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The ubiquitin clan: A protein family essential for life

2011; Wiley; Volume: 585; Issue: 18 Linguagem: Inglês

10.1016/j.febslet.2011.08.020

1873-3468

Dieter H. Wolf,

Cancer-related Molecular Pathways

When in 1978 biochemical experiments uncovered a heat stable polypeptide component of an ATP-dependent proteolytic system from reticulocytes, APF-1 [1], later identified as ubiquitin [2] (a molecule previously discovered as “ubiquitous immunopoietic polypeptide” [3]), nobody in the scientific community would have imagined the impact this 76 amino acids containing molecule will have more than 30 years later in our understanding of cellular regulation. The discovery of ubiquitin function evolved from the search for, a long time enigmatic, energy-dependent degradation process of individual cellular proteins. It was uncovered that ubiquitin is covalently bound to proteins to subject them to proteolysis and that this process requires ATP [4]. An isopeptide linkage is formed between the C-terminal glycin of ubiquitin and a lysine residue of the protein [5]. In case lysine residues are missing also binding of ubiquitin to the amino terminus of a protein can occur [6]. Linking ubiquitin to a protein requires three steps: (i) the C-terminal activation of ubiquitin via an ubiquitin-adenylate and formation of an thioester, (ii) transfer of ubiquitin as thioester to a ubiquitin conjugating enzyme E2 and (iii) the final formation of the isopeptide bond of ubiquitin with the protein together with an ubiquitin ligase, E3 [7, 8]. All the early studies on the ubiquitin triggered proteolytic system had been carried out in a cell-free system. The breakthrough, showing that the ubiquitin system is indeed of tremendous physiological relevance, came from the study of a mammalian mutant cell line conditionally defective in the ubiquitin activating enzyme E1: This mutant exhibits a disturbed cell cycle and a defective degradation of short-lived intracellular proteins under restrictive conditions [9, 10]. This proved the invaluable tool of genetics to unravel the ubiquitin system in its details and initiated genetic and molecular biological experimentation on the best studied eukaryotic model organism, the yeast Saccharomyces cerevisiae [11-13]. The protease finally degrading the ubiquitin tagged proteins remained an enigma for some time. As before, in cell-free extracts a high molecular mass protein was found which was able to degrade ubiquitin tagged proteins [14, 15]. Again, yeast genetics uncovered that this proteinase, called proteasome, is actually the machine required for degradation of ubiquitinated proteins in vivo [16]. At this time the term ubiquitin-proteasome system (UPS) was coined. A multitude of proteolytic processes involving the degradation of specific proteins by the ubiquitin proteasome system was discovered soon thereafter: First reports concerned transcriptional regulation [17, 18], cell cycle progression [19, 20], metabolic regulation [21, 22] and protein quality control of the endoplasmic reticulum (ERAD) [23]. Rapidly thereafter, a multitude of proteins of different cellular pathways were shown to be selectively degraded by the UPS (for review, see [24]). Protein degradation by the UPS was shown not to depend on many single ubiquitin molecules, linked to many different lysine residues of the protein but on a polyubiquitin chain instead, which is built up on the protein target utilizing one of the seven lysine residues of ubiquitin itself, K48 [25]. Most interestingly, a tetrameric ubiquitin chain unit increases the affinity for the proteasome by a factor of about 100 and represents a unique binding determinant, that is not created by shorter chains and is also not present in mono-ubiquitin [26]. Obviously it is the surface that can be created by chain formation, which is recognized by different receptors. The changing localization of the hydrophobic surface patch with isoleucine 44 of ubiquitin through chain formation obviously determines the binding properties. Taking into account the seven lysine residues in ubiquitin (K6, K11, K27, K29, K33, K48 and K63) as well as the N-terminus of methionine 1 and the possibility of forming also mixed chains, one can imagine nature's potential to create a myriad of different surfaces which can be decoded by different receptors (proteins with ubiquitin binding domains, UBDs) all inducing different cellular responses (for review see [27]). One may imagine that evolution has a fantastic playground still here in the future. The presence of ubiquitin chain elongating enzymes (E4 ligases) [28] (for review see [29]) and many different de-ubiquitylating enzymes (DUBs) (for review see [29, 30]) might influence the dynamics of the different processes. Recently also unconventional tagging of proteins with ubiquitin utilizing serine/threonine and cysteine residues has been discovered [31]. The K48 linked ubiquitin chains targeting proteins to proteasomal destruction have been coined the “classical” or “canonical” ubiquitin chains. About 20 years after the discovery of ubiquitin triggering protein degradation, it came as a surprise when it was discovered that ubiquitin is also involved in non-proteolytic processes. Here, non-classical (non-canonical) linkage types formed by M1-, K11- or K63-linked ubiquitin chains were found. Such non-proteolytic processes include DNA repair [32], cell cycle progression, innate immunity and inflammation [27, 33]. Interestingly, however, non-canonical polyubiquitin chains can also be proteolysis signals. For instance, K11 chains generated by the anaphase promoting complex or a mixture of K11, K48 and K63 chains linked to cyclin B1 can be signals for proteasomal degradation [27, 34, 35]. In addition, also mono-ubiquitination of proteins has been shown to be an important signal: Multiple mono-ubiquitins or oligo-ubiquitin chains serve as a sorting signal on plasma membrane proteins for internalization into the endocytic pathway [36]. Mono-ubiquitination is required for sorting membrane embedded cargo proteins to the endosome and lysosome via the ESCRT pathway [37, 38]. One of the future goals must consist in a precise understanding of how the ubiquitin binding partners (receptors) selectively decode the many different ubiquitin signals presented on proteins in time and space and by this regulate the protein orchestra of the cell on the many different levels. After the many surprises the discovery of the ubiquitin system and its widespread cellular functions had provoked, the discovery of protein modifiers of the ubiquitin type came as an additional surprise. All newly discovered ubiquitin-like modifiers (UBLs) share the typical β-grasp fold of ubiquitin. Conjugation of these ubiquitin-like modifiers to proteins utilizes mechanistically similar pathways as found for ubiquitin conjugation. In order to minimize crosstalk with ubiquitin, the conjugation components, however, are distinct. Yet discovered UBLs include ISG15, the SUMO family (SUMO1, SUMO2, SUMO3; Smt3 in yeast), Urm1, NEDD8 (Rub1 in yeast), FUBI (also known as MNSF-β or FAU), FAT10, UFM1, Atg8 and Atg12. Putative UBLs are BUBL1, BUBL2, UBL-1, SF3A120 and oligoadenylate synthetase (for review, see [39, 40]). In the following some functions of the yet best studied UBLs are mentioned: ISG15 was the first UBL found [41]. The 15kD protein is strongly induced by interferons and connected to antiviral responses [42]. SUMO conjugation to target proteins controls a widespread number of cellular activities. Among them are nuclear transport [43, 44], DNA repair [32], cell cycle progression or responses to viral infections [39, 45]. Most interestingly, the SUMO system and the ubiquitin system perform crosstalks [32]. The fact that SUMO can act as a signal for ubiquitination of poly-SUMO chain modified proteins by extending the SUMO chain with an ubiquitin chain and target these proteins for proteasomal degradation was surprising [46] (for review see [47]). The creation of mixed SUMO-ubiquitin chains expands even more the cell's repertoire of signalling surfaces to specifically regulate protein activity. NEDD8 also crosstalks to the ubiquitin system. Its conjugation regulates the activity of cullin-family E3 ubiquitin ligases (for review see [48]). FAT10, originally known as diubiquitin has turned out to be an additional UBL, targeting proteins to the proteasome for degradation. Its induction by cytokines points to the fact that it may play a specific role in the immune system [49] (for review see [50]). Atg8 and Atg12 are UBLs, which provide the link to a different cellular proteolytic pathway, autophagocytosis. Atg12 is conjugated to the Atg5 protein and Atg8 is conjugated to the lipid phosphatidyl-ethanolamine of the membrane of the forming autophagosome. Both conjugation steps are essential for autophagy and finally the degradation of phagocytosed cellular material in the lysosome [51, 52]. The extraordinary mechanistic impact of ubiquitin and the many ubiquitin like proteins in cell regulation have posed the question about ancestor proteins and their evolution. Modern sequence comparison methods, structure determination and mechanistic analysis of the biosynthesis of secondary metabolites and cofactors in procaryotes have led the way. Obviously the biochemical toolbox of the key sulfur incorporation steps in the biosynthetic pathways for thiamine and molybdenum/tungsten cofactors (MoCo/Wco) in bacteria has been evolved to the eukaryotic ubiquitin family conjugation system [40, 53]. As the highly flexible but controlled conjugation of the ubiquitin family proteins to the orchestra of cellular proteins is central to most, if not all physiological processes, it is not surprising that the list of diseases connected to misregulation of the system is steadily growing. Many cancers, severe types of mental retardation, protein folding diseases leading to neurodegenerative disorders (Alzheimer disease, Parkinson disease, Huntingston disease, Creutzfeldt-Jakob disease) or metabolic diseases as type 2 diabetes make the ubiquitin family conjugation system a major target for mechanism based drug intervention [54]. It must therefore be a major goal in the future to understand the nearly universally employed ubiquitin family conjugation system comprehensively. The author thanks Elisabeth Tosta for expert help with the preparation of the manuscript. The work of the author was supported by grants of the Deutsche Forschungsgemeinschaft (Bonn, Germany) and the Fonds der Chemischen Industrie (Frankfurt, Germany). My apology goes to all the colleagues whose work could not be cited because of space limitation.