BSCB Newsletter, Winter 2006

FASEB Summer Research Conference: Ubiquitin & Cellular Regulation
22–27 July 2006 Vermont Academy, Vermont

The Ubiquitin & Cellular Regulation Meeting was organised by Linda Hicke (Northwestern University) and Allan Weissman (NIH/NCI). The meeting was held at the Vermont academy and brought together approximately 200 researchers working on the Ubiquitin-Proteasome System (UPS). It was more like a summer school, including all meals and lodging, this allowed plenty of time for meeting new people, exchanging views and the chance to the seek the insight of other researchers.

The meeting provided the perfect platform for me to learn about the latest research in UPS, and how it is used to control many of the fundamental processes in eukaryotic cells. The UPS is currently one of the hottest topics in biological research with the 2004 Nobel Prize in Chemistry being awarded to Aaron Ciechanover, Avram Hersko and Irwin Rose for the discovery of ubiquitin-proteasome mediated protein degradation. I am very grateful to the British Society of Cell Biology for funding my attendance at this great conference and would like to recommend this meeting, which is held every two years.

The programme of the meeting was designed that only one session ran at a time, ensuring the opportunity to hear every talk. In total during the five days, there were nine oral presentation sessions (five or six invited speakers per session) covering different topics, one workshop, and three poster sessions. I presented my poster entitled 'Proteomic Approach to Study Inclusions Seeded by the Overexpression of a-synuclein' in session 1 – Proteasomes and Degradation. It gave me the opportunity to present my current research, discuss with researchers working on similar projects and look around related posters.

The opening night keynote lecture was entitled 'Physiological regulation by proteolysis: the N-end rule pathway and its functions'. It was given by Alexander Varshavsky (California Institute of Technology). His group was the first to discover that the N-terminal amino acid residue of a protein determines its in vivo half-life; a process now termed the N-end rule. He has since dedicated his scientific career to elucidate the mechanism underlying this process, and described new advances in the understanding of the N-end rule pathway in his talk. One topic he discussed was the N-end rule as a new kind of nitric oxide / oxygen sensor. The in vivo oxidation of N-terminal cysteine is essential for its arginylation and is shown to be controlled by nitric oxide and oxygen. UBR1 and UBR2 are the ubiquitin-protein ligases that are used for the N-end rule regulated degradation. Mutations of UBR1 in humans cause Johanson-Blizzard syndrome (JBS). Knockout mice indicate that the functions of UBR1 and UBR2 are significantly different. UBR1–/– mice are viable with pancreatic insufficiency, similarly to JBS. By contrast, UBR2–/– mice are inviable; defects in male meiosis are observed. UBR1–/–UBR2–/– showed impaired neurogenesis and cardiovascular development in mice.

The main programme of the meeting began with a memorial session dedicated to Professor Cecile Pickart one of the pioneers of ubiquitin research who sadly died recently. The focus of her laboratory at John Hopkins University has been to investigate the assembly and recognition of polyubiquitin signals, focusing on proteasome proteolysis and DNA-damage tolerance. Six speakers in this session were either from the Pickart laboratory or her collaborators. Eric Cooper (Johns Hopkins) described a complex with deubiquitinating activity that is highly specific for K63-linked ubiquitin chains. It includes the lid of the proteasome, the COP9 signalosome, and a novel complex, C6.1A, which includes a poorly characterized JAMM/MPN domain. Yien Che Tsai (NCI) showed that the RING finger protein gp78, autocrine motility factor receptor, is critical for ER-associated degradation, and promotes tumour cell invasion in vitro by its pro-metastatic ubiquitin-protein ligase (E3) activity. Zhijian 'James' Chen (University of Texas) discussed ubiquitin-mediated activation of protein kinases in the NF-kB pathway before leaving to also present the following day at the 'Ubiquitin and signalling' session of Bioscience 2006 in Glasgow, UK.

Then, there was a workshop about Drug Discovery in the Ubiquitin-Proteasome Pathway. David Glass (Novartis Institute for Biomedical Research) described signalling pathways that regulate protein ubiquitylation in skeletal muscle atrophy. Two E3s, MuRF1 and MAFbx, are transcriptionally upregulated. The pathways of regulating these E3s and their potential substrate were discussed. Teresa Soucy (Millennium Pharmaceuticals) described their work to inhibit the activity of NAE (Nedd8 activating enzyme) as an oncology target. The workshop was followed by a panel discussion 'Scientific and Career Opportunities in targeting the Ubiquitin System'. This brainstorming session gave me a few ideas regarding the relative merits of potential careers in academia or industry.

The focus of the second session was ubiquitin and ubiquitin-like (Ubl) binding proteins and their receptors. Ub-binding domains (UBDs) have been identified in many proteins that interact with mono-Ub and/or poly-Ub chains. Ivan Dikic (University of Frankfurt) initially described two UBDs referred to UBM and UBZ. However, during the session it became clear that there were additional UBDs discovered only recently. One of these was the reversed UIM domain (called MIU) described by Simona Polo (IFOM). Colin Gordon (MRC, Edinburgh) described a UBD in Mud1, which recognizes K48-linked multi-Ub chains.

In the third session, the SUMO and cullin neddylation modification pathways were described by Chris Lima (Sloan-Kettering Institute), and Ning Wei (Yale University) and Brenda Schulman (St Jude Children's Research Hospital), respectively. Jon Huibregtse (University of Texas) employed a proteomic approach to identify target proteins for ISG15 (interferon-inducible ubiquitin-like protein) modification. These included ISG15 E1 (Ube1L), ISG15 E2 (UbcH8) and HECT E3 (Herc5). As our group has previously looked at UbcH8 with respect to UPS mediated protein degradation, I found it particularly interesting that this protein has been found to have a role in ubiquitin-like protein modifications too. Also another thought for identification the targets of ubiquitin-like protein, it can be carried out by a comprehensive UPS microarray as presented in the poster session by Hartmut Scheel (Miltenyi Biotec).

The role of Ub in endocytosis and autophagy was discussed in the fourth session. Mono-Ub is required for the internalisation step of endocytosis, and Ub conjugation is required for autophagy. Annie Angers (University of Montreal) described how the E3 Itch was required to ubquitynate its substrate, the endocytic protein, Endophilin. Yoshinori Ohsumi (National Institute for Basic Biology) described two ubiquitin-like conjugation systems, Apg12 and Apg8, which regulate autophagosome formation in yeast.

The role of UPS in transcriptional regulation was discussed in session five, beginning with an overview by William Tansey (Cold Spring Harbor). Subsequently, the role of histone ubiquitylation and sumoylation in yeast transcriptional regulation was presented by Shelley Berger (Wistar institute). Helle Ulrich (CRUK) and Stefan Jentsch (Max Planck Institute of Biochemistry) discussed how the control of DNA damage tolerance mediated by PCNA (proliferating cell nuclear antigen) was dependent on ubiquitylation and SUMOylation.

The focus of the meeting then switched to the proteasome and deubiquitylating enzymes (DUBs). Tingting Yao (Stowers institute) began by describing his work, which indicates that the DUB, Uch37, is a component of the 19S regulatory complex in the cytoplasm, and an ATP-dependent chromatin remodelling complex in the nucleus. Together with Ubp6 (yeast) / Usp14 (human) and Rpn11 (yeast) there are three DUBs now known to associate with the proteasome. Daniel Finley (Harvard Medical School) discussed the regulation of proteasome activity by the E3, Hul5, and the DUB, Ubp6. Ubp6 has a conserved capacity to inhibit the proteasome while Hul5 serves to extend the chains of proteasome-bound ubiquitin conjugates. Therefore, the balance of Hul5 and Ubp6 activity might regulate substrate commitment to degradation.

In session 7, the ways in which substrate ubiquitylation can also be used as a signal to regulate other diverse cellular processes were described. Donald Kirkpatrick (Harvard Medical School) has developed a novel mass spectrometry based method, termed Ubiquitin-AQUA (Absolute QUAntification) to quantify the different linkage types of a given substrate (cyclin B1) ubiquitylated by APC (anaphase promoting complex). It was interesting to hear that the primary Ub linkage type mediated by the APC is the K63-type. In addition, Raymond Deshaies (California Institute of Technology) described another quantitative mass spectrometry method, MudPIT (multidimensional protein identification technology) to identify Ub substrates under different conditions, which permits differential ubiquitylation screens.

Because Parkinson's disease is the focus of my research, the eighth session was of real interest to me and was entitled 'Ubiquitin/Ubl physiology and Disease'. Problems with the ubiquitin system have been associated with many diseases including cancer, autoimmune disease, and neurodegenerative disorders. Kwon-Yul Ryu (Standford University) created UbB (ubiquitin B) knockout mice to show polyubiquitin B gene is required for energy homeostasis. Homozygous disruption of the UbB gene in mice has no effect on embryonic development, but the mice are obese. UbB mutants have increased adiposity, hyperleptinemia, reflex hyperphagia defective and reduced caloric efficiency. Therefore, ubiquitin functions not only cell signalling but also energy homeostasis. Carole LaBonne (Northwestern University) showed that the control of neural crest regulatory factor function requires both ubiquitynation and sumoylation. The neural crest is a population of precursor cells found only in vertebrate embryos, which share phenotypic and molecular characteristic with invasive tumour cells. She discussed how ubiquitin and SUMO plays a role in the processes of neural crest development.

In the final session (session 9), the way in which protein ubiquitylation is used by the cell to eliminate aberrant proteins, 'protein quality control (PQC) degradation', was described. Rich Gardner (Fred Hutchinson Cancer Research Center) recently identified the first PQC degradation pathway in the nucleus, where little if any protein synthesis occurs. Nuclear function requires protein action, but any aberrant proteins could cause damage to cell, therefore, the cell should be able to remove aberrant proteins from the nucleus. Key to PQC degradation in the nucleus is San1 (yeast), a RING-finger E3. ER-quality control system 'ERAD' was discussed by Christian Hirsch (Max-Belbruck Center) and Tom Rapoport (Harvard Medical School).

Overall, I felt that the conference was a brilliant experience. During the conference, my discussions with many of the participants helped me to understand more about the broader aspects of the UPS and provided me with practical hints for my own work.

Chao-Chun Hung
University of Leeds.