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Eukaryotic cells are operational with three different systems to accomplish protein degradation: mitochondrial proteases, which are responsible for the majority of mitochondrial proteins degradation; lysosomes, which mostly degrade membrane and endocytosed protein and ubiquitin-proteasome system (UPS), which degrades a variety of normal and abnormal intracellular proteins. In fact, up to 80–90% of all intracellular proteins are degraded via UPS and it is considered as one of the major pathways of intracellular protein degradation. Protein modification by ubiquitin (Ub) or ubiquitin like modifiers (Ubls) like SUMO, NEDD8, ISG15, and FAT10 control numerous cellular processes including embryonic development, cell cycle progression and signal transduction, transport across the plasma membrane, protein quality control in the endoplasmic reticulum, transcriptional regulation, growth control, immune response, apoptosis, oncogenesis, preimplantation, and processes of the nervous system.
The overview of DUBs
Ubiquitination is an ATP dependent dynamic and reversible process, achieved by sequential enzymatic reactions. The reaction initiates with the activation of Ub by activating enzyme (E1) in ATP dependent manner, subsequently transfers the activated Ub to the cysteine residue of conjugating enzyme and finally attachment to the target protein is achieved by ligases (E3). Almost 600 E3 ligase enzymes have been identified that dictate substrate selection and regulate many cellular events. Like most posttranslational modifications, ubiquitination is reversible; Deubiquitinating enzymes (DUBs) counterbalance ubiquitin ligase activity by removing ubiquitin from target proteins (Figure 1). DUBs play a critical role at the proteasome where they are involved in editing ubiquitin chains and removing and recycling ubiquitin prior to substrate degradation in the proteasome. DUBs also regulate protein targeting and degradation in the lysosome.
Figure 1. The ubiquitin signaling system.
Experimental and bioinformatics approaches have identified almost 100 putative DUBs in human. However, more than 60% of these DUBs are not well characterized or enzymes with unknown functions. DUBs are classified into five distinct families and four out of the five subfamilies identified to date are cysteine proteases and they are classified as ubiquitin C-terminal hydrolases (UCH), the ubiquitin-specific proteases (USP/UBPs), Otubain domain ubiquitin-binding proteins (OTU), Machado-Joseph domain (Josephin domain)-containing (MJD) proteases. Fifth member of DUBs is Jab1/Pab1/MPN domain-containing (JAMM) protease, which belongs to metalloprotease family (Figure 2).
Figure 2. DUBs classification.
DUBs and diseases
Deubiquitinating enzymes and their involvement in diseases are predicted based on the physiological processes, which regulate disease states including neurological disorders and cancer. A number of USPs, which regulate neuronal development, are also associated with neurological disorders. The proteasome-associated protein USP14 plays an important role in synaptic development and dysregulation of USP14 leads to ataxia. USP14 mutant homozygous mice develop growth retardation and exhibit behavioral disorders including, hind limb paralysis. USP46-KO as well as USP46-MT (delta K92) mouse displays shorter tail suspension test immobility, used for assessing antidepressant activity and depression-like behavior, showing loss-of-function phenotype. USP46 is known to regulate pre and post-synaptic GABAergic signaling and maintenance of circadian rhythm. USP9X regulates neural fate.
There are growing numbers of studies demonstrating the role of DUBs in cancers (Figure 3). CYLD and BAP1 are identified as tumor suppressor genes, which undergo frequent mutations in different cancers. Mutations resulting in truncations, missenses, frame shifts in CYLD are linked to familial cylindromatosis and several cancers. Most of the catalytic domain mutations abolish its DUB activity, which is crucial for its tumor suppressor function. Since last five years, more than 400 mutations in BAP1 have been identified in different types of cancer including uveal melanoma mesothelioma, renal cell carcinoma, cholangiocarcinoma, melanocytic tumors, and lung cancers. Loss of deubiquitinase activity of BAP1 has been demonstrated for several mutants and this loss of DUB activity negotiates its tumor suppressor function. OTUB1 up regulation also leads to different types of cancer like bladder, lung, prostrate and breast. A20 acts as a tumor suppressor in B-cell lymphomas. Missense and deletions mutations are frequently found in A20 gene leading to Hodgkin lymphoma, mantle cell lymphoma, MALT lymphoma, and marginal zone lymphoma. A novel role of A20 has been identified in regeneration of muscle fibers and knockdown of A20 impairs muscle differentiation in vitro, suggesting its important role in muscular dystrophy.
Figure 3. Role of DUBs in different aspects of cancer epigenetic regulation.
Therapeutics and future perspective of DUBs
Ubiquitin signaling system is one of the complex signaling systems involved in majority of the cellular processes and deubiquitinating enzymes work as a central component. Aberrant DUBs function endorses human diseases and considerable interest has been emerged in evaluating their therapeutic potential. Even though, data on structural feature of DUBs is going high, still there is a lack of information on substrate specificity, functional, and structural diversity, and tissue specificity of each DUB. This information is necessary before targeting them with specific inhibitors. Therefore, much study and better and efficient technologies are still needed to elucidate both molecular structures and specific substrates for each DUB to validate and develop them in clinic. Using large-scale genomics and proteomics approaches, and carefully designed experimental model systems preceding clinical trials will be necessary for developing efficient DUB-based therapies. Nevertheless, comprehension derived from some studies optimistically will provide new insights into the manifold queries on DUBs, which could direct to the introduction of DUB-targeting strategies for molecular therapies against cancers.