords: Genetic screening; MODY; NGS; Protein function testing