Mechanistic insights into the post-translational regulation of RNA-binding proteins in cell life and disease

Abstract

Cell signaling networks modulate gene expression in response to internal and external stimuli. Rapid cellular adaptation to changing physiological and environmental conditions requires a strict control of the mRNA metabolism. The RNA-binding proteins (RBPs) are posttranscriptional gene regulators that determine the fate of mRNAs at each stage of their life cycle. Generally, the dynamic composition and properties of each cellular compartment exert an important influence on the function of the RBPs. But more specifically, RBP activities are tightly fine-tuned by a diverse array of post-translational modifications (PTMs) that govern protein stability, turnover, subcellular localization and interactions. Here, we have investigated the changes in liquid-liquid phase separation (LLPS) propensity and nucleo-cytoplasmic distribution of T-cell intracellular antigen 1 (TIA-1), a well-known RBP, upon phosphorylation at serines 198 and 199. For this, we resorted to a multidisciplinary approach that combined biochemical and computational methods, as well as the use of phosphomimetic mutations of serine to either aspartate or glutamate. Results uncovered a potential allosteric regulatory pathway for TIA-1 LLPS, by which S198/199 phosphorylation in RNA-recognition motif 3 (RRM3) would induce conformational rearrangements in the prion-related domain (PRD) that promotes stress granules (SGs) formation. Moreover, the same double phosphorylation would also increase TIA-1 cytoplasmic accumulation by impairing its association with target RNAs, presumably due to the reduced capacity of nuclear transcripts to retain this protein. Interestingly, although both aspartate and glutamate substitutions at S198/199 led to similar TIA-1 nucleo-cytoplasmic distributions, only the latter were capable of instigating LLPS and SGs formation, highlighting the importance of testing computationally and/or experimentally different alternatives to mimic phosphorylation. In addition, we have extended our knowledge on the molecular basis underlying the Zn2+-mediated LLPS of TIA-1 by studying the role of its RRM domains. We discovered that the Zn2+-chelating ability of histidines 94 and 96 of RRM2 facilitates the multimerization of TIA-1 in a PRD-independent manner. In sum, we propose herein novel mechanisms regulating TIA-1 subcellular localization, LLPS and recruitment into SGs.