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研究生:林道承
研究生(外文):Lin, Dao-Chen
論文名稱:利用外顯子定序來偵測新生兒糖尿病及第一型B糖尿病之突變和突變葡萄糖激酶之活性分析
論文名稱(外文):Detecting the Mutations of Neonatal Diabetes and Type 1B Diabetes Using Exome Sequencing and Analysis of Mutant Glucokinase Activity
指導教授:李燕晉李燕晉引用關係
指導教授(外文):Lee, Yann-Jinn
口試委員:莊育梩陳涵栩
口試委員(外文):Juang, Yue-LiChen, Harn-Shen
口試日期:2015-07-21
學位類別:碩士
校院名稱:馬偕醫學院
系所名稱:生物醫學研究所
學門:生命科學學門
學類:生物化學學類
論文種類:學術論文
論文出版年:2015
畢業學年度:103
語文別:英文
論文頁數:46
中文關鍵詞:外顯子定序新生兒糖尿病第一型B糖尿病葡萄糖激酶
外文關鍵詞:exome sequencingneonatal diabetes mellitustype 1B diabetes mellitusglucokinase
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背景
糖尿病是一種高血糖的代謝疾病。單基因糖尿病是由於基因突變導致胰島β細胞功能障礙引起的。傳統上,依照病人臨床上的表現和表型,利用Sanger定序檢測突變基因是一個公認的標準方法。次世代定序是一種高通量技術,能夠在短時間的一次操作中找到多個的基因突變。在最近,越來越多的研究使用這種新技術來檢測孟德爾遺傳疾病和單基因糖尿病的基因突變。

方法與材料
我們納入十四個華人患者診斷為新生兒糖尿病或是第1B型糖尿病。我們使用外顯子定序來檢測基因是否有突變。所有發現的變異經由GenePipe的VarioWatch來評估。我們排除在單一鹼基多型性資料庫或千人基因組計畫中已發現存在的變異。我們利用之前有報告過的致病基因及生物資訊學來分析所發現的突變。所有突變都經過Sanger定序確定。新發現的突變需進一步來分析功能確認。

結果
利用外顯子定序,在研究中,我們發現五個突變的基因在五個不同的病患中。
根據之前的研究報告,我們證實兩個突變的基因為致病性。我們利用大腸桿菌重建的野生及突變的葡萄糖激酶來證明了一個新的GCK基因突變會造成葡萄糖激酶活性的減少。在我們的研究中,我們證實了三個致病的突變基因。在我們十四個病人,基因突變的檢出率為21%。

結論
外顯子定序,一種次世代的定序,是一種的高通量技術,可以有效檢測中出新生兒糖尿病及第一型B糖尿病病患的多樣性突變基因。

Background
Diabetes mellitus is a metabolic disease of hyperglycemia and resulted from deficiency of insulin secretion and action. Monogenic diabetes is caused by genes mutations that induce pancreatic β-cell dysfunction. Traditionally, Sanger sequencing is a golden standard method to detect mutated genes by clinical manifestation and phenotype of the probands and their families. Next generation sequencing is a high throughput technology to find more mutations of the genes in one assay in a short time. In recently, more and more studies use this new technology to detect gene mutation in Mendelian disorder and monogenic diabetes.

Method and material
We enrolled forth Han Chinese patients of neonatal diabetes mellitus or type 1B diabetes mellitus. We used exome sequencing by Illumine HiSeq2000 to detect mutations. Variants were identified and assessed in VarioWatch of GenePipe. We excluded the variants in dbSNP or 1000 genomes project. We confirmed the variants as pathogenic by previous reports and bioinformatic analysis. All mutations were rechecked by Sanger sequencing. The finding of novel mutations in our study was proved by functional analysis.

Result
Five mutations genes in 5 individuals were found by exome sequencing in our study. We confirmed two mutations as pathogenicity by previous reports. We proved a novel mutation of GCK gene as pathogenic by reconstruct mutated glucokinase with less enzymatic activity comparing with wild type. Three mutated genes in three patients were discovered and confirmed in our study. The detection rate of mutation was 21% in our forth patients. 

Conclusion
Exome sequencing, a kind of next generation sequencing, is high throughput and robust to detect mutations in patients with neonatal diabetes or type 1B diabetes which is heterogeneous in genetic etiologies.

Abbreviations 1
Index to Tables 2
Index to Figures 3
Chinese Abstract 4
English Abstract 5
Chapter one Introduction 7
1.1 Background 7
1.2 Monogenic Diabetes Mellitus 8
1.3 Neonatal Diabetes Mellitus 9
1.4 Maturity onset diabetes of the young 10
1.5 Type 1B Diabetes Mellitus 11
1.6 Next Generation Sequencing for Diagnosis of Monogenic Diabetes 12
Chapter two Aim and Rationale of the Study 13
Chapter Three Material and Methods 14
3.1 Subjects 14
3.2 Exome Sequencing and Data Analysis 15
3.3 GCK Gene Cloning 16
3.4 Construction of Wild and Mutant Type GST-GCK 17
3.5 Production and Purification of GST-GCK Fusion Protein 18
3.6 Glucokinase Activity 19
Chapter Four Results 20
4.1 Data Analysis of Exome Sequencing 20
4.2 Glucokinase Activity Analysis 22
Chapter Five Discussion 23
Chapter Six Conclusions 26
Chapter Seven References 39



Aguilar-Bryan, L., and Bryan, J. (2008).Neonatal diabetes mellitus.Endocr. Rev. 29, 265–291.

American Diabetes Association (2010).Diagnosis and classification of diabetes mellitus.Diabetes Care 33 (Suppl. 1), S62–S69.

Babenko, A.P., Polak, M., Cavé, H., Busiah, K., Czernichow, P., Scharfmann, R., Bryan, J., Aguilar-Bryan, L., Vaxillaire, M., Froguel, P. et al. (2006).Activating mutations in the ABCC8 gene in neonatal diabetes mellitus. N. Engl. J. Med. 355, 456–466.

Bar, R.S., Muggeo, M., Roth, J., Kahn, C.R., Havrankova, J., andImperato-McGinley, J. (1978). Insulin resistance, acanthosis nigricans, and normal insulin receptors in a young woman: evidence for a postreceptor defect. J. Clin. Endocr.Metab. 47, 620-625.

Cheng, Y.C., Hsiao, F.C., Yeh, E.C., Lin, W.J., Tang, C.Y., Tseng, H.C., Wu, H.T., Liu, C.K., Chen, C.C., Yao, A. et al. (2012). VarioWatch: providing large-scale and comprehensive annotations on human genomic variants in the next generation sequencing era. Nucleic Acids Res. 40(Web Server issue), W76-81.

Colombo, C., Porzio, O., Liu, M., Massa, O., Vasta, M., Salardi, S., Beccaria, L., Monciotti, C., Toni, S., Barbetti, F. et al. (2008). Seven mutations in the human insulin gene linked to permanent neonatal/infancy-onset diabetes mellitus. J. Clin. Invest. 118, 2148–2156.

de Wet, H., Rees, M.G., Shimomura, K., Aittoniemi, J., Patch, A.M., Flanagan, S.E.,
Edghill, E.L., Flanagan, S.E., Patch, A.M., Boustred, C., Parrish, A., Shields, B., Shepherd, M.H., Hussain, K., Kapoor, R.R., Bell, G.I. et al.; Neonatal Diabetes International Collaborative Group, Hattersley, A.T., Ellard, S. (2008). Insulin mutation screening in 1,044 patients with diabetes: mutations in the INS gene are a common cause of neonatal diabetes but a rare cause of diabetes diagnosed in childhood or adulthood. Diabetes 57, 1034–1042.

Ellard, S., Hattersley, A.T., Sansom, M.S., and Ashcroft, F.M. (2007). Increased ATPase activity produced by mutations at arginine-1380 in nucleotide-binding domain 2 of ABCC8 causes neonatal diabetes. Proc. Natl. Acad. Sci. USA 104, 18988–18992.

Ellard, S., Flanagan, S.E., Girard, C.A., Patch, A., Harries, L.W., Parrish, A., Edghill, E.L., Mackay, D.J., Proks, P., Ashcroft F.M. et al. (2007). Permanent neonatal diabetes caused by dominant, recessive, or compound heterozygous SUR1 mutations with opposite functional effects. Am. J. Hum. Genet. 81, 375–382.

Ellard, S., Flanagan, S.E., Girard, C.A., Patch, A.M., Harries, L.W., Parrish, A., Edghill, E.L., Mackay, D.J., Proks, P., Ashcroft, F.M. et al. (2013). Improved genetic testing for monogenic diabetes using targeted next-generation sequencing.Diabetologia 56, 1958-1963.

Flanagan, S.E., Patch, A.M., Mackay, D.J., Edghill, E.L., Gloyn, A.L., Robinson, D., Shield, J.P., Temple, K., Ellard, S., and Hattersley, A.T. (2007). Mutations in ATP-sensitive K+ channel genes cause transient neonatal diabetes and permanent diabetes in childhood or adulthood. Diabetes 56, 1930–1937.

Galán, M., Vincent, O., Roncero, I., Azriel, S., Boix-Pallares, P., Delgado-Alvarez, E., Díaz-Cadórniga, F., Blázquez, E., and Navas, M.A. (2006). Effects of novel maturity-onset diabetes of the young (MODY)-associated mutations on glucokinase activity and protein stability. Biochem. J. 393, 389-96.

Garin, I., Edghill, E.L., Akerman, I., Rubio-Cabezas, O., Rica, I., Locke, J.M., Maestro, M.A., Alshaikh, A., Bundak, R., del Castillo, G. et al. (2010). Recessive mutations in the INS gene result in neonatal diabetes through reduced insulin biosynthesis. Proc. Natl Acad. Sci. USA 107, 3105–3110.

Gloyn, A.L., Pearson, E.R., Antcliff, J.F., Proks, P., Bruining, G.J., Slingerland, A.S., Howard, N., Srinivasan, S., Silva, J.M., Hattersley, A.T. et al. (2004).Activating mutations in the gene encoding the ATP-sensitive potassium-channel subunit Kir6.2 and permanent neonatal diabetes. N. Engl. J. Med. 350, 1838–1849.

Grada, A., and Weinbrecht, K., (2013). Next-Generation Sequencing: Methodology and Application. Journal of Investigative Dermatology 133, e11.

Gribble, F.M., andReimann, F. (2003). Sulfonylurea action revisited: the post-cloning era. Diabetologia 46, 875-891.

Grulich-Henn, J., Wagner, V., Thon, A., Schober, E., Marg, W., Kapellen, T.M., Haberland, H., Raile, K., Ellard, S., Holl, R.W. et al. (2010). Entities and frequency of neonatal diabetes: data from the diabetes documentation and quality management system (DPV). Diabet. Med. 27, 709–712.

Johansson, S., Irgens, H., Chudasama, K.K., Molnes, J., Aerts, J., Roque, F.S., Jonassen, I., Levy, S., Lima, K., Njølstad, P.R. et al. (2012). Exome sequencing and genetic testing for MODY.PLoS One 7, e38050.
Karges, B., Meissner, T., Icks, A., Kapellen, T. and Holl, R.W. (2012).Management of diabetes mellitus in infants. Nature Reviews Endocrinology 8, 201-211.

Klupa, T., Edghill, E.L., Nazim, J., Sieradzki, J., Ellard, S., Hattersley, A.T., and Malecki, M.T. (2005). The identification of a R201H mutation in KCNJ11, which encodes Kir6.2, and successful transfer to sustained-release sulphonylurea therapy in a subject with neonatal diabetes: evidence for heterogeneity of beta cell function among carriers of the R201H mutation. Diabetologia 48, 1029-1031.

Ledermann, H.M. (1995). Is maturity onset diabetes at young age (MODY) more common in Europethan previously assumed.Lancet 345, 648.

Liang, Y., Kesavan, P., Wang, L.Q., Niswender, K., Tanizawa, Y., Permutt, M.A., Magnuson, M.A., and Matschinsky, F.M. (1995). Variable effects of maturity-onset-diabetes-of-youth (MODY)-associated glucokinase mutations on substrate interactions and stability of the enzyme. Biochem. J. 309,167-173.

Mackay, D. J., and Temple, I. K. (2010). Transient neonatal diabetes mellitus type 1. Am. J. Med. Genet. C Semin. Med. Genet. 154C, 335–342.

Mackay, D.J., Callaway, J.L., Marks, S.M., White, H.E., Acerini, C.L., Boonen, S.E., Dayanikli, P., Firth, H.V., Goodship, J.A., Temple, I.K. et al. (2008). Hypomethylation of multiple imprinted loci in individuals with transient neonatal diabetes is associated with mutations in ZFP57. Nat. Genet. 40, 949–951.

Magnuson, M.A. (1990) Glucokinase gene structure.Functional implications of molecular genetic studies.Diabetes 39,523-527.

Matschinsky, F.M. (1990). Glucokinase as glucose sensor and metabolic signal generator in pancreatic beta-cells and hepatocytes. Diabetes 39, 647-652.

Metz, C., Cavé, H., Bertrand, A.M., Deffert, C., Gueguen-Giroux, B., Czernichow, P., and Polak, M.; NDM French Study Group. (2002) Neonatal diabetes mellitus: chromosomal analysis in transient and permanent cases. J. Pediatr. 141, 483–489.

Miller, D.T., Adam, M.P., Aradhya, S., Biesecker, L.G., Brothman, A.R., Carter, N.P., Church, D.M., Crolla, J.A., Eichler, E.E., Ledbetter, D.H. et al. (2010). Consensus statement: chromosomal microarray is a first-tier clinical diagnostic test for individuals with developmental disabilities or congenital anomalies. Am. J. Hum.Genet. 86, 749-764.

Moritani, M., Yokota, I., Tsubouchi, K., Takaya, R., Takemoto, K., Minamitani, K., Urakami, T., Kawamura, T., Kikuchi, N., Amemiya, S. et al.; Japanese Study Group of Insulin Therapy for Childhood and Adolescent Diabetes (JSGIT) (2013).Identification of INS and KCNJ11 gene mutations in type 1B diabetes in Japanese children with onset of diabetes before 5 years of age.Pediatr. Diabetes 14, 112-120.

Njølstad, P.R., Søvik, O., Cuesta-Muñoz, A., Bjørkhaug, L., Massa, O., Barbetti, F., Undlien, D.E., Shiota, C., Magnuson, M.A., Bell, G.I. et al. (2001).Neonatal diabetes mellitus due to complete glucokinase deficiency. N. Engl. J. Med. 344, 1588–1592.

Pearson, E.R., Flechtner, I., Njølstad, P.R., Malecki, M.T., Flanagan, S.E., Larkin, B., Ashcroft, F.M., Klimes, I., Codner, E., Hattersley, A.T. et al.; Neonatal Diabetes International Collaborative Group (2006). Switching from insulin to oral sulfonylureas in patients with diabetes due to Kir6.2 mutations. N. Engl. J. Med. 355, 467–477.

Polak, M., Dechaume, A., Cavé, H., Nimri, R., Crosnier, H., Sulmont, V., de Kerdanet, M., Scharfmann, R., Lebenthal, Y., Vaxillaire, M et al..; French ND (Neonatal Diabetes) Study Group (2008). Heterozygous missense mutations in the insulin gene are linked to permanent diabetes appearing in the neonatal period or in early infancy: a report from the French ND (Neonatal Diabetes) Study Group. Diabetes 57, 1115–1119.

Proks, P., Amanda, L. Arnold, A.L., Jan Bruining, J., Christophe Girard, C., Sarah, E. Flanagan, S.E., Brian Larkin, B., Colclough, K., Ellard, S. et al. (2006). A heterozygous activating mutation in the sulphonylurea receptor SUR1 (ABCC8) causes neonatal diabetes. Hum. Mol. Genet. 15, 1793–1800.

Rafiq, M., Flanagan, S.E., Patch, A.M., Shields, B.M., Ellard, S., and Hattersley, A.T.; Neonatal Diabetes International Collaborative Group (2008). Effective treatment with oral sulfonylureas in patients with diabetes due to sulfonylurea receptor 1 (SUR1) mutations. Diabetes Care 31, 204–209.

Robinson, J.T., Thorvaldsdóttir, H., Winckler, W., Guttman, M., Lander, E.S., Getz, G., and Mesirov, J.P. (2011).Integrative genomics viewer. Nat Biotechnol. 29, 24-26.

Ron, D. (2002). Proteotoxicity in the endoplasmic reticulum:
lessons from the Akita diabetic mouse. J.Clin. Invest. 109, 443–445.

Russo, L., Iafusco, D., Brescianini, S., Nocerino, V., Bizzarri, C., Toni, S., Cerutti, F., Monciotti, C., Pesavento, R., Barbetti, F. et al.; ISPED Early Diabetes Study Group (2011). Permanent diabetes during the first year of life: multiple gene screening in 54 patients. Diabetologia 54, 1693–1701.

Sagen, J.V., Raeder, H., Hathout, E., Shehadeh, N., Gudmundsson, K., Baevre, H., Abuelo, D., Phornphutkul, C., Molnes, J., Njølstad, P.R. et al. (2004). Permanent neonatal diabetes due to mutations in KCNJ11 encoding Kir6.2: patient characteristics and initial response to sulfonylurea therapy. Diabetes 53, 2713–2718.

Sakura, H., Ashcroft, S.J., Terauchi, Y., Kadowaki, T., and Ashcroft, F.M. (1998). Glucose modulation of ATP-sensitive K-currents in wild-type, homozygous and heterozygous glucokinase knock-out mice. Diabetologia 41, 654-659.

Senée, V., Chelala, C., Duchatelet, S., Feng, D., Blanc, H., Cossec, J.C., Charon, C., Nicolino, M., Boileau, P., Julier, C. et al. (2006). Mutations in GLIS3 are responsible for a rare syndrome with neonatal diabetes mellitus and congenital hypothyroidism. Nat Genet. 38, 682-687.

Shaffer, L.G. (2005). American College of Medical Genetics guideline on the cytogenetic evaluation of the individual with developmental delay or mental retardation.; American College of Medical Genetics Professional Practice and Guidelines Committee. Genet. Med. 7, 650-654.

Shendure, J. and Ji, H. (2008). Next-generation DNA sequencing. Nat.Biotechnol. 26, 1135-1145.
Shepherd, M., Sparkes, A.C., and Hattersley, A.T. (2001).Genetic testing in maturity onset diabetes ofthe young (MODY); a new challenge for the diabetic clinic.Pract. Diabetes 18, 16-21.

Shevell, M., Ashwal, S., Donley, D., Flint, J., Gingold, M., Hirtz, D., Majnemer, A., Noetzel, M., and Sheth, R.D. (2003). Practice parameter: evaluation of the child with global developmental delay: report of the Quality Standards Subcommittee of the American Academy of Neurology and The Practice Committee of the Child Neurology Society. Neurology 60, 367-380.

Slingerland, A.S., Shields, B.M., Flanagan, S.E., Bruining, G.J., Noordam, K., Gach, A., Mlynarski, W., Malecki, M.T., Hattersley, A.T., and Ellard, S. (2009). Referral rates for diagnostic testing support an incidence of permanent neonatal diabetes in three European countries of at least 1 in 260,000 live births. Diabetologia 52, 1683–1685.

Steiner, D.F., Tager, H.S., Nanjo, K., Chan, S.J., Rubenstein, A.H. (1995). In The
Metabolic Basis of Inherited Disease, Scriver, C., Beaudet, A., Sly, W., Valle, D. ed.
(McGraw–Hill, New York), Vol 1, 7th Ed, pp 897–904.

Støy, J., Edghill, E.L., Flanagan, S.E., Ye, H., Paz, V.P., Pluzhnikov, A., Below, J.E., Hayes, M.G., Cox, N.J., Bell, G.I. et al.; Neonatal Diabetes International Collaborative Group. (2007). Insulin gene mutations as a cause of permanent neonatal diabetes. Proc. Natl. Acad. Sci. USA 104, 15040–15044.

Tager, H., Given, B., Baldwin, D., Mako, M., Markese, J., Rubenstein, A., Olefsky, J., Kobayashi, M., Kolterman, O., Poucher, R. et al. (1979).A structurally abnormal insulin causing human diabetes. Nature 281, 122–125.

Taha, D., Barbar, M., Kanaan, H., and Williamson Balfe, J. (2003). Neonatal diabetes mellitus, congenital hypothyroidism, hepatic fibrosis, polycystic kidneys, and congenital glaucoma: a new autosomal recessive syndrome? Am. J. Med. Genet A. 15;122A, 269-273.

Temple, I.K., Gardner, R.J., Mackay, D.J., Barber, J.C., Robinson, D.O., and Shield, J.P. (2000). Transient neonatal diabetes: widening the understanding of the etiopathogenesis of diabetes. Diabetes 49, 1359–1366.
Thorvaldsdóttir, H., Robinson, J.T., and Mesirov, J.P. (2013). Integrative Genomics Viewer (IGV): high-performance genomics data visualization and exploration. Brief Bioinform. 14, 178-192.

Turkkahraman, D., Bircan, I., Tribble, N.D., Akçurin, S., Ellard, S., and Gloyn, A.L. (2008) Permanent neonatal diabetes mellitus caused by a novel homozygous (T168A) glucokinase (GCK) mutation: initial response to oral sulphonylurea therapy. J.Pediatr. 153, 122-126.

Wang, J., Takeuchi, T., Tanaka, S., Kubo, S.K., Kayo, T., Lu, D., Takata, K., Koizumi, A., and Izumi, T. (1999). A mutation in the insulin 2 gene induces diabetes with severe pancreatic β-cell dysfunction in the Mody mouse. J.Clin. Invest. 103, 27–37.

Yang, Y., Muzny, D.M., Reid, J.G., Bainbridge, M.N., Willis, A., Ward, P.A., Braxton, A., Beuten, J., Xia, F., Eng, C.M. et al. (2013). Clinical whole-exome sequencing for the diagnosis of mendelian disorders. N. Engl. J. Med. 369, 1502-11.

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