Albright hereditary osteodystrophy (AHO) is a congenital disorder resulting from autosomal dominant inheritance of heterozygous mutations in the GNAS gene which disrupt expression of the heterotrimeric G protein Gs-alpha. Gs-alpha is a ubiquitously expressed signaling molecule that is required for the intracellular generation of the second messenger molecule cyclic AMP in response to stimulation of hormone and other cell-surface receptors. Features of AHO include short stature, subcutaneous ossifications, brachydactyly, and neurobehavioral and developmental abnormalities. Patients who inherit the disease from their mother also develop resistance to multiple hormones which activate Gs-alpha signaling pathways (eg. parathyroid hormone PTH, thyroid stimulating hormone, gonadotropins, and growth hormone releasing hormone) as well as obesity, and this form of the disease is also known as pseudohypoparathyroidism type 1A (PHP1A). In contrast patients who inherit the disease from their father only develop features of AHO, and this is also referred to as pseudopseudohypoparathyroidism (PPHP). We and others have shown in mice and humans that Gs-alpha is imprinted in a tissue-specific manner. In some hormone target tissues Gs-alpha is expressed primarily from the maternal allele, and therefore maternal inheritance of Gs-alpha mutations results in hormone resistance while paternal inheritance of these same mutations does not. A few patients with similar mutations develop a severe form of ectopic ossification called progressive osseous heteroplasia (POH) in which the ossifications form cast-like plates and invade deeper soft tissues. While it has reported that POH only occurs when the mutation is inherited paternally, we have recently studied two cases in which POH resulted from maternal transmission. We have generated mice with Gs-alpha deficiency in chondrocytes or osteoblasts, which provides evidence that the Gs-alpha deficiency in growth plate chondrocytes is likely the direct cause of the short stature and brachydactyly observed in AHO. ? ? GNAS (Gnas in mice) is a very complicated imprinted gene with multiple gene products generated by several alternative promoters and first exons. NESP55 is a chromogranin-like protein that is maternally expressed while XL-alpha-s is a paternally expressed Gs-alpha isoform with a long amino-terminal extension. Both are primarily expressed in neuroendocrine tissues. We have shown that NESP imprinting is not established until postimplantation development. We identified another alternative first exon (exon 1A) that generates paternally expressed untranslated mRNAs and that is a maternal germline imprint mark. We have shown that this region has allele-specific differences in DNA and histone methylation. We also have shown that the Gs-alpha promoter and first exon also has allele-specific differences in histone methylation which correlates to its tissue-specific imprinting, even though this region does not undergo DNA methylation. We have shown that PHPIB (parathyroid hormone resistance in the absence of AHO) is virtually always associated with loss of maternal exon 1A imprinting. A detailed analysis of GNAS imprinting in PHP IB patients showed that familial cases tend to only have abnormal exon 1A imprinting associated with a deletion mutation within a closely-linked gene, while sporadic cases often have additional imprinting defects involving NESP and XL-alpha-s. We have generated exon 1A knockout mice, and show that this region is not required for Nesp and XL-alpha-s imprinting, but is required for tissue-specific Gs-alpha imprinting. Mice with paternal exon 1A deletion, which have Gs-alpha overexpression in renal proximal tubules due to loss of paternal Gs-alpha imprinting, have increased parathyroid hormone sensitivity with low circulating parathyroid hormone levels. ? ? Although obesity has been previously considered to be a general feature of AHO present in both PHP1A and PPHP patients, we have recently shown that obesity is a specific feature of PHP1A and therefore is a result of loss of Gs-alpha expression in one or more tissues due to the combined effects of maternal mutation and paternal imprinting. These clinical observations are consistent with findings in mice with germline Gs-alpha mutations showing that mice with maternal mutations develop severe obesity with lower energy expenditure and insulin resistance and that these effects are reversed by the presence of a paternal 1A deletion which reverses Gs-alpha imprinting (Z01-DK043313-03). Preliminary results in a brain-specific Gs-alpha knockout model suggest that this imprinting effect is localized to one or more regions in the central nervous system (Z01-DK043315-01). Based upon these observations we are initiating studies in the NIH Obesity/Clinical Phenotyping Center examining the metabolic characteristics of AHO and related patients in detail to better characterize the metabolic defect and gain understanding of its pathogenesis.
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