title: Lower Circulating Folate Induced by a Fidgetin Intronic Variant Is Associated With Reduced Congenital Heart Disease Susceptibility.
abstract: BACKGROUND
  Folate deficiency is an independent risk factor for congenital heart disease (CHD); however, the maternal plasma folate level is paradoxically not a good diagnostic marker. Genome-wide surveys have identified variants of nonfolate metabolic genes associated with the plasma folate level, suggesting that these genetic polymorphisms are potential risk factors for CHD.
  
  
  METHODS
  To examine the effects of folate concentration-related variations on CHD risk in the Han Chinese population, we performed 3 independent case-control studies including a total of 1489 patients with CHD and 1745 control subjects. The expression of the Fidgetin (FIGN) was detected in human cardiovascular and decidua tissue specimens with quantitative real-time polymerase chain reaction and Western blotting. The molecular mechanisms were investigated by luciferase reporter assays, surface plasmon resonance, and chromatin immunoprecipitation. FIGN-interacting proteins were confirmed by tandem affinity purification and coimmunoprecipitation. Proteasome activity and metabolite concentrations in the folate pathway were quantified with a commercial proteasome activity assay and immunoassays, respectively.
  
  
  RESULTS
  The +94762G>C (rs2119289) variant in intron 4 of theFIGNgene was associated with significant reduction in CHD susceptibility (P=5.1×10-14for the allele,P=8.5×10--13for the genotype). Analysis of combined samples indicated that CHD risks in individuals carrying heterozygous (GC) or homozygous (CC) genotypes were reduced by 44% (odds ratio [OR]=0.56; 95% confidence interval [CI]=0.47-0.67) and 66% (OR=0.34; 95% CI=0.23-0.50), respectively, compared with those with the major GG genotype. Minor C allele carriers who had decreased plasma folate levels exhibited significantly increasedFIGNexpression because the transcription suppressor CREB1 did not bind the alternative promoter ofFIGNisoform X3. Mechanistically, increased FIGN expression led to the accumulation of both reduced folate carrier 1 and dihydrofolate reductase via inhibition of their proteasomal degradation, which promoted folate absorption and metabolism.
  
  
  CONCLUSIONS
  We report a previously undocumented finding that decreased circulating folate levels induced by increased folate transmembrane transport and utilization, as determined by theFIGNintronic variant, serves as a protective mechanism against CHD. Our results may explain why circulating folate levels do not have a good diagnostic value.
authors.affiliation: From Obstetrics and Gynecology Hospital of Fudan University, State Key Laboratory of Genetic Engineering, School of Life Sciences and Institutes of Biomedical Sciences, Fudan University, Shanghai, China (D.W., Y.L., R.Z., H.L., S.-M.Z., H.W., J.-Y.Z.); Key Laboratory of Reproduction Regulation of NPFPC, Institute of Reproduction and Development and Children's Hospital of Fudan University, Fudan University, Shanghai, China (D.W., F.W., Y.L., R.Z., S.-M.Z., H.W., J.-Y.Z.); MOE Key Laboratory of Contemporary Anthropology, Collaborative Innovation Center for Genetics and Development, Fudan University, Shanghai, China (S.-M.Z., H.W., J.-Y.Z.); Department of Cardiothoracic Surgery, Second Hospital of Anhui Medical University, Anhui Medical University, Hefei, China (K.-H.S., H.T.); Cardiovascular Research Center, Anhui Medical University, Hefei, China (K.-H.S., H.T.); Institute of Cardiovascular Disease, General Hospital of Jinan Military Region, Jinan, China (W.D., B.Q.); Collaborative Innovation Center for Biotherapy, West China Hospital, Sichuan University, Chengdu, China (S.M.-Z., J.-Y.Z.); and Department of Neonatology, First Affiliated Hospital of Wenzhou Medical University, Wenzhou, China (D.W.).
authors.forename: Dan
authors.initials: D
authors.lastname: Wang
authors.affiliation: From Obstetrics and Gynecology Hospital of Fudan University, State Key Laboratory of Genetic Engineering, School of Life Sciences and Institutes of Biomedical Sciences, Fudan University, Shanghai, China (D.W., Y.L., R.Z., H.L., S.-M.Z., H.W., J.-Y.Z.); Key Laboratory of Reproduction Regulation of NPFPC, Institute of Reproduction and Development and Children's Hospital of Fudan University, Fudan University, Shanghai, China (D.W., F.W., Y.L., R.Z., S.-M.Z., H.W., J.-Y.Z.); MOE Key Laboratory of Contemporary Anthropology, Collaborative Innovation Center for Genetics and Development, Fudan University, Shanghai, China (S.-M.Z., H.W., J.-Y.Z.); Department of Cardiothoracic Surgery, Second Hospital of Anhui Medical University, Anhui Medical University, Hefei, China (K.-H.S., H.T.); Cardiovascular Research Center, Anhui Medical University, Hefei, China (K.-H.S., H.T.); Institute of Cardiovascular Disease, General Hospital of Jinan Military Region, Jinan, China (W.D., B.Q.); Collaborative Innovation Center for Biotherapy, West China Hospital, Sichuan University, Chengdu, China (S.M.-Z., J.-Y.Z.); and Department of Neonatology, First Affiliated Hospital of Wenzhou Medical University, Wenzhou, China (D.W.).
authors.forename: Feng
authors.initials: F
authors.lastname: Wang
authors.affiliation: From Obstetrics and Gynecology Hospital of Fudan University, State Key Laboratory of Genetic Engineering, School of Life Sciences and Institutes of Biomedical Sciences, Fudan University, Shanghai, China (D.W., Y.L., R.Z., H.L., S.-M.Z., H.W., J.-Y.Z.); Key Laboratory of Reproduction Regulation of NPFPC, Institute of Reproduction and Development and Children's Hospital of Fudan University, Fudan University, Shanghai, China (D.W., F.W., Y.L., R.Z., S.-M.Z., H.W., J.-Y.Z.); MOE Key Laboratory of Contemporary Anthropology, Collaborative Innovation Center for Genetics and Development, Fudan University, Shanghai, China (S.-M.Z., H.W., J.-Y.Z.); Department of Cardiothoracic Surgery, Second Hospital of Anhui Medical University, Anhui Medical University, Hefei, China (K.-H.S., H.T.); Cardiovascular Research Center, Anhui Medical University, Hefei, China (K.-H.S., H.T.); Institute of Cardiovascular Disease, General Hospital of Jinan Military Region, Jinan, China (W.D., B.Q.); Collaborative Innovation Center for Biotherapy, West China Hospital, Sichuan University, Chengdu, China (S.M.-Z., J.-Y.Z.); and Department of Neonatology, First Affiliated Hospital of Wenzhou Medical University, Wenzhou, China (D.W.).
authors.forename: Kai-Hu
authors.initials: KH
authors.lastname: Shi
authors.affiliation: From Obstetrics and Gynecology Hospital of Fudan University, State Key Laboratory of Genetic Engineering, School of Life Sciences and Institutes of Biomedical Sciences, Fudan University, Shanghai, China (D.W., Y.L., R.Z., H.L., S.-M.Z., H.W., J.-Y.Z.); Key Laboratory of Reproduction Regulation of NPFPC, Institute of Reproduction and Development and Children's Hospital of Fudan University, Fudan University, Shanghai, China (D.W., F.W., Y.L., R.Z., S.-M.Z., H.W., J.-Y.Z.); MOE Key Laboratory of Contemporary Anthropology, Collaborative Innovation Center for Genetics and Development, Fudan University, Shanghai, China (S.-M.Z., H.W., J.-Y.Z.); Department of Cardiothoracic Surgery, Second Hospital of Anhui Medical University, Anhui Medical University, Hefei, China (K.-H.S., H.T.); Cardiovascular Research Center, Anhui Medical University, Hefei, China (K.-H.S., H.T.); Institute of Cardiovascular Disease, General Hospital of Jinan Military Region, Jinan, China (W.D., B.Q.); Collaborative Innovation Center for Biotherapy, West China Hospital, Sichuan University, Chengdu, China (S.M.-Z., J.-Y.Z.); and Department of Neonatology, First Affiliated Hospital of Wenzhou Medical University, Wenzhou, China (D.W.).
authors.forename: Hui
authors.initials: H
authors.lastname: Tao
authors.affiliation: From Obstetrics and Gynecology Hospital of Fudan University, State Key Laboratory of Genetic Engineering, School of Life Sciences and Institutes of Biomedical Sciences, Fudan University, Shanghai, China (D.W., Y.L., R.Z., H.L., S.-M.Z., H.W., J.-Y.Z.); Key Laboratory of Reproduction Regulation of NPFPC, Institute of Reproduction and Development and Children's Hospital of Fudan University, Fudan University, Shanghai, China (D.W., F.W., Y.L., R.Z., S.-M.Z., H.W., J.-Y.Z.); MOE Key Laboratory of Contemporary Anthropology, Collaborative Innovation Center for Genetics and Development, Fudan University, Shanghai, China (S.-M.Z., H.W., J.-Y.Z.); Department of Cardiothoracic Surgery, Second Hospital of Anhui Medical University, Anhui Medical University, Hefei, China (K.-H.S., H.T.); Cardiovascular Research Center, Anhui Medical University, Hefei, China (K.-H.S., H.T.); Institute of Cardiovascular Disease, General Hospital of Jinan Military Region, Jinan, China (W.D., B.Q.); Collaborative Innovation Center for Biotherapy, West China Hospital, Sichuan University, Chengdu, China (S.M.-Z., J.-Y.Z.); and Department of Neonatology, First Affiliated Hospital of Wenzhou Medical University, Wenzhou, China (D.W.).
authors.forename: Yang
authors.initials: Y
authors.lastname: Li
authors.affiliation: From Obstetrics and Gynecology Hospital of Fudan University, State Key Laboratory of Genetic Engineering, School of Life Sciences and Institutes of Biomedical Sciences, Fudan University, Shanghai, China (D.W., Y.L., R.Z., H.L., S.-M.Z., H.W., J.-Y.Z.); Key Laboratory of Reproduction Regulation of NPFPC, Institute of Reproduction and Development and Children's Hospital of Fudan University, Fudan University, Shanghai, China (D.W., F.W., Y.L., R.Z., S.-M.Z., H.W., J.-Y.Z.); MOE Key Laboratory of Contemporary Anthropology, Collaborative Innovation Center for Genetics and Development, Fudan University, Shanghai, China (S.-M.Z., H.W., J.-Y.Z.); Department of Cardiothoracic Surgery, Second Hospital of Anhui Medical University, Anhui Medical University, Hefei, China (K.-H.S., H.T.); Cardiovascular Research Center, Anhui Medical University, Hefei, China (K.-H.S., H.T.); Institute of Cardiovascular Disease, General Hospital of Jinan Military Region, Jinan, China (W.D., B.Q.); Collaborative Innovation Center for Biotherapy, West China Hospital, Sichuan University, Chengdu, China (S.M.-Z., J.-Y.Z.); and Department of Neonatology, First Affiliated Hospital of Wenzhou Medical University, Wenzhou, China (D.W.).
authors.forename: Rui
authors.initials: R
authors.lastname: Zhao
authors.affiliation: From Obstetrics and Gynecology Hospital of Fudan University, State Key Laboratory of Genetic Engineering, School of Life Sciences and Institutes of Biomedical Sciences, Fudan University, Shanghai, China (D.W., Y.L., R.Z., H.L., S.-M.Z., H.W., J.-Y.Z.); Key Laboratory of Reproduction Regulation of NPFPC, Institute of Reproduction and Development and Children's Hospital of Fudan University, Fudan University, Shanghai, China (D.W., F.W., Y.L., R.Z., S.-M.Z., H.W., J.-Y.Z.); MOE Key Laboratory of Contemporary Anthropology, Collaborative Innovation Center for Genetics and Development, Fudan University, Shanghai, China (S.-M.Z., H.W., J.-Y.Z.); Department of Cardiothoracic Surgery, Second Hospital of Anhui Medical University, Anhui Medical University, Hefei, China (K.-H.S., H.T.); Cardiovascular Research Center, Anhui Medical University, Hefei, China (K.-H.S., H.T.); Institute of Cardiovascular Disease, General Hospital of Jinan Military Region, Jinan, China (W.D., B.Q.); Collaborative Innovation Center for Biotherapy, West China Hospital, Sichuan University, Chengdu, China (S.M.-Z., J.-Y.Z.); and Department of Neonatology, First Affiliated Hospital of Wenzhou Medical University, Wenzhou, China (D.W.).
authors.forename: Han
authors.initials: H
authors.lastname: Lu
authors.affiliation: From Obstetrics and Gynecology Hospital of Fudan University, State Key Laboratory of Genetic Engineering, School of Life Sciences and Institutes of Biomedical Sciences, Fudan University, Shanghai, China (D.W., Y.L., R.Z., H.L., S.-M.Z., H.W., J.-Y.Z.); Key Laboratory of Reproduction Regulation of NPFPC, Institute of Reproduction and Development and Children's Hospital of Fudan University, Fudan University, Shanghai, China (D.W., F.W., Y.L., R.Z., S.-M.Z., H.W., J.-Y.Z.); MOE Key Laboratory of Contemporary Anthropology, Collaborative Innovation Center for Genetics and Development, Fudan University, Shanghai, China (S.-M.Z., H.W., J.-Y.Z.); Department of Cardiothoracic Surgery, Second Hospital of Anhui Medical University, Anhui Medical University, Hefei, China (K.-H.S., H.T.); Cardiovascular Research Center, Anhui Medical University, Hefei, China (K.-H.S., H.T.); Institute of Cardiovascular Disease, General Hospital of Jinan Military Region, Jinan, China (W.D., B.Q.); Collaborative Innovation Center for Biotherapy, West China Hospital, Sichuan University, Chengdu, China (S.M.-Z., J.-Y.Z.); and Department of Neonatology, First Affiliated Hospital of Wenzhou Medical University, Wenzhou, China (D.W.).
authors.forename: Wenyuan
authors.initials: W
authors.lastname: Duan
authors.affiliation: From Obstetrics and Gynecology Hospital of Fudan University, State Key Laboratory of Genetic Engineering, School of Life Sciences and Institutes of Biomedical Sciences, Fudan University, Shanghai, China (D.W., Y.L., R.Z., H.L., S.-M.Z., H.W., J.-Y.Z.); Key Laboratory of Reproduction Regulation of NPFPC, Institute of Reproduction and Development and Children's Hospital of Fudan University, Fudan University, Shanghai, China (D.W., F.W., Y.L., R.Z., S.-M.Z., H.W., J.-Y.Z.); MOE Key Laboratory of Contemporary Anthropology, Collaborative Innovation Center for Genetics and Development, Fudan University, Shanghai, China (S.-M.Z., H.W., J.-Y.Z.); Department of Cardiothoracic Surgery, Second Hospital of Anhui Medical University, Anhui Medical University, Hefei, China (K.-H.S., H.T.); Cardiovascular Research Center, Anhui Medical University, Hefei, China (K.-H.S., H.T.); Institute of Cardiovascular Disease, General Hospital of Jinan Military Region, Jinan, China (W.D., B.Q.); Collaborative Innovation Center for Biotherapy, West China Hospital, Sichuan University, Chengdu, China (S.M.-Z., J.-Y.Z.); and Department of Neonatology, First Affiliated Hospital of Wenzhou Medical University, Wenzhou, China (D.W.).
authors.forename: Bin
authors.initials: B
authors.lastname: Qiao
authors.affiliation: From Obstetrics and Gynecology Hospital of Fudan University, State Key Laboratory of Genetic Engineering, School of Life Sciences and Institutes of Biomedical Sciences, Fudan University, Shanghai, China (D.W., Y.L., R.Z., H.L., S.-M.Z., H.W., J.-Y.Z.); Key Laboratory of Reproduction Regulation of NPFPC, Institute of Reproduction and Development and Children's Hospital of Fudan University, Fudan University, Shanghai, China (D.W., F.W., Y.L., R.Z., S.-M.Z., H.W., J.-Y.Z.); MOE Key Laboratory of Contemporary Anthropology, Collaborative Innovation Center for Genetics and Development, Fudan University, Shanghai, China (S.-M.Z., H.W., J.-Y.Z.); Department of Cardiothoracic Surgery, Second Hospital of Anhui Medical University, Anhui Medical University, Hefei, China (K.-H.S., H.T.); Cardiovascular Research Center, Anhui Medical University, Hefei, China (K.-H.S., H.T.); Institute of Cardiovascular Disease, General Hospital of Jinan Military Region, Jinan, China (W.D., B.Q.); Collaborative Innovation Center for Biotherapy, West China Hospital, Sichuan University, Chengdu, China (S.M.-Z., J.-Y.Z.); and Department of Neonatology, First Affiliated Hospital of Wenzhou Medical University, Wenzhou, China (D.W.). zhaosm@fudan.edu.cn wanghy@fudan.edu.cn zhaojy@fudan.edu.cn.
authors.forename: Shi-Min
authors.initials: SM
authors.lastname: Zhao
authors.affiliation: From Obstetrics and Gynecology Hospital of Fudan University, State Key Laboratory of Genetic Engineering, School of Life Sciences and Institutes of Biomedical Sciences, Fudan University, Shanghai, China (D.W., Y.L., R.Z., H.L., S.-M.Z., H.W., J.-Y.Z.); Key Laboratory of Reproduction Regulation of NPFPC, Institute of Reproduction and Development and Children's Hospital of Fudan University, Fudan University, Shanghai, China (D.W., F.W., Y.L., R.Z., S.-M.Z., H.W., J.-Y.Z.); MOE Key Laboratory of Contemporary Anthropology, Collaborative Innovation Center for Genetics and Development, Fudan University, Shanghai, China (S.-M.Z., H.W., J.-Y.Z.); Department of Cardiothoracic Surgery, Second Hospital of Anhui Medical University, Anhui Medical University, Hefei, China (K.-H.S., H.T.); Cardiovascular Research Center, Anhui Medical University, Hefei, China (K.-H.S., H.T.); Institute of Cardiovascular Disease, General Hospital of Jinan Military Region, Jinan, China (W.D., B.Q.); Collaborative Innovation Center for Biotherapy, West China Hospital, Sichuan University, Chengdu, China (S.M.-Z., J.-Y.Z.); and Department of Neonatology, First Affiliated Hospital of Wenzhou Medical University, Wenzhou, China (D.W.). zhaosm@fudan.edu.cn wanghy@fudan.edu.cn zhaojy@fudan.edu.cn.
authors.forename: Hongyan
authors.initials: H
authors.lastname: Wang
authors.affiliation: From Obstetrics and Gynecology Hospital of Fudan University, State Key Laboratory of Genetic Engineering, School of Life Sciences and Institutes of Biomedical Sciences, Fudan University, Shanghai, China (D.W., Y.L., R.Z., H.L., S.-M.Z., H.W., J.-Y.Z.); Key Laboratory of Reproduction Regulation of NPFPC, Institute of Reproduction and Development and Children's Hospital of Fudan University, Fudan University, Shanghai, China (D.W., F.W., Y.L., R.Z., S.-M.Z., H.W., J.-Y.Z.); MOE Key Laboratory of Contemporary Anthropology, Collaborative Innovation Center for Genetics and Development, Fudan University, Shanghai, China (S.-M.Z., H.W., J.-Y.Z.); Department of Cardiothoracic Surgery, Second Hospital of Anhui Medical University, Anhui Medical University, Hefei, China (K.-H.S., H.T.); Cardiovascular Research Center, Anhui Medical University, Hefei, China (K.-H.S., H.T.); Institute of Cardiovascular Disease, General Hospital of Jinan Military Region, Jinan, China (W.D., B.Q.); Collaborative Innovation Center for Biotherapy, West China Hospital, Sichuan University, Chengdu, China (S.M.-Z., J.-Y.Z.); and Department of Neonatology, First Affiliated Hospital of Wenzhou Medical University, Wenzhou, China (D.W.). zhaosm@fudan.edu.cn wanghy@fudan.edu.cn zhaojy@fudan.edu.cn.
authors.forename: Jian-Yuan
authors.initials: JY
authors.lastname: Zhao
mesh_terms: D000074183:ATPases Associated with Diverse Cellular Activities; D000251:Adenosine Triphosphatases / Q000235:genetics* / Q000378:metabolism; D000818:Animals; D044466:Asian People / Q000235:genetics*; D015415:Biomarkers / Q000097:blood; D016022:Case-Control Studies; D016009:Chi-Square Distribution; D002675:Child, Preschool; D002681:China / Q000453:epidemiology; D017362:Cyclic AMP Response Element-Binding Protein / Q000235:genetics / Q000378:metabolism; D015536:Down-Regulation; D005260:Female; D005492:Folic Acid / Q000097:blood*; D005787:Gene Frequency; D056726:Genetic Association Studies; D020022:Genetic Predisposition to Disease; D057809:HEK293 Cells; D006330:Heart Defects, Congenital / Q000175:diagnosis / Q000208:ethnology / Q000235:genetics / Q000517:prevention & control*; D006579:Heterozygote; D006720:Homozygote; D006801:Humans; D007438:Introns; D016015:Logistic Models; D008297:Male; D026901:Membrane Transport Proteins / Q000378:metabolism; D008869:Microtubule-Associated Proteins; D016017:Odds Ratio; D010641:Phenotype; D020641:Polymorphism, Single Nucleotide; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D065840:Protective Factors; D059748:Proteolysis; D034622:RNA Interference; D051381:Rats; D018570:Risk Assessment; D012307:Risk Factors; D013762:Tetrahydrofolate Dehydrogenase / Q000378:metabolism; D014158:Transcription, Genetic; D014162:Transfection
chemical_list: D015415:Biomarkers; C115163:CREB1 protein, human; D017362:Cyclic AMP Response Element-Binding Protein; D026901:Membrane Transport Proteins; D008869:Microtubule-Associated Proteins; C120043:SLC19A2 protein, human; D005492:Folic Acid; D013762:Tetrahydrofolate Dehydrogenase; D046988:Proteasome Endopeptidase Complex; D000251:Adenosine Triphosphatases; D000074183:ATPases Associated with Diverse Cellular Activities; C000613810:FIGN protein, human
keywords: congenital heart diseases; folate transmembrane transport; folate utilization; proteasome