Brief Genetics Report No Evidence for Linkage Or For
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Brief Genetics Report No Evidence for Linkage or for Diabetes-Associated Mutations in the Activin Type 2B Receptor Gene (ACVR2B) in French Patients With Mature-Onset Diabetes of the Young or Type 2 Diabetes Sophie Dupont,1 El Habib Hani,1 Corentin Cras-Me´neur,2 Fre´de´rique De Matos,1 Ste´phane Lobbens,1 Ce´cile Lecoeur,1 Martine Vaxillaire,1 Raphae¨l Scharfmann,2 and Philippe Froguel1 Activins are members of the transforming growth fac- pancreatic buds from gut endoderm (2). Furthermore, tor- superfamily. They have a wide range of biological activins, which are expressed in rat islets and rat pancre- effects on cell growth and differentiation. For trans- atic anlage, stimulate insulin secretion even in the absence membrane signaling, activins bind directly to activin of glucose by inhibiting the activity of ATP-sensitive Kϩ receptor type 2A (ACVR2A) or 2B (ACVR2B). Trans- channels and modulating voltage-dependent Ca2ϩ chan- genic and knock-out mice for the ACVR2B gene display nels (3,4). For transmembrane signaling, activin requires various endocrine pancreas-related abnormalities, in- two types of activin receptors: type 1 (ACVR1) and type 2 cluding islet hypoplasia and glucose intolerance, dem- (ACVR2). Activin binds directly to ACVR2, and this com- onstrating the crucial role of ACVR2B in the regulation plex associates with and phophorylates ACVR1, which in of pancreas development. We have thus examined the contribution of this factor to the development of ma- turn activates the downstream signaling pathways impli- ture-onset diabetes of the young (MODY) and type 2 cating SMAD 2 and 3 (5). Two type 2 receptors (ACVR2A diabetes. No evidence of linkage at the ACVR2B locus and ACVR2B) that are expressed in the primordia of has been detected in MODY families with unknown pancreas and mature pancreatic cells in mice have been etiology for diabetes or found in affected sib pairs from identified (6). In a recent study, Kim et al. (7) described the families with type 2 diabetes. Mutation screening of the pancreatic phenotype of knockout mice for Acvr2a and/or coding sequence in MODY probands and in a family with Acvr2b. They showed that Acvr2b–/– adult mice had hypo- severe type 2 diabetes, including a case of pancreatic plastic islets and higher glycemia 30 min after an oral agenesis, showed single nucleotide polymorphisms that glucose tolerance test, whereas Acvr2b–/– had no obvious did not cosegregate with MODY and were not associated pancreatic abnormalities. Acvr2aϩ/–b–/– embryos were not with type 2 diabetes. Our results indicate that ACVR2B does not represent a common cause of either MODY or viable and showed reduced islet size and number, misex- type 2 diabetes in the French Caucasian population. pression of SHH, and reduced expression of insulin and ϩ/– –/– ϩ/– Diabetes 50:1219–1221, 2001 glucagon. Whereas Acvr2a , Acvr2a , and Acvr2b adult mice displayed no detectable malformations of the foregut-derived organs, Acvr2aϩ/–bϩ/– mice showed hypo- plastic islets and impaired glucose tolerance (resulting ctivins, members of the transforming growth from inadequate insulin production or secretion) in a more factor- superfamily, exert a diverse range of severe manner than the Acvr2b–/– mice. The endocrine biological effects on cell growth and differenti- pancreas is thus particularly sensitive to the type and A ation: they especially govern embryonic axial extent of activin receptor defective function. In addition, patterning (1) and may restrict sonic hedgehog (SHH) various abnormalities of the pancreas were reported in expression in embryonic chicks, allowing evagination of two studies of transgenic mice expressing in the pancreas a dominant-negative form of Acvr2b gene under the insulin and the -actin promoter, respectively (8,9). In both cases, From the 1Institute of Biology-Centre National de la Recherche Scientifique (CNRS) 8090, Institut Pasteur, Lille, France; and 2 Institut National de la Sante´ mice had severely hypoplastic islets. In the first study, the et de la Recherche Me´dicale (INSERM) U457, Hospital Robert Debre´, Paris, mice showed impairment of the insulin response to glu- France. cose; in the second one, abnormal endocrine cells were Address correspondence and reprint requests to Philippe Froguel, Institut Pasteur de Lille, 1 rue du Pr Calmette, 59000 Lille, France. E-mail: observed outside the islets. The functional studies impli- [email protected]. cating Acvr2b in the regulation of pancreas development Received for publication 19 September 2000 and accepted in revised form 5 February 2001. prompted us to examine the contribution of the ACVR2B Additional information can be found in an online appendix at www. gene to the development of MODY and type 2 diabetes in diabetes.org/diabetes/appendix.asp. the French population. In addition to their pancreatic ACVR1, activin receptor type 1; GI, glucose intolerant; LOD, logarithm of –/– odds; MLB, maximum-likelihood binomial; MODY, mature-onset diabetes of phenotype, Acvr2b mice have other severe complica- the young; SHH, sonic hedgehog. tions: indeed, they also show hypoplastic spleen, abnormal DIABETES, VOL. 50, MAY 2001 1219 NO T2DM-ASSOCIATED MUTATIONS IN ACVR2B IN FRENCH PATIENTS TABLE 1 TABLE 2 Genotypic distribution of the IVS6nt-13T/C variant found in the Results of type 2 diabetic sib-pair analyses performed in 143 unrelated MODY probands, type 2 diabetic subjects, and control pedigrees subjects studied Marker Phenotypic group* LOD-MLB P MLS P n T/T T/C C/C D3S1277 Large (T2DM ϩ GI) 0.02 0.37 0.11 0.32 Unrelated MODY probands 11 4 5 2 Strict (T2DM) 0.00 0.50 0.00 0.50 Unrelated T2DM probands 100 28 51 21 D3S3521 Large (T2DM ϩ GI) 0.01 0.42 0.00 0.50 Unrelated control subjects 100 35 40 25 Strict (T2DM) 0.00 0.50 0.15 0.27 Data are n. T2DM, type 2 diabetic. *The affection status was determined by taking into account the patients’ diabetic status. MLS, maximum LOD score; T2DM, type 2 stomach, defects in axial patterning, and lateral asymme- diabetes. try; 70% of these mice die before weaning, probably because of severe cardiac defects (10). Therefore, we pean and American populations by Kosaki et al. (12) searched for mutations in the ACVR2B gene in a family through the screening of the ACVR2B gene in normogly- with a severe form of type 2 diabetes treated with insulin, cemic patients with left-right axis malformations. It is including one member presenting with cardiac defects and noteworthy that we cannot exclude the presence of addi- pancreatic agenesis. The latter was not caused by muta- tional ACVR2B variations located in the promoter and tions in IPF1, which has been already screened for muta- introns in the MODY families tested. tions in this pedigree (data not shown). The markers D3S1277 and D3S3521 were also typed in We first checked the normal expression of ACVR2B in 143 French Caucasian pedigrees with type 2 diabetes (13). human embryonic pancreas by reverse transcriptase– We found no evidence of linkage between diabetes and polymerase chain reaction (details are available in the these microsatellites (Table 2) (details are available in the online appendix at www.diabetes.org/diabetes/appendix. online appendix at www.diabetes.org/diabetes/appendix. asp). Then we studied 11 MODY families of French ances- asp). The design of our linkage studies in type 2 diabetes try that were previously found to have no mutations in the pedigrees allowed us to reach the exclusion standard five known MODY genes (“MODY X” families) (11). We score of –2 for a recurrent sibling risk (s) of 1.3 (calcu- performed linkage analyses using markers located in the lated using the Mapmaker/Sib program). vicinity of the ACVR2B gene. We placed the gene between By linkage and/or screening studies, we have found no markers D3S1277 (60 cM) and D3S3521 (62 cM) by radia- evidence for a predisposing role of the ACVR2B gene in tion hybrid mapping. D3S1277 and D3S3521 were typed in MODY or type 2 diabetic French Caucasian families. 9 of the 11 French MODY X families (2 of the 11 MODY X However, studies of patients from other ethnic origins may families were not suitable for linkage analyses because be of interest to the genetics of type 2 diabetes. DNA was available for only 2 patients in those two pedi- grees). Cumulative logarithm of odds (LOD) scores showed RESEARCH DESIGN AND METHODS exclusion of linkage with diabetes at ACVR2B (LOD Subjects. The 11 MODY families are of French ancestry and have been previously described (11). The French family F4854 was recruited at Hospital scores at 0 for D3S1277 and D3S3521 were –11.74 and –17.05, respectively). The families-individual LOD scores Robert Debre´, Paris, France. All of the families and patients with type 2 diabetes came from 550 type 2 ranged from –4.71 to 0.63 for D3S1277 and from –4.28 to diabetic French Caucasian pedigrees recruited through a multimedia cam- 0.33 for D3S3521, indicating no evidence of linkage with paign. All families studied here had no known etiology for diabetes. A total of diabetes (details of the results are available in the online 143 French Caucasian pedigrees with type 2 diabetes were used for sib-pair appendix at www.diabetes.org/diabetes/appendix.asp). analyses and were primarily selected for a genome-wide search for type 2 diabetes–susceptibility genes (13). At least two diabetic subjects in each The 11 exons and flanking introns of ACVR2B were thus sibship had to be undergoing treatment for diabetes (to load the families with screened for mutations in one diabetic proband from each severe form of type 2 diabetes). The sibships presented no bilineal inheritance of the 11 MODY X pedigrees. The screening was also of type 2 diabetes, and only one of the subjects was diagnosed before the age performed in seven members of the family mentioned of 25 years.