Deubiquitylating enzymes and disease.

Deubiquitylating enzymes (DUBs) can hydrolyze a peptide, amide, ester or thiolester bond at the C-terminus of UBIQ (ubiquitin), including the post-translationally formed branched peptide bonds in mono- or multi-ubiquitylated conjugates. DUBs thus have the potential to regulate any UBIQ-mediated cellular process, the two best characterized being proteolysis and protein trafficking. Mammals contain some 80-90 DUBs in five different subfamilies, only a handful of which have been characterized with respect to the proteins that they interact with and deubiquitylate. Several other DUBs have been implicated in various disease processes in which they are changed by mutation, have altered expression levels, and/or form part of regulatory complexes. Specific examples of DUB involvement in various diseases are presented. While no specific drugs targeting DUBs have yet been described, sufficient functional and structural information has accumulated in some cases to allow their rapid development. PUBLICATION HISTORY : Republished from Current BioData's Targeted Proteins database (TPdb; http://www.targetedproteinsdb.com).

Von Hippel-Lindau-coupled and transcription-coupled nucleotide excision repair-dependent degradation of RNA polymerase II in response to trabectedin.

PURPOSE: Ecteinascidin 743 (Et743; trabectedin, Yondelis) has recently been approved in Europe for the treatment of soft tissue sarcomas and is undergoing clinical trials for other solid tumors. Et743 selectively targets cells proficient for TC-NER, which sets it apart from other DNA alkylating agents. In the present study, we examined the effects of Et743 on RNA Pol II. EXPERIMENTAL DESIGN AND RESULTS: We report that Et743 induces the rapid and massive degradation of transcribing Pol II in various cancer cell lines and normal fibroblasts. Pol II degradation was abrogated by the proteasome inhibitor MG132 and was dependent on TC-NER. Cockayne syndrome (CS) cells and xeroderma pigmentosum (XP) cells (XPD, XPA, XPG, and XPF) were defective in Pol II degradation, whereas XPC cells whose defect is limited to global genome NER in nontranscribing regions were proficient for Pol II degradation. Complementation of the CSB and XPD cells restored Pol II degradation. We also show that cells defective for the VHL complex were defective in Pol II degradation and that complementation of those cells restores Pol II degradation. Moreover, VHL deficiency rendered cells resistant to Et743-induced cell death, a similar effect to that of TC-NER deficiency. CONCLUSION: These results suggest that both TC-NER-induced and VHL-mediated Pol II degradation play a role in cell killing by Et743.

Transcriptional regulation of phenylethanolamine N-methyltransferase in pheochromocytomas from patients with von Hippel-Lindau syndrome and multiple endocrine neoplasia type 2.

Pheochromocytomas in multiple endocrine neoplasia type 2 (MEN-2) express phenylethanolamine N-methyltransferase (PNMT), the enzyme that catalyzes conversion of norepinephrine to epinephrine, whereas those in von Hippel-Lindau (VHL) syndrome do not. Consequently, pheochromocytomas in MEN-2 produce epinephrine, whereas those in VHL syndrome produce mainly norepinephrine. This study examined whether transcription factors known to regulate expression of PNMT explain the different tumor phenotypes in these syndromes. Quantitative polymerase chain reaction (PCR) and Western blotting were used to assess levels of mRNA and protein for the glucocorticoid receptor, early growth response 1 (Egr-1), the Sp1 transcription factor (Sp1), and MYC-associated zinc finger protein (MAZ) in 6 MEN-2 and 13 VHL tumors. Results were cross-checked with data obtained using microarray gene expression profiling in a further set of 10 MEN-2 and 12 VHL tumors. Pheochromocytomas in MEN-2 and VHL syndrome did not differ in expression of the glucocorticoid receptor, Egr-1, Sp1, or MAZ as assessed by quantitative PCR and Western blotting. Microarray data also indicated no relevant differences in expression of the glucocorticoid receptor, Egr-1, MAZ, and the AP2 transcription factor. Thus, our results do not support a role for the above transcription factors in determining differences in expression of PNMT in pheochromocytomas from patients with VHL syndrome and MEN-2. Microarray analysis, however, did indicate differences in expression of genes involved in neural crest cell lineage and chromaffin cell development, consistent with differential survival of PNMT-expressing cells in the two syndromes.

Human disorders of ubiquitination and proteasomal degradation.

PURPOSE OF REVIEW: The goal of this review is to provide an overview of rapidly evolving information on a new group of genetic inborn errors affecting ubiquitination and proteasomal degradation of proteins and to suggest a classification scheme for these disorders. The relevant genes encode ubiquitin, ubiquitin enzymes (E1 and many E2s and E3s), deubiquitinating enzymes, proteasomal subunits, and substrates undergoing ubiquitination. RECENT FINDINGS: Since the initial recognition that Angelman syndrome is caused by maternal deficiency of the E6-AP ubiquitin E3 ligase (gene symbol UBE3A), several. other disorders of E3 ligases have been identified, including autosomal recessive juvenile Parkinson disease, the APECED form of autoimmune polyendocrinopathy syndrome, von Hippel-Lindau syndrome, and congenital polycythemia. Disorders that disturb ubiquitin regulatory signaling include at least two subtypes of Fanconi anemia, the BRCA1 and BRCA2 forms of breast and ovarian cancer susceptibility, incontinentia pigmenti, and cylindromatosis. Many disorders affect ubiquitin pathways secondarily. SUMMARY: The authors propose both a genetic and a functional classification for disorders of ubiquitination and proteasomal degradation, as follows. Genetic classes include mutations in (1) the UBB ubiquitin gene; (2) enzymes of ubiquitination including E1, E2, E3, and related proteins; (3) deubiquitinases; (4) proteasomal subunits; and (5) substrates of ubiquitination. Functional classes include defects in (1) proteolytic degradation, (2) ubiquitin signaling, and (3) subcellular localization of substrates. Additional functional classes are likely to be defined, and individual disorders may involve multiple functional defects.

A variety of phenotype with R161Q germline mutation of the von Hippel-Lindau tumor suppressor gene in Japanese kindred.

Von Hippel-Lindau (VHL) syndrome is an autosomal dominant neoplastic disorder characterized by hemangioblastomas of the central nervous system and retina, renal cell carcinomas, pheochromocytoma, and islet cell tumors. This syndrome is closely related with the VHL, a tumor suppressor gene, implying that loss of function or inactivating mutations of both alleles or copies of this gene cause tumor formation. The product of the VHL gene, pVHL, is known to be a component of ubiquitin ligase which targets the transcription factor such as hypoxia-inducible factor (HIF) for degradation in the presence of oxygen. Different VHL mutations confer different site-specific risks of cancer. However, the precise role of pVHL to develop only some specified tumors, especially pheochromocytoma, is not fully understood. We identified a missense mutation of VHL gene, 695 G --> A (R161Q), in a Japanese kindred with type 2A VHL syndrome. We analysed 16 members of this family and detected the same mutation in 8 individuals. All 5 members with tumors possessed the same mutation. Interestingly, one of the identical twins, who had the same R161Q germline mutation, did not show any visible tumors throughout the body, while another of the twins developed a huge pheochromocytoma and retinal angioma. Moreover, one of the affected members in the kindred developed pancreatic neuro-endocrine tumors without pheochromocytoma in spite of possessing the identical germline mutation of the VHL gene. These findings suggest that there are some other additional factors including environmental exposures to initiate and develop tumor formation in the VHL syndrome although abnormalities of VHL gene might be involved.

Tyrosine hydroxylase gene expression in varying forms of human pheochromocytoma.

We performed a comparative study of catecholamine content, tyrosine hydroxylase (TH) activity, and TH mRNA levels in normal human adrenals and various clinical forms of human pheochromocytoma. We studied sporadic, benign intra-adrenal chromaffin tumors and other non-malignant intra-adrenal tumors associated with multiple endocrine neoplasia type 2B (MEN 2B) and von Hippel-Lindau disease along with one extra-adrenal malignant pheochromocytoma. Our findings suggest substantial differences in TH transcriptional rates or the stability of TH mRNA or both may contribute to altered TH expression in human chromaffin cells associated with "normal" adrenal tissues and various forms of pheochromocytoma and distinctive patterns of expression in the different settings in which these tumors arise.

Malignant brain tumours associated with adenylate kinase in cerebrospinal fluid.

Adenylate-kinase activity was measured in cerebrospinal fluid (C.S.F.) from 35 healthy control subjects and 11 patients with brain tumours, of which 9 were malignant and 2 benign. No adenylate kinase could be detected in C.S.F. from the controls or from the patients with benign brain tumours. The enzyme was consistently found in C.S.F. from patients with malignant brain tumours.