Adrenal GIPR expression and chromosome 19q13 microduplications in GIP-dependent Cushing’s syndrome
Anne-Lise Lecoq, Constantine A. Stratakis, Say Viengchareun, Ronan Chaligné, Lucie Tosca, Vianney Deméocq, Mirella Hage, Annabel Berthon, Fabio R. Faucz, Patrick Hanna, Hadrien-Gaël Boyer, Nicolas Servant, Sylvie Salenave, Gérard Tachdjian, Clovis Adam, Vanessa Benhamo, Eric Clauser, Anne Guiochon-Mantel, Jacques Young, Marc Lombès, Isabelle Bourdeau, Dominique Maiter, Antoine Tabarin, Jérôme Bertherat, Hervé Lefebvre, Wouter de Herder, Estelle Louiset, André Lacroix, Philippe Chanson, Jérôme Bouligand, Peter Kamenický
Anne-Lise Lecoq, Constantine A. Stratakis, Say Viengchareun, Ronan Chaligné, Lucie Tosca, Vianney Deméocq, Mirella Hage, Annabel Berthon, Fabio R. Faucz, Patrick Hanna, Hadrien-Gaël Boyer, Nicolas Servant, Sylvie Salenave, Gérard Tachdjian, Clovis Adam, Vanessa Benhamo, Eric Clauser, Anne Guiochon-Mantel, Jacques Young, Marc Lombès, Isabelle Bourdeau, Dominique Maiter, Antoine Tabarin, Jérôme Bertherat, Hervé Lefebvre, Wouter de Herder, Estelle Louiset, André Lacroix, Philippe Chanson, Jérôme Bouligand, Peter Kamenický
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Research Article Endocrinology

Adrenal GIPR expression and chromosome 19q13 microduplications in GIP-dependent Cushing’s syndrome

Abstract

GIP-dependent Cushing’s syndrome is caused by ectopic expression of glucose-dependent insulinotropic polypeptide receptor (GIPR) in cortisol-producing adrenal adenomas or in bilateral macronodular adrenal hyperplasias. Molecular mechanisms leading to ectopic GIPR expression in adrenal tissue are not known. Here we performed molecular analyses on adrenocortical adenomas and bilateral macronodular adrenal hyperplasias obtained from 14 patients with GIP-dependent adrenal Cushing’s syndrome and one patient with GIP-dependent aldosteronism. GIPR expression in all adenoma and hyperplasia samples occurred through transcriptional activation of a single allele of the GIPR gene. While no abnormality was detected in proximal GIPR promoter methylation, we identified somatic duplications in chromosome region 19q13.32 containing the GIPR locus in the adrenocortical lesions derived from 3 patients. In 2 adenoma samples, the duplicated 19q13.32 region was rearranged with other chromosome regions, whereas a single tissue sample with hyperplasia had a 19q duplication only. We demonstrated that juxtaposition with cis-acting regulatory sequences such as glucocorticoid response elements in the newly identified genomic environment drives abnormal expression of the translocated GIPR allele in adenoma cells. Altogether, our results provide insight into the molecular pathogenesis of GIP-dependent Cushing’s syndrome, occurring through monoallelic transcriptional activation of GIPR driven in some adrenal lesions by structural variations.

Authors

Anne-Lise Lecoq, Constantine A. Stratakis, Say Viengchareun, Ronan Chaligné, Lucie Tosca, Vianney Deméocq, Mirella Hage, Annabel Berthon, Fabio R. Faucz, Patrick Hanna, Hadrien-Gaël Boyer, Nicolas Servant, Sylvie Salenave, Gérard Tachdjian, Clovis Adam, Vanessa Benhamo, Eric Clauser, Anne Guiochon-Mantel, Jacques Young, Marc Lombès, Isabelle Bourdeau, Dominique Maiter, Antoine Tabarin, Jérôme Bertherat, Hervé Lefebvre, Wouter de Herder, Estelle Louiset, André Lacroix, Philippe Chanson, Jérôme Bouligand, Peter Kamenický

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Figure 3

Chromosome rearrangements in adrenocortical adenoma of two patients with glucose-dependent insulinotropic polypeptide–dependent (GIP-dependent) Cushing’s syndrome drive ectopic GIP receptor (GIPR) expression in adenoma cells.

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Chromosome rearrangements in adrenocortical adenoma of two patients with...
(A) Genomic rearrangements in adenoma samples from patient #1 and #2, as revealed by mate-pair analysis. Next-generation sequencing (NGS) data identified abnormally mapped reads corresponding in tumor from patient #1 to interchromosomal rearrangement between chromosome segments 19q13.32 and 20q13.12, and in tumor from patient #2 to intrachromosomal rearrangement between chromosome segments 19q13.32 and 19q13.43. The genomic coordinates of the rearranged genomic segments are shown above the boxes. White and blue arrows indicate abnormal reads corresponding to break points. Selected genes in the rearranged chromosome segments are shown below the boxes (adapted from the UCSC Genome Browser). The break point at position hg19 chr20:45,950,228 in adenoma from patient #1 corresponds to the second intron of the zinc finger and the MYND (myeloid, Nervy, and DEAF-1) domain containing 8 (ZMYND8) gene, which is consequently truncated. In the adenoma of patient #2, the break point at position hg19 chr19:57,454,276 falls within a noncoding region of chromosome 19. (B) The breakpoints in the proximity of the GIPR gene at base-pair resolution, characterized by conventional Sanger sequencing of the PCR fragments, showing one base-pair insertion (Ins) typical of canonical nonhomologous end-joining. Microhomologies are indicated in red. (C) DNA (top) and RNA FISH (bottom) analyses in adenoma of patient #1. On DNA FISH, 3 GIPR loci (green) and two ZMYND8 loci (red) are visible in each interphase nucleus. Thus, DNA FISH confirms duplication of the 19q13.32 region and its insertion in close proximity to ZMYND8 in this sample. Representative examples of GIPR expression (green) and of ZMYND8 expression (white) in adenoma of patient #1, as analyzed by RNA FISH, are also shown. Arrows indicate GIPR expression from a single allele in each nucleus. The RNA FISH signal corresponds to the translocated GIPR allele fused with intron 2 of the ZMYND8 gene located in chromosome region 20q13.12. Scale bars: 5 μm.