CCAAT/enhancer binding protein α,β and δ gene variants: associations with obesity related phenotypes in the Leeds Family Study

Abstract

Objective: To identify novel polymorphisms in the genes encoding the transcription factors CCAAT/enhancer binding protein α, β and δ ( CEBPA, CEBPB, CEBPD) and investigate associations between polymorphisms and obesity-related phenotypes. Methods: Denaturing high-performance liquid chromatography (HPLC) was used to screen for novel gene variants and polymorphisms were genotyped in stored DNA from participants of the Leeds Family Study (537 subjects from 89 families). Genotype and haplotype analyses were carried out in STATA and PBAT, respectively. Results: Twenty-five polymorphisms were identified; 11 in CEBPA, 12 in CEBPB and 2 in CEBPD. Several allelic variants were associated at a nominal 5% level with waist-to-hip ratio (-919G>A in CEBPA, -412G>T and 646C>T in CEBPB), leptin (1558G>A in CEBPA, -1051A>G and 1383T>- in CEBPB) and adiponectin (1382G>T and 1903G>T in CEBPB). Effects of CEBPA and CEBPB allelic variants were independent, but variants within each gene were in linkage disequilibrium. Several associations were observed between other obesity-related traits and allelic variants in CEBPA and CEBPB, but not CEBPD. Conclusion: These findings suggest that common allelic variants in CEBPA and CEBPB could influence abdominal obesity and related metabolic abnormalities associated with type 2 diabetes and cardiovascular disease in healthy White Northern European families, although results require independent confirmation.

Keywords

CCAAT enhancer binding proteins polymorphism obesity metabolic abnormalities

References

Despres JP , Lemieux I. ( 2006) Abdominal obesity and metabolic syndrome. Nature 444: 881-7.

Freeman MS , Mansfield MW , Barrett JH , Grant PJ ( 2002) Heritability of features of the insulin resistance syndrome in a community-based study of healthy families. Diabet Med 19: 994-9.

Farooqi IS , O’Rahilly S. ( 2007) Genetic factors in human obesity. Obes Rev 8(Suppl 1): 37-40.

Rose KM , Newman B. , Mayer-Davis EJ , Selby JV ( 1998) Genetic and behavioral determinants of waist-hip ratio and waist circumference in women twins. Obes Res 6: 383-92.

Clement K. , Vaisse C. , Lahlou N. , et al. (1998) A mutation in the human leptin receptor gene causes obesity and pituitary dysfunction. Nature 392: 398-401.

Savage DB , Agostini M. , Barroso I. , et al. (2002) Digenic inheritance of severe insulin resistance in a human pedigree. Nat Genet 31: 379-84.

Farooqi IS , Yeo GS , Keogh JM , et al. (2000) Dominant and recessive inheritance of morbid obesity associated with melanocortin 4 receptor deficiency. J Clin Invest 106: 271-9.

Frayling TM , Timpson NJ , Weedon MN , et al. (2007) A common variant in the FTO gene is associated with body mass index and predisposes to childhood and adult obesity. Science 316: 889-94.

Thorleifsson G. , Walters GB , Gudbjartsson DF , et al. (2009) Genome-wide association yields new sequence variants at seven loci that associate with measures of obesity . Nat Genet 41: 18-24.

10.

Jonsson A. , Renstrom F. , Lyssenko V. , et al. (2009) Assessing the effect of interaction between an FTO variant (rs9939609) and physical activity on obesity in 15,925 Swedish and 2,511 Finnish adults. Diabetologia 52: 1334-8.

11.

Cao Z. , Umek RM , McKnight SL ( 1991) Regulated expression of three C/EBP isoforms during adipose conversion of 3T3-L1 cells. Genes Dev 5: 1538-52.

12.

Ramji DP , Foka P. ( 2002) CCAAT/enhancer-binding proteins: structure, function and regulation. Biochem J 365: 561-75.

13.

Westmacott A. , Burke ZD , Oliver G. , Slack JM , Tosh D. (2006) C/EBPalpha and C/EBPbeta are markers of early liver development. Int J Dev Biol 50: 653-57.

14.

Tanaka T. , Yoshida N. , Kishimoto T. , Akira S. ( 1997) Defective adipocyte differentiation in mice lacking the C/EBPbeta and/ or C/EBPdelta gene. EMBO J 16: 7432-43.

15.

Flodby P. , Barlow C. , Kylefjord H. , Ahrlund-Richter L. , Xanthopoulos KG ( 1996) Increased hepatic cell proliferation and lung abnormalities in mice deficient in CCAAT/ enhancer binding protein alpha. J Biol Chem 271: 24753-60.

16.

Wang ND , Finegold MJ , Bradley A. , et al. (1995) Impaired energy homeostasis in C/EBP alpha knockout mice. Science 269: 1108-12.

17.

Jurado LA , Song S. , Roesler WJ , Park EA ( 2002) Conserved amino acids within CCAAT enhancer-binding proteins (C/ EBP(alpha) and beta) regulate phosphoenolpyruvate carboxykinase (PEPCK) gene expression. J Biol Chem 277: 27606-12.

18.

El Jack AK , Hamm JK , Pilch PF , Farmer SR (1999) Reconstitution of insulin-sensitive glucose transport in fibroblasts requires expression of both PPARgamma and C/ EBPalpha. J Biol Chem 274: 7946-51.

19.

Kaestner KH , Christy RJ , Lane MD ( 1990) Mouse insulin-responsive glucose transporter gene: characterization of the gene and trans-activation by the CCAAT/enhancer binding protein. Proc Natl Acad Sci U S A 87: 251-5.

20.

Park SK , Oh SY , Lee MY , Yoon S. , Kim KS , Kim JW ( 2004) CCAAT/enhancer binding protein and nuclear factor-Y regulate adiponectin gene expression in adipose tissue. Diabetes 53: 2757-66.

21.

Seo JB , Noh MJ , Yoo EJ , et al. (2003) Functional characterization of the human resistin promoter with adipocyte determination- and differentiation-dependent factor 1/sterol regulatory element binding protein 1c and CCAAT enhancer binding protein-alpha. Mol Endocrinol 17: 1522-33.

22.

Miller SG , De Vos P. , Guerre-Millo M. , et al. (1996) The adipocyte specific transcription factor C/EBPalpha modulates human ob gene expression. Proc Natl Acad Sci U S A 93: 5507-11.

23.

Alam T. , An MR , Mifflin RC , Hsieh CC , Ge X. , Papaconstantinou J. ( 1993) Trans-activation of the alpha 1-acid glycoprotein gene acute phase responsive element by multiple isoforms of C/EBP and glucocorticoid receptor. J Biol Chem 268: 15681-8.

24.

Akira S. , Isshiki H. , Sugita T. , et al. (1990) A nuclear factor for IL-6 expression (NF-IL6) is a member of a C/EBP family. EMBO J 9: 1897-906.

25.

Fernandez-Rachubinski FA , Weiner JH , Blajchman MA (1996) Regions flanking exon 1 regulate constitutive expression of the human antithrombin gene. J Biol Chem 271: 29502-12.

26.

Figueiredo MS , Brownlee GG (1995) Cis-acting elements and transcription factors involved in the promoter activity of the human factor VIII gene. J Biol Chem 270: 11828-38.

27.

Freeman MS , Mansfield MW , Barrett JH , Grant PJ ( 2003) Insulin resistance: an atherothrombotic syndrome. The Leeds Family Study. Thromb Haemost 89: 161-8.

28.

Freeman MS , Mansfield MW , Barrett JH , Grant PJ ( 2002) Genetic contribution to circulating levels of hemostatic factors in healthy families with effects of known genetic polymorphisms on heritability. Arterioscler Thromb Vasc Biol 22: 506-10.

29.

Hudson BI , Stickland MH , Futers TS , Grant PJ ( 2001) Effects of novel polymorphisms in the RAGE gene on transcriptional regulation and their association with diabetic retinopathy. Diabetes 50: 1505-11.

30.

Barrett JC , Fry B. , Maller J. , Daly MJ ( 2005) Haploview: analysis and visualization of LD and haplotype maps. Bioinformatics 21: 263-5.

31.

Lange C. , DeMeo D. , Silverman EK , Weiss ST , Laird NM ( 2004) PBAT: tools for family-based association studies. Am J Hum Genet 74: 367-9.

32.

Timchenko N. , Wilson DR , Taylor LR , et al. (1995) Autoregulation of the human C/EBP alpha gene by stimulation of upstream stimulatory factor binding. Mol Cell Biol 15: 1192-202.

33.

Cleutjens CB , van Eekelen CC , van Dekken H. , et al. (1993) The human C/EBP delta (CRP3/CELF) gene: structure and chromosomal localization. Genomics 16: 520-3.

34.

Chambers JC , Elliott P. , Zabaneh D. , et al. (2008) Common genetic variation near MC4R is associated with waist circumference and insulin resistance . Nat Genet 40: 716-18.

35.

Linhart HG , Ishimura-Oka K. , DeMayo F. , et al. (2001) C/ EBPalpha is required for differentiation of white, but not brown, adipose tissue. Proc Natl Acad Sci U S A 98: 12532-7.

36.

Hendricks-Taylor LR , Bachinski LL , Siciliano MJ , et al. (1992) The CCAAT/enhancer binding protein (C/EBP alpha) gene (CEBPA) maps to human chromosome 19q13.1 and the related nuclear factor NF-IL6 (C/EBP beta) gene (CEBPB) maps to human chromosome 20q13.1. Genomics 14: 12-17.

37.

Lee JH , Reed DR , Li WD , et al. (1999) Genome scan for human obesity and linkage to markers in 20q13. Am J Hum Genet 64: 196-209.

38.

Dong C. , Wang S. , Li WD , Li D. , Zhao H. , Price RA ( 2003) Interacting genetic loci on chromosomes 20 and 10 influence extreme human obesity. Am J Hum Genet 72: 115-24.

39.

Klupa T. , Malecki MT , Pezzolesi M. , et al. (2000) Further evidence for a susceptibility locus for type 2 diabetes on chromosome 20q13.1-q13.2. Diabetes 49: 2212-16.

40.

Feitosa MF , Rice T. , North KE , et al. Pleiotropic QTL on chromosome 19q13 for triglycerides and adiposity: the HERITAGE Family Study. Atherosclerosis 185: 426-32.

41.

Van Tilburg JH , Sandkuijl LA , Strengman E. , et al. (2003) A genome-wide scan in type 2 diabetes mellitus provides independent replication of a susceptibility locus on 18p11 and suggests the existence of novel Loci on 2q12 and 19q13 . J Clin Endocrinol Metab 88: 2223-30.

42.

Lembertas AV , Perusse L. , Chagnon YC , et al. (1997) Identification of an obesity quantitative trait locus on mouse chromosome 2 and evidence of linkage to body fat and insulin on the human homologous region 20q. J Clin Invest 100: 1240-7.

43.

An P. , Freedman BI , Hanis CL , et al. (2005) Genome-wide linkage scans for fasting glucose, insulin, and insulin resistance in the National Heart, Lung, and Blood Institute Family Blood Pressure Program: evidence of linkages to chromosome 7q36 and 19q13 from meta-analysis. Diabetes 54: 909-14.

44.

Yang J. , Croniger CM , Lekstrom-Himes J. , et al. (2005) Metabolic response of mice to a postnatal ablation of CCAAT/enhancer-binding protein alpha. J Biol Chem 280: 38689-99.

45.

Schroeder-Gloeckler JM , Rahman SM , Janssen RC , et al. (2007) CCAAT/enhancer-binding protein beta deletion reduces adiposity, hepatic steatosis, and diabetes in Lepr(db/ db) mice. J Biol Chem 282: 15717-29.

46.

Millward CA , Heaney JD , Sinasac DS , et al. (2007) Mice with a deletion in the gene for CCAAT/enhancer-binding protein beta are protected against diet-induced obesity. Diabetes 56: 161-7.

47.

Ling H. , Waterworth DM , Stirnadel HA , et al. (2009) Genome-wide linkage and association analyses to identify genes influencing adiponectin levels: the GEMS Study. Obesity (Silver Spring) 17: 737-44.

48.

Dai F. , Sun G. , Aberg K. , et al. (2008) A whole genome linkage scan identifies multiple chromosomal regions influencing adiposity-related traits among Samoans. Ann Hum Genet 72: 780-92.

49.

Salma N. , Xiao H. , Imbalzano AN ( 2006) Temporal recruitment of CCAAT/enhancer-binding proteins to early and late adipogenic promoters in vivo. J Mol Endocrinol 36: 139-51.

50.

Valle M. , Martos R. , Gascon F. , Canete R. , Zafra MA , Morales R. ( 2005) Low-grade systemic inflammation, hypoadiponectinemia and a high concentration of leptin are present in very young obese children, and correlate with metabolic syndrome. Diabetes Metab 31: 55-62.

51.

Friedman JM , Halaas JL (1998) Leptin and the regulation of body weight in mammals. Nature 395: 763-70.

52.

Lindsay RS , Funahashi T. , Krakoff J. , et al. (2003) Genome-wide linkage analysis of serum adiponectin in the Pima Indian population. Diabetes 52: 2419-25.

53.

Wertheim N. , Cai Z. , McGraw TE (2004) The transcription factor CCAAT/enhancer-binding protein alpha is required for the intracellular retention of GLUT4. J Biol Chem 279: 41468-76.

54.

Wu Z. , Rosen ED , Brun R. , et al. (1999) Cross-regulation of C/EBP alpha and PPAR gamma controls the transcriptional pathway of adipogenesis and insulin sensitivity . Mol Cell 3: 151-8.

55.

Christy RJ , Kaestner KH , Geiman DE , Lane MD ( 1991) CCAAT/enhancer binding protein gene promoter: binding of nuclear factors during differentiation of 3T3-L1 preadipocytes. Proc Natl Acad Sci U S A 88: 2593-7.

56.

Seki T. , Miyasu T. , Noguchi T. , et al. (2001) Reciprocal regulation of tissue-type and urokinase-type plasminogen activators in the differentiation of murine preadipocyte line 3T3-L1 and the hormonal regulation of fibrinolytic factors in the mature adipocytes. J Cell Physiol 189: 72-8.

57.

Juhan-Vague I. , Alessi MC , Mavri A. , Morange PE (2003) Plasminogen activator inhibitor-1, inflammation, obesity, insulin resistance and vascular risk. J Thromb Haemost 1: 1575-9.

58.

Olofsson LE , Orho-Melander M. , William-Olsson L. , et al. (2008) CCAAT/enhancer binding protein alpha (C/EBPalpha) in adipose tissue regulates genes in lipid and glucose metabolism and a genetic variation in C/EBPalpha is associated with serum levels of triglycerides. J Clin Endocrinol Metab 93: 4880-6.