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In an effort to investigate how loss of deneddylation affects SCF activity, …


Biology Articles » Biochemistry » Protein Biochemistry » Targeted silencing of Jab1/Csn5 in human cells downregulates SCF activity through reduction of F-box protein levels

Abstract
- Targeted silencing of Jab1/Csn5 in human cells downregulates SCF activity through reduction of F-box protein levels

Targeted silencing of Jab1/Csn5 in human cells downregulates SCF activity through reduction of F-box protein levels

Gregory A Cope1,3 and Raymond J Deshaies1,2

1Department of Biology, California Institute of Technology Pasadena, CA 91125, USA
2Howard Hughes Medical Institute, 4000 Jones Bridge Road, Chevy Chase, MD, 20815-6789, USA
3Department of Biology, Stanford University, Palo Alto, CA 94305, USA

 

Background

SCF ubiquitin ligases target numerous proteins for ubiquitin dependent proteolysis, including p27 and cyclin E. SCF and other cullin-RING ligases (CRLs) are regulated by the ubiquitin-like protein Nedd8 that covalently modifies the cullin subunit. The removal of Nedd8 is catalyzed by the Jab1/MPN domain metalloenzyme (JAMM) motif within the Csn5 subunit of the Cop9 Signalosome.

Results

Here, we conditionally knock down Csn5 expression in HEK293 human cells using a doxycycline-inducible shRNA system. Cullin levels were not altered in CSN-deficient human cells, but the levels of multiple F-box proteins were decreased. Molecular analysis indicates that this decrease was due to increased Cul1- and proteasome-dependent turnover. Diminished F-box levels resulted in reduced SCF activity, as evidenced by accumulation of two substrates of the F-box protein Fbw7, cyclin E and c-myc, in Csn5-depleted cells.

Conclusion

We propose that deneddylation of Cul1 is required to sustain optimal activity of SCF ubiquitin ligases by repressing 'autoubiquitination' of F-box proteins within SCF complexes, thereby rescuing them from premature degradation.

BMC Biochemistry 2006, 7:1. This is an Open Access article distributed under the terms of the Creative Commons Attribution License.


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