Publications

BACKGROUND

We looked for mutations of the HRPT2 gene, which encodes the parafibromin protein, in sporadic parathyroid carcinoma because germ-line inactivating HRPT2 mutations have been found in a type of familial hyperparathyroidism--hyperparathyroidism-jaw tumor (HPT-JT) syndrome--that carries an increased risk of parathyroid cancer.

METHODS

We directly sequenced the full coding and flanking splice-junctional regions of the HRPT2 gene in 21 parathyroid carcinomas from 15 patients who had no known family history of primary hyperparathyroidism or the HPT-JT syndrome at presentation. We also sought to confirm the somatic nature of the identified mutations and tested the carcinomas for tumor-specific loss of heterozygosity at HRPT2.

RESULTS

Parathyroid carcinomas from 10 of the 15 patients had HRPT2 mutations, all of which were predicted to inactivate the encoded parafibromin protein. Two distinct HRPT2 mutations were found in tumors from five patients, and biallelic inactivation as a result of a mutation and loss of heterozygosity was found in one tumor. At least one HRPT2 mutation was demonstrably somatic in carcinomas from six patients. Unexpectedly, HRPT2 mutations in the parathyroid carcinomas of three patients were identified as germ-line mutations.

CONCLUSIONS

Sporadic parathyroid carcinomas frequently have HRPT2 mutations that are likely to be of pathogenetic importance. Certain patients with apparently sporadic parathyroid carcinoma carry germ-line mutations in HRPT2 and may have the HPT-JT syndrome or a phenotypic variant.

Tumor necrosis factor-alpha (TNF-alpha) has been shown to reduce endothelial nitric-oxide synthase (eNOS) gene expression through post-transcriptional regulation of mRNA stability. The current study documented an independent effect of the cytokine on the eNOS gene promoter. TNF-alpha effected a time- and dose-dependent reduction in activity of a transiently transfected human -1197 eNOS-luciferase reporter. This reduction was inhibited by co-transfection of dominant negative IKKbeta as well as a nonphosphorylatable constitutively suppressive mutant of IkappaB implying involvement of the NFkappaB cascade in the inhibitory effect. The locus of the TNF-alpha-dependent inhibition was traced to two Sp1-binding sites positioned between -109 and -95 and -81 and -67 relative to the transcription start site. Electrophoretic mobility shift analysis and immunoperturbation studies showed evidence for Sp1 and Sp3 binding to each element. TNF-alpha treatment had no effect on the binding pattern to the downstream (-81 to -67) site but did suppress association of Sp1 and Sp3 to the upstream (-109 to -95) site. Collectively, these data indicate that TNF-alpha exerts transcriptional, as well as post-transcriptional, effects on eNOS gene expression and suggest a potential mechanism to account for the endothelial dysfunction that accompanies disorders such as diabetes mellitus and heart failure.

Microarray experiments have identified novel candidate genes in animal models of asthma. In the near future, genomics may have a profound impact on the way we think about this common and complex disease.