This review discusses the regulation and cellular roles of ubiquitin-specific deubiquitinating enzymes (DUBs). DUBs are proteases that reverse the modification of proteins by a single ubiquitin and remodel polyubiquitin chains on target proteins. The human genome encodes nearly 100 DUBs, which are classified into five families: UCH, USP, OTU, Josephin, and JAMM. Most DUB activity is cryptic, requiring specific binding to substrates or scaffolding proteins to achieve catalytically competent conformation. DUBs contain multiple domains that modulate substrate specificity, protein-protein interactions, and cellular localization. Binding partners and multi-protein complexes modulate DUB activity and substrate specificity. Recent studies using quantitative assays, genetic approaches, and RNAi have provided new insights into the function of yeast and human DUBs. The review covers the general properties of DUBs, including their modular structure, substrate-induced conformational changes, and regulation by scaffold or adapter binding. It also discusses the roles of DUBs in various cellular processes, such as protein degradation, DNA repair, chromatin remodeling, cell cycle regulation, endocytosis, and signaling kinase modulation. Specific examples are provided from studies on protein degradation, DNA repair, chromatin remodeling, cell cycle regulation, endocytosis, and signaling kinases.This review discusses the regulation and cellular roles of ubiquitin-specific deubiquitinating enzymes (DUBs). DUBs are proteases that reverse the modification of proteins by a single ubiquitin and remodel polyubiquitin chains on target proteins. The human genome encodes nearly 100 DUBs, which are classified into five families: UCH, USP, OTU, Josephin, and JAMM. Most DUB activity is cryptic, requiring specific binding to substrates or scaffolding proteins to achieve catalytically competent conformation. DUBs contain multiple domains that modulate substrate specificity, protein-protein interactions, and cellular localization. Binding partners and multi-protein complexes modulate DUB activity and substrate specificity. Recent studies using quantitative assays, genetic approaches, and RNAi have provided new insights into the function of yeast and human DUBs. The review covers the general properties of DUBs, including their modular structure, substrate-induced conformational changes, and regulation by scaffold or adapter binding. It also discusses the roles of DUBs in various cellular processes, such as protein degradation, DNA repair, chromatin remodeling, cell cycle regulation, endocytosis, and signaling kinase modulation. Specific examples are provided from studies on protein degradation, DNA repair, chromatin remodeling, cell cycle regulation, endocytosis, and signaling kinases.