Intrinsically disordered proteins (IDPs) regularly constitute components of larger protein assemblies contributing to architectural stability. Two small, highly acidic IDPs have been linked to the so-called PCI complexes carrying PCI-domain subunits, including the proteasome lid and the COP9 signalosome. These two IDPs, DSS1 and CSNAP, have been proposed to have similar structural propensities and functions, but they display differences in their interactions and interactome sizes. Here we characterized the structural properties of human DSS1 and CSNAP at the residue level using NMR spectroscopy and probed their propensities to bind ubiquitin. We find that distinct structural features present in DSS1 are completely absent in CSNAP, and vice versa, with lack of relevant ubiquitin binding to CSNAP, suggesting the two proteins to have diverged in both structure and function. Our work additionally highlights that different local features of seemingly similar IDPs, even subtle sequence variance, may endow them with different functional traits. Such traits may underlie their potential to engage in multiple interactions thereby impacting their interactome sizes.
Intrinsically disordered proteins (IDPs) are often multifunctional and frequently posttranslationally modified. Deleted in split hand/split foot 1 (Dss1 – Sem1 in budding yeast) is a highly multifunctional IDP associated with a range of protein complexes. However, it remains unknown if the different functions relate to different modified states. In this work we show that S. pombe Dss1 is a substrate for casein kinase 2 in vitro and we identify three phosphorylated threonines in its linker region separating two known disordered ubiquitin binding motifs. Phosphorylations of the threonines had no effect on ubiquitin binding, but caused a slight destabilization of the C‐terminal α‐helix and mediated a direct interaction with the forkhead‐associated domain (FHA) of the RING‐FHA E3‐ubiquitin ligase defective in mitosis 1 (Dma1). The phosphorylation sites are not conserved and are absent in human Dss1. Sequence analyses revealed that the Txx(E/D) motif, which is important for phosphorylation and Dma1 binding, is not linked to certain branches of the evolutionary tree. Instead, we find that the motif appears randomly, supporting the mechanism of ex nihilo evolution of novel motifs. In support of this, other threonine‐based motifs, although frequent, are non‐conserved in the linker, pointing to additional functions connected to this region. We suggest that Dss1 acts as an adaptor protein that docks to Dma1 via the phosphorylated FHA‐binding motifs, while the C‐terminal α‐helix is free to bind mitotic septins, thereby stabilizing the complex. The presence of Txx(D/E) motifs in the disordered regions of certain septin subunits may be of further relevance to the formation and stabilization of these complexes.This article is protected by copyright. All rights reserved.
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