Biology subjects

Boag, P. R.

Publications and source records attributed to Boag, P. R..

5 recordsLinked to original sources

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↗

bioRxiv2019

WormCat: an online tool for annotation and visualization of Caenorhabditis elegans genome-scale data

The emergence of large gene expression datasets has revealed the need for improved tools to identify enriched gene categories and visualize enrichment patterns. While Gene Ontogeny (GO) provides a valuable tool for gene set enrichment analysis, it has several limitations. First, it is difficult to graphically compare multiple GO analyses. Second, genes from some model systems are not well represented. For example, around 30% of Caenorhabditis elegans genes are missing from analysis in commonly used databases. To allow categorization and visualization of enriched C. elegans gene sets in different types of genome-scale data, we developed WormCat, a web-based tool that uses a near-complete annotation of the C. elegans genome to identify co-expressed gene sets and scaled heat map for enrichment visualization. We tested the performance of WormCat using a variety of published transcriptomic datasets and show that it reproduces major categories identified by GO. Importantly, we also found previously unidentified categories that are informative for interpreting phenotypes or predicting biological function. For example, we analyzed published RNA-seq data from C. elegans treated with combinations of lifespan-extending drugs where one combination paradoxically shortened lifespan. Using WormCat, we identified sterol metabolism as a category that was not enriched in the single or double combinations but emerged in a triple combination along with the lifespan shortening. Thus, WormCat identified a gene set with potential phenotypic relevance that was not uncovered with previous GO analysis. In conclusion, WormCat provides a powerful tool for the analysis and visualization of gene set enrichment in different types of C. elegans datasets.

bioinformatics↗

bioRxiv2018

Widespread cytoplasmic polyadenylation programs asymmetry in the germline and early embryo

BACKGROUNDThe program of embryonic development is launched by selective activation of a silent maternal transcriptome. In Caenorhabditis elegans, nuclei of the adult germline are responsible for the synthesis of at least two distinct mRNA populations; those required for housekeeping functions, and those that program the oocyte-to-embryo transition. We mapped this separation by changes to the length-distribution of poly(A)-tails that depend on GLD-2 mediated cytoplasmic polyadenylation and its regulators genome-wide.\n\nRESULTSMore than 1000 targets of cytoplasmic polyadenylation were identified by differential polyadenylation. Amongst mRNA with the greatest dependence on GLD-2 were those encoding RNA binding proteins with known roles in spatiotemporal patterning such as mex-5 and pos-1. In General, the 3 UTR of GLD-2 targets were longer, contained cytosine-patches, and were enriched for non-standard polyadenylation-motifs. To identify the deadenylase that initiated transcript silencing, we depleted the known deadenylases in the gld-2(0) mutant background. Only the loss of CCF-1 suppressed the short-tailed phenotype of GLD-2 targets suggesting that in addition to its general role in RNA turnover, this is the major deadenylase for regulatory silencing of maternal mRNA. Analysis of poly(A)-tail length-change in the embryo lacking specific RNA-binding proteins revealed new candidates for asymmetric expression in the first embryonic divisions.\n\nCONCLUSIONThe concerted action of RNA binding proteins exquisitely regulates GLD-2 activity in space and time. We present our data as interactive web resources for a model where GLD-2 mediated cytoplasmic polyadenylation regulates target mRNA at each stage of worm germline and early embryonic development.

developmental biology↗

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↗

bioRxiv2018

The chromatin modifiers SET-25 and SET-32 are required for initiation but not long-term maintenance of transgenerational epigenetic inheritance

Some epigenetic modifications are inherited from one generation to the next, providing a potential mechanism for the inheritance of environmentally acquired traits. Transgenerational inheritance of RNA interference phenotypes in Caenorhabditis elegans provides an excellent model to study this phenomenon, and whilst studies have implicated both chromatin modifications and small RNA pathways in heritable silencing their relative contributions remain unclear. Here we demonstrate that the histone methyltransferases SET-25 and SET-32 are required for the establishment of a transgenerational silencing signal but not for long-term maintenance of this signal between subsequent generations, suggesting that transgenerational epigenetic inheritance is a multi-step process with distinct genetic requirements for establishment and maintenance of heritable silencing. Furthermore, small RNA sequencing reveals that the abundance of secondary siRNAs (thought to be the effector molecules of heritable silencing) does not correlate with silencing phenotypes. Together, our results suggest that the current mechanistic models of epigenetic inheritance are incomplete.

genetics↗