Biology subjects

Wilce, J. A.

Publications and source records attributed to Wilce, J. A..

3 recordsLinked to original sources

bioRxiv2025

Allosteric modulation of TIA-1 phase separation by double serine phosphorylation

In response to diverse harmful stimuli, eukaryotic cells generate cytoplasmic stress granules (SGs), mainly composed of mRNAs and RNA-binding proteins (RBPs). RBPs are fine-tuned by a diverse array of post-translational modifications (PTMs), with important consequences for the assembly, dynamics and clearance of SGs. One of the best characterized SG nucleators is the RBP T-cell intracellular antigen 1 (TIA-1), although knowledge about the structural and functional impact of its identified PTMs is very limited. TIA-1 is organized into three RNA-recognition motifs (RRMs) and a C-terminal prion-related domain (PRD) that drives its phase separation from the cytosol. Here, we analyzed the effect of TIA-1 double phosphorylation in RRM3, at serines 198 and 199. Microscopic observations revealed an increased propensity of the phosphomimetic TIA-1 S198/199E to undergo liquid-liquid phase separation (LLPS) and self-assemble into SGs independently of stress stimuli. Our computational simulations, supported by NMR data, have suggested that such phosphorylations promote the formation of a {beta}-hairpin motif at the beginning of the PRD. Moreover, the ALS-associated mutation V283M in TIA-1 was predicted to lead to the formation of an aberrant structure in the {beta}-hairpin region, highlighting the fine balance between physiological and pathogenic TIA-1 phase transition, and the importance of a better understanding of the molecular mechanisms underlying the liquid demixing of this RBP.

biophysics↗

bioRxiv2024

The RNA-binding activity of the TRIM-NHL protein NHL-2 is essential for miRNA-mediated gene regulation

The conserved TRIM-NHL protein, NHL-2, plays a key role in small RNA pathways in Caenorhabditis elegans. NHL-2 has been shown to interact with U-rich RNA through its NHL domain, but the importance to its biological function is unknown. We defined the crystal structure of the NHL domain to 1.4 [A] resolution and identified residues that affect affinity for U-rich RNA. Functional analysis of an NHL-2 RNA-binding loss-of-function mutant demonstrated defects in the heterochronic pathway, suggesting that RNA binding is essential for its role in this miRNA pathway. Processing bodies were enlarged in the NHL-2 RNA-binding mutant, suggesting a defect in mRNA decay. We also identified the eIF4E binding protein IFET-1 as a strong synthetic interactor with NHL-2 and the DEAD box RNA helicase CGH-1 (DDX6), linking NHL-2 function to translation repression. We demonstrated that in the absence of NHL-2, there was an enrichment of miRNA transcripts associated with the miRNA pathway Argonaute proteins ALG-2 and ALG-2. We demonstrate that NHL-2 RNA-binding activity is essential for let-7 family miRNA-mediated translational repression. We conclude that the NHL-2, CGH-1, and IFET-1 regulatory axes work with the core miRISC components to form an effector complex that is required for some, but not all, miRNAs.

genetics↗

bioRxiv2018

The TRIM-NHL protein NHL-2 is a Novel Co-Factor of the CSR-1 and HRDE-1 22G-RNA Pathways

Proper regulation of germline gene expression is essential for fertility and maintaining species integrity. In the C. elegans germline, a diverse repertoire of regulatory pathways promote the expression of endogenous germline genes and limit the expression of deleterious transcripts to maintain genome homeostasis. Here we show that the conserved TRIM-NHL protein, NHL-2, plays an essential role in the C. elegans germline, modulating germline chromatin and meiotic chromosome organization. We uncover a role for NHL-2 as a co-factor in both positively (CSR-1) and negatively (HRDE-1) acting germline 22G-small RNA pathways and the somatic nuclear RNAi pathway. Furthermore, we demonstrate that NHL-2 is a bona fide RNA binding protein and, along with RNA-seq data point to a small RNA independent role for NHL-2 in regulating transcripts at the level of RNA stability. Collectively, our data implicate NHL-2 as an essential hub of gene regulatory activity in both the germline and soma.

genetics↗