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Downregulation of COP9 signalosome subunits differentially affects the CSN complex and target protein stability

DOI: 10.1186/1471-2091-8-27

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Permanent knockdowns of CSN1 and CSN3 led to a reduction of the subunits to approximately 40%, which is accompanied by a proportional decrease of the CSN holocomplex. In contrast, downregulation of CSN5 in HeLa cells reduced the CSN5 protein below 20% without significant effects on the remaining complex. The CRL component Rbx1 was characterized by accelerated proteolysis in siCSN1 and siCSN3 and also in siCSN5 cells, however, with lesser extent. Immunoprecipitated CSN complex from siCSN5 cells was less effective in phosphorylating c-Jun and p27. Accelerated degradation of c-Jun in siCSN5 cells was rescued by overexpression of CSN5 as well as of the deneddylation mutant CSN5D151N. Overexpression of CSN5 cannot rescue c-Jun destabilization in siCSN1.There exists a coordinated downregulation of CSN subunits in the CSN1 and CSN3 knockdowns. The underlying regulatory mechanisms are obscure. CSN5 seems to possess a specific status in HeLa cells. Its reduction is not connected with coordinated downregulation of other subunits. CSN knockdowns confirm that the stabilization of the CRL component Rbx1 is a major CSN function. In addition, downregulation of CSN subunits influences the stability of important cellular regulators such as c-Jun and p27.The COP9 signalosome (CSN) is a conserved protein complex, which controls eukaryotic protein degradation via the ubiquitin (Ub) proteasome system (UPS) [1,2]. In mammals the core complex consists of 8 subunits (CSN1 to CSN8) [3], the exact function of which is not exactly known. CSN5 exhibits a MPN+/JAMM domain [4,5] responsible for metalloprotease activity. As a complex-bound protein CSN5 removes NEDD8, an ubiquitin-like protein, from cullins. This cleavage of an isopeptide bond called deneddylation controls the ubiquitination by cullin-RING Ub ligases (CRLs), a large family of multisubunit E3s [6,7]. Cullin proteins (Cul1 to Cul7) are components of CRLs functioning as scaffolds of the Ub ligase complexes. Cullin neddylation and den

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