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Integration of the catalytic subunit activates deneddylase activity in vivo as final step in fungal COP 9 signalosome assembly
Author(s) -
Beckmann Elena A.,
Köhler Anna M.,
Meister Cindy,
Christmann Martin,
Draht Oliver W.,
Rakebrandt Nikolas,
Valerius Oliver,
Braus Gerhard H.
Publication year - 2015
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1111/mmi.13017
Subject(s) - protein subunit , biology , ubiquitin , microbiology and biotechnology , mutant , biochemistry , gene
Summary The eight‐subunit COP 9 signalosome ( CSN ) is conserved from filamentous fungi to humans and functions at the interface between cellular signalling and protein half‐life control. CSN consists of six PCI and two MPN domain proteins and forms a scaffold for additional interacting proteins. CSN controls protein stability in the ubiquitin‐proteasome system where the MPN domain CSN 5/ CsnE subunit inactivates cullin‐ RING ligases. The CSN 5/ CsnE isopeptidase functions as deneddylase and removes the ubiquitin‐like protein N edd8. The six PCI domain proteins of human CSN form a horseshoe‐like ring and all eight subunits are connected by a bundle of C ‐terminal α‐helices. We show that single deletions of any csn subunit of A spergillus nidulans resulted in the lack of deneddylase activity and identical defects in the coordination of development and secondary metabolism. The CSN 1/ CsnA N ‐terminus is dispensable for deneddylase activity but required for asexual spore formation. Complex analyses in mutant strains revealed the presence of a seven‐subunit pre‐ CSN without catalytic activity. Reconstitution experiments with crude extracts of deletion strains and recombinant proteins allowed the integration of CSN 5/ CsnE into pre‐ CSN resulting in an active deneddylase. This supports a stable seven subunit pre‐ CSN intermediate where deneddylase activation in vivo can be controlled by CSN 5/ CsnE integration as final assembly step.