Investigating the essential roles of Ubiquitin-like molecules and the Ubiquitin Proteasome System

Who we are

Area of research

The biochemical process of ubiquitylation is an efficient way to label proteins that will be targeted to degradation by the proteasome at the right time and cellular compartment, according to the cell necessities. In addition, this post-translational modification also offers to the cell the possibility to control other processes related or not to proteolytic functions. Indeed, the attachment of ubiquitin (Ub) or ubiquitin-like (UBL) molecules such as SUMO and NEDD8 does not always result in proteolysis, but can induce conformational and/or interaction changes driving a large diversity of effects. Considering the interconnectivity among Ub and UBL molecules, we end up with an extremely complex system regulating protein activity and stability. Given the implications of the ubiquitin proteasome system (UPS) in many essential cellular processes, the technology required to study this system is very diverse. As part of the proposed network the latest technological developments will be used to explore chemical, biochemical, molecular, developmental and genetic aspects of the UPS. This research area has become an area of investigation by itself, very attractive for fundamental scientists as well as for the pharmaceutical industry aiming to identify potential targets for drug development to be used in many diseases where this system has been shown to be vital.

Objectives

The main objective of the international training network UPStream is the training of a novel generation of European scientist in a critical and complex field of modern biology: the understanding of the regulation of the ubiquitin proteasome system and its potential use for drug development, as well as ubiquitin-like molecules.

State of the art

Tagging protein substrates with Ubiquitin or Ubiquitin-like molecules is a highly dynamic and reversible process.

Ubiquitin (Ub) and ubiquitin-like molecules (UBLs) play key roles in the control of many vital cellular processes, such as cell cycle control, stress response, DNA repair, signal transduction, transcription and gene silencing [1].

The active form of ubiquitin is generated through the processing of a C-terminal hydrolase (UCH). Conjugation of Ub to a protein substrate is accomplished via a thioester cascade involving an Ub-activating enzyme (E1), an Ub-conjugating enzyme (E2) and an Ub-protein ligase (E3) (Fig 1). The result of this cascade is the formation of an isopeptide bond between the C-terminus of Ub and a lysine (Lys) in the target protein. Most of the specificity of the system derives from the high fidelity of protein-protein interactions between E3 and the substrate [1, 2]. Ub can be attached to proteins as a monomer or as polymers. PolyUb chains are formed when additional Ub molecules are attached to one of the 7 Lys residues existing in the Ub molecule (K6, K11, K27, K29, K33, K48 or K63), or in a head-to-tail manner (linear Ub chains), resulting in chain-specific functions. For example, K48 and K11 Ub chains have been associated with proteasomal degradation [1, 2].

UBLs have been reported to play a role in proteasomal degradation, SUMO-2 being shown to be able to recruit an Ub E3 ligase that can then target proteins to degradation [3, 4]. The covalent attachment of UBLs to Lys residues is biochemically similar to, but functionally distinct from, ubiquitylation. So far the best-characterised UBL systems are SUMO (SUMOylation) and NEDD8 conjugation (NEDDylation) pathways [5, 6]. Attachment of Ub, SUMO and NEDD8 molecules is a highly dynamic process, while the reverse reaction (de-ubiquitylation) is mediated by specific isopeptidases (de-conjugating enzymes), such as de-ubiquitylating enzymes (DUBs) or SUMO specific proteases (SUSPs) [7, 8] (Figure 1).

E3 ligases and DUBs, could be therapeutic targets, as they recognize a discrete subset of substrate proteins.

Figure 1. Conjugation and deconjugation of ubiquitylated proteins

The proteasome

The presentation and degradation of substrates by the proteasome is a highly regulated process. Composed of a catalytic core (20S) and one or two regulatory particle(s) (19S), the 26S proteasome recognizes and drives the degradation of ubiquitylated proteins (Figure 2).

Figure 2. Electron Microscopy 3D images of purified 26S proteasomes. By Krutauz D, Rosenzweig R, Dafonseca P, Morris E, Glickman MH, (unpublished)

The successful use of proteasome inhibitors for the treatment of Multiple Myelomas (MM) and Mantle Cell Lymphomas (MCL) revealed the potential of the UPS as a drug target. The importance of these findings was recognized with the attribution of the Nobel Prize in Chemistry to Irwin Rose, Avram Hershko and Aaron Ciechanover in 2004. Since then, a frenetic course for the development of drugs acting at different levels of the UPS has started.

Ubiquitin and ubiquitin-like binding domains

A growing number of Ub- and UBL-binding domains (UBDs) has been identified in the last decade. UBDs are found in members of many different protein families that link them to Ub-related processes. These motifs, of 20 to 40 amino acid residues, are highly conserved among various species. Some of the most studied UBDs include the CUE (coupling Ub to the endoplasmic reticulum), the UIM (Ub-interacting motif) and the UBA (Ub-associated) domains [9]. A significant characteristic of the UBDs is their ability to fold into an active UBL-interacting form independent of the rest of their protein. A single SUMO-interaction motif (SIM) has been reported to date and it has been associated with protein degradation although it might not be a general function for this motif [3, 4, 10].

References:

  • Fang S, and Weissman AM (2004) Cell Mol. Life Sci. 61: 1546-61.
  • Herpe R, and Rodriguez MS (2008) Int. J. Biochem. Cell Biol. 40: 1126-40.
  • Sun H, et al (2007) EMBO J. 26: 4102-12.
  • Prudden J, et al (2007) EMBO J. 26: 4089-101.
  • Johnson ES (2004) Annu. Rev. Biochem. 73: 355-82.
  • Pan ZQ, et al (2004) Oncogene 23: 1985-97.
  • Guterman A and Glickman MH (2004) Curr Protein Pept Sci. 5(3):201-11.
  • Melchior F, et al (2003) Trends Biochem Sci. 28: 612-8.
  • Buchberger A (2002) Trends Cell Biol. 12: 216-21
  • Perry JJ, et al (2008). Trends Biochem Sci. 33: 201-8.

Management structure

Supervisory Board

Members: All 14 Scientists in-charge including the coordinator as chair and a representative of the Fellow Board. The Supervisory Board is the highest and final decision making body of the consortium. Each institution has one vote. Prof. Ron Hay (UNIVDUN) has the casting vote in case of a tie.

The Supervisory Board (and sub-boards) meets (preferably in person, but also by videoconference if required) every six months. The Supervisory Board is supported by the Coordinator and its Project Management Team. The Project Management Team is responsible for the overall management of the project, communication between all partners, communication with the EU commission, distribution of funds, collating of annual reports and financial statements and forwarding these to the commission, decisions regarding Intellectual Property Rights, including arbitration where necessary, and the official representation of the project.

Ombudsman

The advise of the ombudsman to the Supervisor with regard to matters raised by an ESR or ER via or independently of the Fellow Board is a binding advice. The Ombudsman will be P8-Goethe: Ivan Dikic.

Scientific Executive Board - SEB

Members: P1-CICbioGUNE: Manuel S. Rodriguez; P7A-IGMM: Marc Piechazyck; P8-Goethe: Ivan Dikic; P9-UNIVDUN: Ron T. Hay.

Tasks of the SEB with regard to WP2 (RP and IRP as described in B2) and WP6:

  • Monitoring milestones and deliverables. Managing progress and interaction between the different RPs and IRPs; Monitoring worldwide state of the art and suggesting changes to the Scientist-in-charge; Monitoring ethical aspects

Training Executive Board - TEB

Members: P1-CICbioGUNE: Manuel S. Rodriguez; P4-NKI: Huib Ovaa; P7B-CRMB: Olivier Coux; P11-GSK: Julio J. Martin. The presence of GSK as private partner ensures that the training is geared towards providing the trained researchers the widest possible employment prospects.

  • Members: P1-CICbioGUNE: Manuel S. Rodriguez; P4-NKI: Huib Ovaa; P7B-CRMB: Olivier Coux; P11-GSK: Julio J. Martin. The presence of GSK as private partner ensures that the training is geared towards providing the trained researchers the widest possible employment prospects.

Recruitment Executive Board - REB

Members: P1-CICbioGUNE: Rosa Barrio; P5-UCPH: Jesper Olsen; P6-LUMC: Alfred Vertegaal; P7B-CRBM: Olivier Coux.

  • The main objective of this board is with regard to WP 1 Recruitment, the overall recruitment and secondment of researchers (both ESRs and ERs) into the UPStream work programme.

Fellow Board

The Fellow Board consists of all recruited ESRs and ERs, and advises the Training Executive Board on training needs and takes part in the organization of network meetings. The Ombudsman will be available for all Fellows as an independent arbiter and can be consulted on all confidential matters.