Multiple Arkadia/RNF111 structures coordinate its Polycomb body association and transcriptional control

The RING domain protein Arkadia/RNF111 is a ubiquitin ligase in the transforming growth factor beta (TGFß) pathway. We previously identified Arkadia as a small ubiquitin-like modifier (SUMO)-binding protein with clustered SUMO-interacting motifs (SIMs) that together form a SUMO-binding domain (SBD). However, precisely how SUMO interaction contributes to the function of Arkadia was not resolved. Through analytical molecular and cell biology, we found that the SIMs share redundant function with Arkadia''s M domain, a region distinguishing Arkadia from its paralogs ARKL1/ARKL2 and the prototypical SUMO-targeted ubiquitin ligase (STUbL) RNF4. The SIMs and M domain together promote both Arkadia''s colocalization with CBX4/Pc2, a component of Polycomb bodies, and the activation of a TGFbeta pathway transcription reporter. Transcriptome profiling through RNA sequencing showed that Arkadia can both promote and inhibit gene expression, indicating that Arkadia''s activity in transcriptional control may depend on the epigenetic context, defined by Polycomb repressive complexes and DNA methylation [Sun, Liu, and Hunter (2014) Mol Cell Biol 34(16):2981-2995]. Overall design: To determine the role of Arkadia in TGFß signaling at the transcriptome level, the profiles of TGFß-stimulated gene expression were examined in Ark+/+ (Ark_WT), Ark-/- (Ark_null), and Arkadia (WT and sim mutant)-reconstituted Ark-/- MEFs. RNA sequencing was carried out using poly(A)-enriched RNA samples from unstimulated cells and cells treated with TGFß for 1h, 4h, or 16h as indicated. Two batches of sequencing data for a total of 16 independent samples were submitted.

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Sun H, Liu Y, Hunter T