Visualizing Mutation-Specific Differences in the Trafficking-Deficient Phenotype of Kv11.1 Proteins Linked to Long QT Syndrome Type 2

Targeting the ubiquitin-proteasome pathway has emerged as a rational approach in the treatment of human cancer. Based on positive preclinical and clinical studies, bortezomib was subsequently approved for the clinical use as a front-line treatment for newly diagnosed multiple myeloma patients and for the treatment of relapsed/refractory multiple myeloma and mantle cell lymphoma, for which this drug has become the staple of treatment. The approval of bortezomib by the US Food and Drug Administration (FDA) represented a significant milestone as the first proteasome inhibitor to be implemented in the treatment of malignant disease. Bortezomib has shown a positive clinical benefit either alone or as a part of combination therapy to induce chemo-/radio-sensitization or overcome drug resistance. One of the major mechanisms of bortezomib associated with its anticancer activity is through upregulation of NOXA, which is a proapoptotic protein, and NOXA may interact with the anti-apoptotic proteins of Bcl-2 subfamily Bcl-X(L) and Bcl-2, and result in apoptotic cell death in malignant cells. Another important mechanism of bortezomib is through suppression of the NF-κB signaling pathway resulting in the down-regulation of its anti-apoptotic target genes. Although the majority of success achieved with bortezomib has been in hematological malignancies, its effect toward solid tumors has been less than encouraging. Additionally, the widespread clinical use of bortezomib continues to be hampered by the appearance of dose-limiting toxicities, drug-resistance and interference by some natural compounds. These findings could help guide physicians in refining the clinical use of bortezomib, and encourage basic scientists to generate next generation proteasome inhibitors that broaden the spectrum of efficacy and produce a more durable clinical response in cancer patients. Other desirable applications for the use of proteasome inhibitors include the development of inhibitors against specific E3 ligases, which act at an early step in the ubiquitin-proteasome pathway, and the discovery of less toxic and novel proteasome inhibitors from natural products and traditional medicines, which may provide more viable drug candidates for cancer chemoprevention and the treatment of cancer patients in the future.

After insertion into the endoplasmic reticulum (ER), proteins that fail to fold there are destroyed. Through a process termed dislocation such misfolded proteins arrive in the cytosol, where ubiquitination, deglycosylation and finally proteasomal proteolysis dispense with the unwanted polypeptides. The machinery involved in the extraction of misfolded proteins from the ER is poorly defined. The human cytomegalovirus-encoded glycoproteins US2 and US11 catalyse the dislocation of class I major histocompatibility complex (MHC) products, resulting in their rapid degradation. Here we show that US11 uses its transmembrane domain to recruit class I MHC products to a human homologue of yeast Der1p, a protein essential for the degradation of a subset of misfolded ER proteins. We show that this protein, Derlin-1, is essential for the degradation of class I MHC molecules catalysed by US11, but not by US2. We conclude that Derlin-1 is an important factor for the extraction of certain aberrantly folded proteins from the mammalian ER.

Using a pulse-chase approach combined with immunoprecipitation, we showed that newly synthesized influenza virus hemagglutinin (HA) and vesicular stomatitis virus G protein associate transiently during their folding with calnexin, a membrane-bound endoplasmic reticulum (ER) chaperone. Inhibitors of N-linked glycosylation (tunicamycin) and glucosidases I and II (castanospermine and 1-deoxynojirimycin) prevented the association, whereas inhibitors of ER alpha-mannosidases did not. Our results indicated that binding of these viral glycoproteins to calnexin correlated closely with the composition of their N-linked oligosaccharide side chains. Proteins with monoglucosylated oligosaccharides were the most likely binding species. On the basis of our data and existing information concerning the role of monoglucosylated oligosaccharides on glycoproteins, we propose that the ER contains a unique folding and quality control machinery in which calnexin acts as a chaperone that binds proteins with partially glucose-trimmed carbohydrate side chains. In this model glucosidases I and II serve as signal modifiers and UDP-glucose:glycoprotein glucosyltransferase, as a folding sensor.