Cloning and sequence analysis of SLC25A13 transcripts in human amniocytes

ZHANG Zhan-Hui; ZHAO Xin-Jing; SONG Yuan-Zong; TANG Xiao-Mei; ZHA Qing-Bing

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Chinese Journal of Contemporary Pediatrics ›› 2012, Vol. 14 ›› Issue (3) : 221-225.

EXPERIMENTAL RESEARCH

Cloning and sequence analysis of SLC25A13 transcripts in human amniocytes

ZHANG Zhan-Hui, ZHAO Xin-Jing, SONG Yuan-Zong, TANG Xiao-Mei, ZHA Qing-Bing

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Abstract

OBJECTIVE: This research intends to amplify the entire coding region sequences of SLC25A13 mRNA which encodes citrin, and to investigate sequence features of the transcripts for this gene in cultured human amniocytes. This study will provide laboratory evidence for prenatal diagnosis of neonatal intrahepatic cholestasis caused by citrin deficiency (NICCD) at mRNA level. METHODS: One amniocyte sample was collected from a pregnant woman who underwent prenatal diagnosis of citrin deficiency and whose fetus has proven a carrier of 851del4 mutation by genomic DNA analysis. Another amniocyte sample, as a control, was from a fetus without family history of citrin deficiency. Total RNA was extracted from cultured amniocytes, cDNA was synthesized, and then nested-PCR was performed to amplify the entire coding region sequences of SLC25A13. The PCR products were cloned and analyzed by sequencing. RESULTS: The entire coding region of SLC25A13 gene was successful amplified from two cultured human amniocytes. The splice variant of SLC25A13, SLCA (normal mRNA), was identified in the two samples. SLCB (CAG insertion between exon 9-10) was identified in the control. SLCC (exon 5-11 skipping), but not transcriptional product from the allele with 851del4 mutation, was identified in the 851del4 mutation carrier. CONCLUSIONS: This study demonstrated that the entire coding region of SLC25A13 cDNA can be successfully amplified from two cultured human amniocytes, and revealed exon 5-11 skipping as a novel SLC25A13 transcript. Normal mRNA predominated in the transcripts in normal control and 851del4 mutation carrier, suggesting that the two fetuses were not at risk for NICCD. These SLC25A13 transcription features provided laboratory evidence for prenatal diagnosis of NICCD.

Key words

SLC25A13 / Nest PCR / Neonatal intrahepatic cholestasis caused by citrin deficiency / Amniocyte

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ZHANG Zhan-Hui, ZHAO Xin-Jing, SONG Yuan-Zong, TANG Xiao-Mei, ZHA Qing-Bing. Cloning and sequence analysis of SLC25A13 transcripts in human amniocytes[J]. Chinese Journal of Contemporary Pediatrics. 2012, 14(3): 221-225

References

［1］Kobayashi K, Sinasac DS, Iijima M, Boright AP, Begum L, Lee JR, et al. The gene mutated in adult-onset type II citrullinaemia encodes a putative mitochondrial carrier protein［J］. Nat Genet, 1999, 22(2): 159-163.

［2］Saheki T, Kobayashi K. Mitochondrial aspartate glutamate carrier (citrin) deficiency as the cause of adult-onset type II citrullinemia (CTLN2) and idiopathic neonatal hepatitis (NICCD)［J］. J Hum Genet, 2002, 47(7): 333-341.

［3］宋元宗, 小林圭子. citrin缺陷病［J］. 实用儿科临床杂志, 2008, 23(20):1564-1565.

［4］Lu YB, Kobayashi K, Ushikai M, Tabata A, Iijima M, Li MX, et al. Frequency and distribution in East Asia of 12 mutations identified in the SLC25A13 gene of Japanese patients with citrin deficiency［J］. J Hum Genet, 2005, 50(7): 338-346.

［5］Kobayshi K, Ushikai M, Song YZ, Gao HZ, Sheng JS, Tabata A, et al. Overview of citrin deficiency: SLC25A13 mutations and the frequency［J］. J Appl Clin Pediatr, 2008, 23(20): 1553-1557.

［6］Song YZ, Li BX, Hao H, Xin RL, Zhang T, Zhang CH, et al. Selective screening for inborn errors of metabolism and secondary methylmalonic aciduria in pregnancy at high risk district of neural tube defects: a human metabolome study by GC-MS in China［J］. Clin Biochem, 2008, 41(7-8): 616-620.

［7］宋元宗，牛饲美晴，小林圭子，佐伯武赖. 小儿胆汁淤积性肝病的病因学特征［J］. 中华儿科杂志, 2009, 47(8):624-627.

［8］Dimmock D, Maranda B, DionisiVici C, Wang J, Kleppe S, Fiermonte G, et al. Citrin deficiency, a perplexing global disorder［J］. Mol Genet Metab, 2009, 96(1): 44-49.

［9］宋元宗, 郝虎, 牛饲美晴, 柳国胜, 肖昕, 佐伯武赖, 等. 疑难病研究-citrin缺陷导致的新生儿肝内胆汁淤积症［J］. 中国当代儿科杂志, 2006, 8(2):125-128.

［10］Song YZ, Deng M, Chen FP, Wen F, Guo L, Cao SL, et al. Genotypic and phenotypic features of citrin deficiency: five-year experience in a Chinese pediatric center［J］. Int J Mol Med, 2011, 28(1): 33-40.

［11］Song YZ, Li BX, Chen FP, Liu SR, Sheng JS, Ushikai M, et al. Neonatal intrahepatic cholestasis caused by citrin deficiency: clinical and laboratory investigation of 13 subjects in mainland of China［J］. Dig Liver Dis, 2009, 41(9): 683-689.

［12］Tabata A, Sheng JS, Ushikai M, Song YZ, Gao HZ, Lu YB, et al. Identification of 13 novel mutations including a retrotransposal insertion in SLC25A13 gene and frequency of 30 mutations found in patients with citrin deficiency［J］. J Hum Genet, 2008, 53(6): 534-545.

［13］温鹏强, 王国兵, 陈占玲, 崔冬, 袁泉, 宋萍, 等. Citrin 缺陷导致的新生儿肝内胆汁淤积症SLC25A13基因分析［J］. 中国当代儿科杂志, 2011, 13(4):306-308.

［14］宋元宗, 郭丽, 杨艳玲, 韩连书, 小林圭子, 佐伯武赖. citrin缺陷导致的生长发育落后和血脂异常:一种新的临床表现型［J］. 中国当代儿科杂志, 2009, 11(5):328-332.

［15］Zhao XJ, Tang XM, Zha QB, Shi SS, Song YZ, Xiao XM. Prenatal diagnosis of citrin deficiency in a Chinese family with a fatal proband［J］. Tohoku J Exp Med, 2011, 225(4): 273-276.

［16］邢雅智, 邱文娟, 叶军, 韩连书, 许姗姗, 张惠文, 等. citrin缺陷导致的新生儿肝内胆汁淤积症临床和SLC25A13基因突变的研究［J］. 中华医学遗传学杂志, 2010, 27(2):180-185.

［17］Shigeta T, Kasahara M, Kimura T, Fukuda A, Sasaki K, Arai K, et al. Liver transplantation for an infant with neonatal intrahepatic cholestasis caused by citrin deficiency using heterozygote living donor［J］. Pediatr Transplant, 2010, 14(7): E86-E88.

［18］Ohura T, Kobayashi K, Tazawa Y, Abukawa D, Sakamoto O, Tsuchiya S, et al. Clinical pictures of 75 patients with neonatal intrahepatic cholestasis caused by citrin deficiency (NICCD)［J］. J Inherit Metab Dis, 2007, 30(2): 139-144.

［19］Tokuhara D, Iijima M, Tamamori A, Ohura T, Takaya J, Maisawa S, et al. Novel diagnostic approach to citrin deficiency: analysis of citrin protein in lymphocytes［J］. Mol Genet Metab, 2007, 90(1): 30-36.

［20］陈剑虹, 林静吟. 羊膜腔穿刺术在产前诊断地中海贫血中的应用［J］. 国际医药卫生导报, 2009, 15(13): 44-46.

［21］黄莹, 李东明. 改良羊水原位培养技术在产前诊断中的应用［J］. 广西医学, 2008, 30(12): 1852-1853.

［22］Wang ET, Sandberg R, Luo S, Khrebtukova I, Zhang L, Mayr C, et al. Alternative isoform regulation in human tissue transcriptomes［J］. Nature, 2008, 456(7221): 470-476.

［23］Weiss C, Zeng Y, Huang J, Sobocka MB, Rushbrook JI. Bovine NAD+-dependent isocitrate dehydrogenase: alternative splicing and tissue-dependent expression of subunit 1［J］. Biochemistry, 2000, 39(7): 1807-1816.

［24］Hosler R, Kerick M, Mah N, Hultschig C, Richter G, Bretz F, et al. Alterations of pre-mRNA splicing in human inflammatory bowel disease［J］. Eur J Cell Biol, 2011, 90(6-7): 603-611.

［25］Hansson O, Zhou Y, Renstrom E, Osmark P. Molecular function of TCF7L2: Consequences of TCF7L2 splicing for molecular function and risk for type 2 diabetes［J］. Curr Diab Rep, 2010, 10(6): 444-451.

［26］Maquat LE. Nonsense-mediated mRNA decay: splicing, translation and mRNP dynamics［J］. Nat Rev Mol Cell Biol, 2004, 5(2): 89-99.

［27］Frischmeyer PA, Dietz HC. Nonsense-mediated mRNA decay in health and disease［J］. Hum Mol Genet, 1999, 8(10): 1893-1900.