Disordered Motifs and Domains in Cell Control - October 11-15, 2014

Disordered Motifs and Domains in Cell Control

Poster Session I

18-POS Board 18 Multivalent Interaction of Higher-order Oligomers of SPOP with the Transcriptional Activator Gli3

Jihun Lee , Melissa R. Marzahn, Wendy K. Pierce, Tanja Mittag. St. Jude Children's Research Hospital, Memphis, TN, USA.

Multivalent interactions, in which each binding partner has multiple binding sites or binding motifs for the other protein, can lead to the assembly of large higher-order complexes allowing for subcellular organization. Speckle-type POZ protein (SPOP), recently identified as a novel tumor suppressor, localizes to nuclear puncta and is a substrate adaptor of a cullin3-RING ubiquitin ligase (CRL). It recruits substrates to the CRL and promotes their ubiquitination and downstream degradation. SPOP self-associates into large higher-order homo-oligomers through its two independent oligomerization domains, rendering it multivalent for substrates. Some SPOP substrates, such as Gli3, reportedly harbor many SPOP binding motifs. Here, we identified SPOP binding motifs in Gli3 and determined that SPOP self-associates indefinitely through an isodesmic self-association mechanism. Using a variety of biophysical methods including dynamic light scattering (DLS), composition gradient multi-angle light scattering (CG-MALS), biolayer interferometry (BLI) and cellular assays we studied the multivalent interaction between SPOP and Gli3. We investigated self-association and substrate binding of SPOP mutants found in cancers comparing physiological and pathological SPOP properties. We determined factors promoting SPOP localization in nuclear punctate structures. The formation of higher-order complexes may result in subcellular organization but may also lead to activity enhancement through high local concentrations of components, and ultrasensitive regulation of signaling pathways.

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