Proteasome-mediated Turnover Research Articles

Overview of the clinical efficacy of investigational anticancer drugs.

The purpose of this overview was to make a broad inventory of investigational drugs for medicinal cancer treatment and, specifically, to indicate the evidence of clinical efficacy. Information was retrieved from electronic database searches in Medline and CANCERLIT and relevant published reviews. As the most recent findings are first reported as conference abstracts, an important basis for identification of new drugs and clinical results was a hand search of 13,392 abstracts from five major recent cancer conferences. A total of 209 investigational approaches or drugs were identified and classified into one of eight groups according to proposed mechanism of action. For 28 drugs/approaches survival data were available from randomized controlled trials. Statistically significant benefit was observed for only 12. In earlier phases no or modest anticancer activity was reported. It is speculated that the expanding knowledge in tumour biology might not easily translate into new substantially better anticancer drugs.

HRUL138, a novel human RNA-binding RING-H2 ubiquitin-protein ligase.

Cellular as well as viral RNAs are usually found complexed with proteins. In an attempt to identify proteins that interact with transcripts of hepatitis B virus (HBV), a DNA virus that replicates through reverse transcription, a partial cDNA was isolated from a human cDNA expression library whose gene product bound to an HBV-derived RNA. Using an overlapping clone from a molecular hybridization screen a full-length cDNA was assembled. It contained a large open reading frame for a 1208 amino-acid protein of 138 kDa identical to the hypothetical product of the KIAA0675 clone. Closely related sequences are present in mouse cDNA libraries but not in the genomes of lower organisms. The protein sequence contained no known RNA-binding domain and, apart from a probable coiled-coil domain, the only significant homology involved a complete RING-H2 motif. This suggested that the protein might be a novel RNA-binding RING-dependent ubiquitin-protein ligase or E3 enzyme. A motif critical for RNA binding was experimentally mapped to a central Lys-rich region. Binding specificity is either broad or the protein has as yet unknown physiological targets; hence, at present, a potential importance for HBV biology remains open. The RING-H2 domain was functional in and essential for self- and trans-ubiquitylation in vitro and for proteasome-mediated turnover of the protein in vivo. We therefore termed it hRUL138 for human RNA-binding ubiquitin ligase of 138 kDa. hRUL138 mRNAs are expressed at low levels in most tissues. GFP-tagged hRUL138 derivatives were found associated with cytoplasmic structures, possibly the ER, but excluded from the nucleus. The combined presence of RNA binding and E3 activity in hRUL138 raises the possibility that both are mechanistically linked.

Parkin Protects against the Toxicity Associated with Mutant α-Synuclein: Proteasome Dysfunction Selectively Affects Catecholaminergic Neurons

One hypothesis for the etiology of Parkinson's disease (PD) is that subsets of neurons are vulnerable to a failure in proteasome-mediated protein turnover. Here we show that overexpression of mutant α-synuclein increases sensitivity to proteasome inhibitors by decreasing proteasome function. Overexpression of parkin decreases sensitivity to proteasome inhibitors in a manner dependent on parkin's ubiquitin-protein E3 ligase activity, and antisense knockdown of parkin increases sensitivity to proteasome inhibitors. Mutant α-synuclein also causes selective toxicity to catecholaminergic neurons in primary midbrain cultures, an effect that can be mimicked by the application of proteasome inhibitors. Parkin is capable of rescuing the toxic effects of mutant α-synuclein or proteasome inhibition in these cells. Therefore, parkin and α-synuclein are linked by common effects on a pathway associated with selective cell death in catecholaminergic neurons.

C-Myc hot spot mutations in lymphomas result in inefficient ubiquitination and decreased proteasome-mediated turnover

AbstractThe c-myc proto-oncogene encodes a short-lived transcription factor that plays an important role in cell cycle regulation, differentiation and apoptosis. c-myc is often rearranged in tumors resulting in deregulated expression. In addition, mutations in the coding region of c-myc are frequently found in human lymphomas, a hot spot being the Thr58 phosphorylation site, a mutation shown to enhance the transforming capacity of c-Myc. It is, however, still unclear in what way this mutation affects c-Myc activity. Our results show that proteasome-mediated turnover of c-Myc is substantially impaired in Burkitt's lymphoma cells with mutated Thr58 or other mutations that abolish Thr58 phosphorylation, whereas endogenous or ectopically expressed wild type c-Myc proteins turn over at normal rates in these cells. Myc Thr58 mutants expressed ectopically in other cell types also exhibit reduced proteasome-mediated degradation, which correlates with a substantial decrease in their ubiquitination. These results suggest that ubiquitin/proteasome-mediated degradation of c-Myc is triggered by Thr58 phosphorylation revealing a new important level of control of c-Myc activity. Mutation of Thr58 in lymphoma thus escapes this regulation resulting in accumulation of c-Myc protein, likely as part of the tumor progression.

9 - Lymphomas and Dysproteinaemias

The channel-kinase transient receptor potential melastatin 7 (TRPM7) is a bifunctional protein with ion channel and kinase domains. The kinase activity of TRPM7 has been linked to the regulation of a broad range of cellular activities, but little is understood as to how the channel itself is regulated by its own kinase activity. Here, using several mammalian cell lines expressing WT TRPM7 or kinase-inactive variants, we discovered that compared with the cells expressing WT TRPM7, cells in which TRPM7's kinase activity was inactivated had faster degradation, elevated ubiquitination, and increased intracellular retention of the channel. Mutational analysis of TRPM7 autophosphorylation sites further revealed a role for Ser-1360 of TRPM7 as a key residue mediating both TRPM7 stability and intracellular trafficking. Additional trafficking roles were uncovered for Ser-1403 and Ser-1567, whose phosphorylation by TRPM7's kinase activity mediated the interaction of the channel with the signaling protein 14-3-3θ. In summary, our results point to a critical role for TRPM7's kinase activity in regulating proteasome-mediated turnover of the TRPM7 channel and controlling its cellular localization in polarized epithelial cells. Overall, these findings improve our understanding of the significance of TRPM7's kinase activity for functional regulation of its channel activity.