Type of presentation: Poster

LS-4-P-5839 Structural Investigations of COP9 signalosome binding to Cullin-RING ligases:promiscuous interactions sharing common regulatory principles

Cavadini S.^1,2, Fischer E. S.^1,2, Goldie K. N.³, Böhm K.^1,2, Lingaraju G. M.^1,2, Bunker R. D.^1,2, Pantelic R. S.³, Mohamed W. I.^1,2, Stahlberg H.³, Thomä N. H.^1,2

¹Friedrich Miescher Institute for Biomedical Research, Maulbeerstrasse 66, 4058 Basel, Switzerland., ²University of Basel, Petersplatz 10, 4003 Basel, Switzerland., ³Center for Cellular Imaging and NanoAnalytics, Biozentrum, University of Basel, 4058 Basel, Switzerland.

k.goldie@unibas.ch

The CUL4A-RBX1-DDB1-DDB2 ligase (CRL4A^DDB2) is the primary DNA damage-sensing complex in the repair of UV light induced pyrimidine dimers¹. The class of cullin-RING ubiquitin ligases (CRLs) comprises six canonical cullin families (Cul1, Cul2, Cul3, Cul4A, Cul4B, and Cul5) which together with the RING-domain proteins Rbx1 or Rbx2 mediate the ubiqutination ~20% of the proteins degraded by the proteasome². CRLs assembly is modular, with sets of receptors and adaptors giving rise to hundreds of distinct cullin-RING E3 ubiquitin ligase complexes³. Regulation of this structurally diverse family of ligases depends on the COP9 signalosome (CSN), an 8-subunit isopeptidase that removes the covalently conjugated Nedd8 activator from the cullin^4-6. In addition to its catalytic function, CSN form tight complexes with the unmodified CRL, a process implicated in preventing the CRL substrate adaptor from undergoing cycles of futile auto-ubiquitination and degradation in vivo^7-9. CSN inhibition is overcome once the CRL binds a substrate. The mechanism underlying CSN binding to structurally diverse CRL complexes across families remains unclear. Here we present the structure of CRL4 family and a dimeric CRL3 in complex with CSN using single-particle electron microscopy (EM). We find that the conserved cullin C-terminus together with Rbx1 is held by CSN2 and CSN4 subunits. In the CRL1, CRL3 and CRL4 family the divergent receptors and adaptors form a variety of contacts that are unexpectedly plastic in nature involving CSN1, CSN3 and CSN helical bundle. Moreover, we show that: (i) the ligase substrates can access the CSN-bound cullin receptors in a similar fashion. (ii) Depending on the size of the substrate, CSN is released by steric means. Altogether our findings imply a model where steric repulsion by the cognate substrate allow CSN-mediated regulation of ~300 different CRL enzymes in response to different cues, without the need for dedicated interactions or common motifs.

References;

1. Scrima, A. et al. FEBS letters 585 (18), 2818-25. (2011).

2. Soucy, T. A. T. et al. Audio, Transactions of the IRE Professional Group 458, 732-736, (2009).

3. Deshaies, R. J. & Joazeiro, C. A. P. Annual review of biochemistry 78, 399-434, (2009).

4. Schwechheimer, C. et al. Science (New York, NY) 292, 1379-1382, (2001).

5. Lyapina, S. et al. Science (New York, NY) 292, 1382-1385, (2001).

6. Wei, N., Chamovitz, D. A. & Deng, X. W. Cell 78, 117-124, (1994).

7. Fischer, E. S. et al. Cell 147, 1024-1039, (2011).

8. Enchev, R. I. et al. Cell Reports, 1-23, (2012).

9. Emberley, E. D., Mosadeghi, R. & Deshaies, R. J. Journal of biological chemistry 287, 29679-29689, (2012).

Fig. 1: Cryo-EM structure and pseudo-atomic model of the CSN-N8CRL4^DDB2 complex.