Muscle atrophy complicates many diseases as well as aging, and its presence predicts both decreased quality of life and survival. Much work has been conducted to define the molecular mechanisms involved in maintaining protein homeostasis in muscle. To date, the ubiquitin proteasome system (UPS) has been shown to play an important role in mediating muscle wasting. In this review, we have collated the enzymes in the UPS whose roles in muscle wasting have been confirmed through loss-of-function studies. We have integrated information on their mechanisms of action to create a model of how they work together to produce muscle atrophy. These enzymes are involved in promoting myofibrillar disassembly and degradation, activation of autophagy, inhibition of myogenesis as well as in modulating the signaling pathways that control these processes. Many anabolic and catabolic signaling pathways are involved in regulating these UPS genes, but none appear to coordinately regulate a large number of these genes. A number of catabolic signaling pathways appear to instead function by inhibition of the insulin/IGF-I/protein kinase B anabolic pathway. This pathway is a critical determinant of muscle mass, since it can suppress key ubiquitin ligases and autophagy, activate protein synthesis, and promote myogenesis through its downstream mediators such as forkhead box O, mammalian target of rapamycin, and GSK3β, respectively. Although much progress has been made, a more complete inventory of the UPS genes involved in mediating muscle atrophy, their mechanisms of action, and their regulation will be useful for identifying novel therapeutic approaches to this important clinical problem.

During spermatogenesis, a large fraction of cellular proteins is degraded as the spermatids evolve to their elongated mature forms. In particular, histones must be degraded in early elongating spermatids to permit chromatin condensation. Our laboratory previously demonstrated the activation of ubiquitin conjugation during spermatogenesis. This activation is dependent on the ubiquitin-conjugating enzyme (E2) UBC4, and a testis-particular isoform, UBC4-testis, is induced when histones are degraded. Therefore, we tested whether there are UBC4-dependent ubiquitin protein ligases (E3s) that can ubiquitinate histones. Indeed, a novel enzyme, E3^Histone, which could conjugate ubiquitin to histones H1, H2A, H2B, H3, and H4 in vitro, was found. Only the UBC4/UBC5 family of E2s supported E3^Histone-dependent ubiquitination of histone H2A, and of this family, UBC4-1 and UBC4-testis are the preferred E2s. We purified this ligase activity 3,600-fold to near homogeneity. Mass spectrometry of the final material revealed the presence of a 482-kDa HECT domain-containing protein, which was previously named LASU1. Anti-LASU1 antibodies immunodepleted E3^Histone activity. Mass spectrometry and size analysis by gel filtration and glycerol gradient centrifugation suggested that E3^Histone is a monomer of LASU1. Our assays also show that this enzyme is the major UBC4-1-dependent histone-ubiquitinating E3. E3^Histone is therefore a HECT domain E3 that likely plays an important role in the chromatin condensation that occurs during spermatid maturation.

P27(Kip1) is a cyclin-dependent kinase inhibitor that regulates the G(1)/S transition. Increased degradation of p27(Kip1) is associated with cellular transformation. Previous work demonstrated that the ubiquitin ligases KPC1/KPC2 and SCF(Skp2) ubiquitinate p27(Kip1) in G(1) and early S, respectively. The regulation of these ligases remains unclear. We report here that the USP19 deubiquitinating enzyme interacts with and stabilizes KPC1, thereby modulating p27(Kip1) levels and cell proliferation. Cells depleted of USP19 by RNA interference exhibited an inhibition of cell proliferation, progressing more slowly from G(0)/G1 to S phase, and accumulated p27(Kip1). This increase in p27(Kip1) was associated with normal levels of Skp2 but reduced levels of KPC1. The overexpression of KPC1 or the use of p27(-/-) cells inhibited significantly the growth defect observed upon USP19 depletion. KPC1 was ubiquitinated in vivo and stabilized by proteasome inhibitors and by overexpression of USP19, and it also coimmunoprecipitated with USP19. Our results identify USP19 as the first deubiquitinating enzyme that regulates the stability of a cyclin-dependent kinase inhibitor and demonstrate that progression through G(1) to S phase is, like the metaphase-anaphase transition, controlled in a hierarchical, multilayered fashion.

Improvements in health in the past decades have resulted in increased numbers of the elderly in both developed and developing regions of the world. Advances in therapy have also increased the prevalence of patients with chronic and degenerative diseases. Muscle wasting, a feature of most chronic diseases, is prominent in the elderly and contributes to both morbidity and mortality. A major research goal has been to identify the proteolytic system(s) that is responsible for the degradation of proteins that occurs in muscle atrophy. Findings over the past 20 years have clearly confirmed an important role of the ubiquitin proteasome system in mediating muscle proteolysis, particularly that of myofibrillar proteins. However, recent observations have provided evidence that autophagy, calpains and caspases also contribute to the turnover of muscle proteins in catabolic states, and furthermore, that these diverse proteolytic systems interact with each other at various levels. Importantly, a number of intracellular signaling pathways such as the IGF1/AKT, myostatin/Smad, PGC1, cytokine/NFκB, and AMPK pathways are now known to interact and can regulate some of these proteolytic systems in a coordinated manner. A number of loss of function studies have identified promising therapeutic approaches to the prevention and treatment of wasting. However, additional biomarkers and other approaches to improve early identification of patients who would benefit from such treatment need to be developed. The current data suggests a network of interacting proteolytic and signaling pathways in muscle. Future studies are needed to improve understanding of the nature and control of these interactions and how they work to preserve muscle function under various states of growth and atrophy.