Endocrine Abstracts (2004) 7 P45

Transcriptional interference by novel human PPARgamma mutants associated with lipodystrophic insulin resistance

M Agostini1, E Schoenmakers1, AG Smith1, I Szatmari3, O Rajanayagam1, DB Savage1,2, C Mitchell1, M Clarke4, AM Zalin4, RC Trembath5, S Kumar6, JWR Schwabe7, L Nagy3, S O'Rahilly1,2, M Gurnell1 & VKK Chatterjee1

1Department of Medicine, University of Cambridge, Cambridge, UK; 2Department of Clinical Biochemistry, University of Cambridge, Cambridge, UK; 3Department of Biochemistry and Molecular Biology, University of Debrecen, Hungary; 4Wordsley Hospital, Wordsley, UK; 5Department of Medicine, University of Leicester, Leicester, UK; 6Department of Medicine, University of Warwick, Warwick, UK; 7MRC Laboratory of Molecular Biology, Cambridge, UK;

The nuclear receptor PPARgamma is important for biological processes including adipogenesis and glucose homeostasis. In subjects with severe insulin resistance, we have previously reported two types of human PPARgamma gene defect: heterozygous, missense mutations (P467L, V290M) in the ligand binding domain (LBD) which inhibit wild type (WT) receptor action in a dominant negative manner by recruitment of transcriptional corepressors; or double heterozygosity for a frameshift/premature stop (FSX mutation in the DNA binding domain (DBD) of PPARgamma leading to loss of receptor function, together with a defect in a second gene (PPP1R3) involved in skeletal muscle glycogen metabolism.

Here, we describe three novel heterozygous mutations in the DNA (C114R, C131R) or ligand (R357X) binding domains of PPARgamma in unrelated subjects with the characteristic clinical phenotype including a stereotyped pattern of partial lipodystrophy, insulin resistance, dyslipidaemia and hypertension. Consonant with their location, the novel PPARgamma mutations disrupt DNA binding (C114R, C131Y) or dimerisation (R357X) with retinoid X receptor and are transcriptionally inactive like the FSX mutation. However, in contrast to the FSX mutant, the C114R, C131Y and R357X mutant proteins retain the ability to translocate to the nucleus. Furthermore, the C114R, C131Y and R357X mutants inhibit WT receptor action when coexpressed, whereas the FSX mutant lacks dominant negative activity. Induction of a PPARgamma target gene (aP2) in response to receptor agonist was markedly attenuated in primary monocyte-derived dendritic cells from subjects harbouring the C114R, C131Y and R357X mutations whereas cells with the FSX mutation responded fully, providing in vivo evidence for the differing dominant negative properties of mutant receptors. Our observations suggest that dominant negative inhibition by the new PPARgamma mutants occurs via a novel mechanism, with competition for coactivators interfering with transcriptional activation by wild type receptor.