528 The ATP-dependent, ubiquitin–proteasome system (UPS) Over the past two decades, progress in understanding the action and regulation of the UPS has been at the center of attempts to understand the control of protein turnover. The UPS includes concerted actions of enzymes that link ubiquitin (Ub), a member of the heat-shock protein family, to protein substrates that are destined for degradation (Fig. 1). When a chain of at least four to five ubiquitins are linked to a protein, it is marked for degradation in a second step mediated by the proteasome [12, 13]. Specif- ically, the tagged protein will be recognized by the 26S proteasome, a very large multicatalytic protease complex that not only recognizes Ub-conjugated proteins but also removes Ub, unwinds the protein, and injects it into the central core of the 26S proteasome. Once inside this central, tube-like structure, the protein substrate is degraded into small peptides [14]. Three enzymatic components are required to link Ub to proteins that are destined for degradation. There appears to be only one E1 (Ub-activating) enzyme and around 40 E2 (Ub-carrier or conjugating) proteins. The key enzyme, however, is the E3 enzyme, which accounts for the exquisite specificity of proteins to be degraded. There are at least a thousand E3 enzymes (Ub ligase), and each can recognize a specific protein substrate and catalyze the transfer of an activated Ub from the E2 carrier protein to the substrate protein [10, 15]. Fig. 1 The ubiquitin–proteasome pathway of protein degradation. Ubiquitin (Ub) is conjugated to proteins destined for degradation by an ATP-dependent process that involves three enzymes (E1–E3). A chain of five Ub molecules attached to the protein substrate is recognized by the 26S proteasome, which removes Ub and digests the protein into peptides. The peptides are degraded to amino acids by peptidases in the cytoplasm or used in antigen presentation. (Reproduced with permission from [15]) Pediatr Nephrol (2008) 23:527–535 Since the initial reports that the UPS recognizes specific proteins and tags them for destruction, knowledge about proteolytic processes in the proteasome has exploded. Thousands of proteins have been recognized as being degraded by the UPS, and novel cellular functions are now known to be regulated by Ub conjugation. In terms of protein breakdown, the major functions of the pathway are: Rapid removal of proteins Protein degradation is irrevers- ible, and hence, destruction of a protein generally leads to a complete termination of cellular process mediated by the protein. Consequently, protein degradation is critical for the regulation of metabolism and cell turnover. The rapid degradation of specific proteins also permits cells (as well as the organism) to rapidly adapt to a change in physiological conditions (e.g. requiring a switch to glucose as an energy source involves converting protein stores into amino acids that can be used in gluconeogenesis). Regulation of gene transcription Ub conjugation affects gene transcription because many transcription factors become conjugated to Ub, and transcription activators are degraded by the proteasome [16]. This process regulates transcriptional activity by removing “spent” activators and resetting a promoter for additional rounds of transcription [17]. Second, the ability of transcription factors to function varies with their location within the cell. For example, nuclear factor (NF)-κB, a proinflammatory transcriptional factor, is kept outside the nucleus because movement into the nucleus is blocked by its association with an inhibitory protein, IκB. Destruction of IκB, initiated by the IKK kinase and carried out by the UPS, frees NF-κB, which then translocates to the nucleus to stimulate gene transcription [18]. A clinical application of this function of the UPS has developed in oncology. Bortezomib (Velcade, PS-341), a proteasome inhibitor, has proven to be beneficial in patients with multiple myeloma and is currently in clinical trials for the treatment of other cancers [19, 20]. The proposed mechanism of action involves the ability of bortezomib to prevent the UPS-induced destruction of IκB, thereby blocking the activation of NF-κB (an antiapoptosis tran- scription factor), leading to an increase in apoptosis [21]. Inhibition of the proteasome, therefore, will induce apopto- sis of the neoplastic cells [22]. In addition, myeloma cells are also particularly dependent upon NF-κB to produce essential growth factors [especially inerleukin 6 (Il-6)]; when NF-κB is inactive, the growth of myeloma cells is depressed. Quality-control mechanism The UPS selectively eliminates abnormally folded or damaged proteins that have arisen because of missense or nonsense mutations, biosynthetic errors, proteins damaged by oxygen radicals, or by