Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Catalytic residues of microRNA Argonautes play a modest role in microRNA star strand destabilization in C. elegans [miRNA]

ABSTRACT: Many Argonaute proteins can cleave RNA (“slicing”) as part of the microRNA-induced silencing complex (miRISC), even though miRNA-mediated target repression is generally independent of target cleavage. Here we use C. elegans to examine the role of catalytic residues of miRNA Argonautes in organismal development. In contrast to previous work, mutations in presumed catalytic residues did not interfere with normal development when introduced by CRISPR. We find that unwinding and decay of miRNA star strands is weakly defective in the catalytic residue mutants, with the largest effect observed in embryos. Argonaute-Like Gene 2 (ALG-2) is more dependent on slicing for unwinding than ALG-1. The miRNAs that displayed the greatest (albeit minor) dependence on catalytic residues for unwinding tend to form stable duplexes with their star strand, and in some cases, lowering duplex stability alleviates dependence on catalytic residues. While a few miRNA guide strands are reduced in the mutant background, the basis of this is unclear since changes were not dependent on EBAX-1, an effector of Target-Directed miRNA Degradation (TDMD). Overall, this work defines a role for the catalytic residues of miRNA Argonautes in star strand decay; future work should examine whether this role may contribute to the selection pressure to conserve catalytic activity of miRNA Argonautes across the metazoan phylogeny.

ORGANISM(S): Caenorhabditis elegans

PROVIDER: GSE256489 | GEO | 2024/02/26

REPOSITORIES: GEO

ACCESS DATA

Json Xml

Shared Molecules

Only show the datasets with similarity scores above: 0.5

Threshold

0.5

Similar Datasets

Catalytic residues of microRNA Argonautes play a modest role in microRNA star strand destabilization in C. elegans

Project description:Many Argonaute proteins can cleave RNA (“slicing”) as part of the microRNA-induced silencing complex (miRISC), even though miRNA-mediated target repression is generally independent of target cleavage. Here we use C. elegans to examine the role of catalytic residues of miRNA Argonautes in organismal development. In contrast to previous work, mutations in presumed catalytic residues did not interfere with normal development when introduced by CRISPR. We find that unwinding and decay of miRNA star strands is weakly defective in the catalytic residue mutants, with the largest effect observed in embryos. Argonaute-Like Gene 2 (ALG-2) is more dependent on slicing for unwinding than ALG-1. The miRNAs that displayed the greatest (albeit minor) dependence on catalytic residues for unwinding tend to form stable duplexes with their star strand, and in some cases, lowering duplex stability alleviates dependence on catalytic residues. While a few miRNA guide strands are reduced in the mutant background, the basis of this is unclear since changes were not dependent on EBAX-1, an effector of Target-Directed miRNA Degradation (TDMD). Overall, this work defines a role for the catalytic residues of miRNA Argonautes in star strand decay; future work should examine whether this role may contribute to the selection pressure to conserve catalytic activity of miRNA Argonautes across the metazoan phylogeny.

2024-02-26 | GSE256488 | GEO

Defining the contribution of microRNA-specific slicing Argonautes in animals

Project description:microRNAs regulate gene expression through interaction with an Argonaute protein family member. While some members of this protein family retain an enzymatic activity capable of cleaving RNA molecules complementary to Argonaute-bound small RNAs, the role of the slicing activity in the canonical microRNA pathway is still unclear in animals. To address the importance of slicing Argonautes in animals, we created Caenorhabditis elegans mutant strains carrying catalytically dead endogenous ALG-1 and ALG-2, the only two slicing Argonautes essential for the miRNA pathway in this animal model. We observe that the loss of ALG-1 and ALG-2 slicing activity affects overall animal fitness and causes phenotypes reminiscent of miRNA defects only when grown and maintained at restrictive temperature. Furthermore, the analysis of global miRNA expression show that the catalytic activity of ALG-1 and ALG-2 differentially regulate the level of specific subsets of miRNAs in young adults. We also demonstrate that altering the slicing activity of those miRNA-specific Argonautes does not result in any defect in the production of canonical miRNAs. Together, these data support that the slicing activity of miRNA-specific Argonautes functions to maintain levels of a set of miRNAs for optimal viability and fitness in animals particularly exposed to specific growing conditions.

2024-03-15 | GSE222733 | GEO

Dipeptidyl peptidase DPF-3 is a gatekeeper of microRNA Argonaute compensation in animals

Project description:MicroRNAs (miRNAs) are essential regulators involved in multiple biological processes. To achieve their gene repression function, they are loaded in miRNA-specific Argonautes to form the miRNA-induced silencing complex (miRISC). Mammals and C. elegans possess more than one paralog of miRNA-specific Argonautes but the dynamic between them remains unclear. Here, we report the conserved dipeptidyl peptidase DPF-3 as a new interactor of the miRNA-specific Argonautes ALG-1 and ALG-2 in C. elegans. Knockout of dpf-3 increases ALG-2 levels and miRISC formation in alg-1 null animals, thereby compensating for ALG-1 loss and rescuing miRNA-related defects observed. DPF-3 can cleave an ALG-2 N-terminal peptide in vitro but does not appear to rely on this catalytic activity to regulate ALG-2 in vivo. This study uncovers the importance of DPF-3 in the miRNA pathway and provides insights on how multiple miRNA Argonautes contribute to achieve proper miRNA-mediated gene regulation in animals.

2025-03-06 | PXD055867 | Pride

A specific Argonaute phosphorylation regulates the binding to microRNAs during C. elegans development.

Project description:Argonaute proteins are at the core of the microRNA-mediated gene silencing pathway essential for animals. In C. elegans, the microRNA-specific Argonautes ALG-1 and ALG-2 regulate multiple processes required for proper animal developmental timing and viability. Here, we identified a new phosphorylation site, serine 642, on ALG-1 that modulates microRNA association. Mutating ALG-1 serine 642 into a phospho-mimicking residue impairs microRNA binding and causes embryonic lethality and post-embryonic phenotypes that are common with alteration of microRNA functions. Monitoring microRNA levels in alg-1 phosphorylation mutant animals reveal that miRNA passenger strands strongly increase but are not preferentially loaded into ALG-1, indicating that the miRNA binding defects could also lead to miRNA duplexes accumulation. Our genetic and biochemical experiments support the protein kinase A KIN-1 as the putative kinase that phosphorylates ALG-1 serine 642. Altogether, our data indicate that PKA triggers the ALG-1 phosphorylation to regulate its microRNAs association during C. elegans development.

2022-09-10 | GSE198352 | GEO

Dipeptidyl peptidase DPF-3 is a gatekeeper of microRNA Argonaute compensation in animals

Project description:MicroRNAs (miRNAs) are essential regulators involved in multiple biological processes. To achieve their gene repression function, they are loaded in miRNA-specific Argonautes to form the miRNA-induced silencing complex (miRISC). Mammalians and C. elegans intriguingly possess more than one paralog of miRNA-specific Argonautes and the dynamic between them remains unclear. Here, using C. elegans as a model system, we report dipeptidyl peptidase DPF-3 as a new interactor of miRNA-specific Argonautes ALG-1 and ALG-2. Knockout of dpf-3 rescues miRNA-related defects observed in alg-1 null animals and allows ALG-2 to better compensate for the lack of its paralog through higher expression and activity. While DPF-3 can cleave a N-terminal dipeptide of ALG-2 in vitro, we show that it is likely not the molecular mechanism used by DPF-3 to regulate ALG-2. This study uncovers the importance of a dipeptidyl peptidase in the miRNA pathway and provide insights in miRNA-specific Argonautes regulation dynamics in animals.

2025-03-06 | GSE276186 | GEO

The Making of a Slicer: Activation of Human Argonaute-1

Project description:The Argonautes are the central protein component in small RNA silencing pathways. Of the four human Argonautes (hAgo1-4) only hAgo2 is an active slicer. We determined the structure of hAgo1 bound to endogenous copurified RNAs to 1.75 Å resolution and hAgo1 loaded with let-7 miRNA to 2.1 Å. Both structures are strikingly similar to the structures of hAgo2. A conserved DEDH catalytic tetrad within the PIWI domain of hAgo2 is required for its slicing activity. hAgo3, with an intact tetrad, could be activated by swapping in the N domain of hAgo2, without additional PIWI domain mutations. An additional mutation on a loop adjacent to the active site of hAgo1 results in slicer activity that is substantially enhanced by swapping in the N domain of hAgo2. Intriguingly, the elements that make Argonaute an active slicer involve a sophisticated interplay between the active site and more distant regions of the enzyme. RNAsprofiling of small RNAs associated with recombinant protein used for chrystallographic studies

2013-05-20 | E-GEOD-46870 | biostudies-arrayexpress

Caenorhabditis elegans ALG-1 antimorphic mutations uncover functions for Argonaute in microRNA guide strand selection and passenger strand disposal

Project description:MicroRNAs are regulators of gene expression whose functions are critical for normal development and physiology. We have previously characterized mutations in a Caenorhabditis elegans microRNA-specific Argonaute ALG-1 (Argonaute-like gene) that are antimorphic [alg-1(anti)]. alg-1(anti) mutants have dramatically stronger microRNA-related phenotypes than animals with a complete loss of ALG-1. ALG-1(anti) miRISC (microRNA induced silencing complex) fails to undergo a functional transition from microRNA processing to target repression. To better understand this transition, we characterized the small RNA population associated with ALG-1(anti) complexes in vivo. alg-1(anti) mutants dramatically overaccumulated microRNA* (passenger) strands, and immunoprecipitated ALG-1(anti) complexes contained nonstoichiometric yields of mature microRNA and microRNA* strands, with some microRNA* strands present in the ALG-1(anti) Argonaute far in excess of the corresponding mature microRNAs. We show complex and microRNA-specific defects in microRNA strand selection and microRNA* strand disposal. For certain microRNAs (for example mir-58), microRNA guide strand selection by ALG-1(anti) appeared normal, but microRNA* strand release was inefficient. For other microRNAs (such as mir-2), both the microRNA and microRNA* strands were selected as guide by ALG-1(anti), indicating a defect in normal specificity of the strand choice. Our results suggest that wild-type ALG-1 complexes recognize structural features of particular microRNAs in the context of conducting the strand selection and microRNA* ejection steps of miRISC maturation.

2015-09-07 | GSE72659 | GEO

Modeling neurodevelopmental disorder-associated human AGO1 mutations in C. elegans Argonaute alg-1. [RNA-Seq]

Project description:MicroRNAs (miRNA) associate with Argonaute proteins and negatively regulate gene expression by base pairing with complementary sequences in the 3’ UTRs of target genes. De novo coding variants in the human Argonaute gene AGO1 were reported to cause neurodevelopmental disorder (NDD) with intellectual disability (ID). Most of the altered amino acids are conserved between the miRNA associated Argonautes in H. sapiens and C. elegans, suggesting that the human AGO1 mutations could disrupt evolutionarily conserved functions in miRNA biogenesis or target repression. We genetically modeled four human AGO1 mutations in C. elegans by introducing identical mutations into the C. elegans AGO1 homolog, ALG-1. These alg-1 NDD mutations caused phenotypes in C. elegans indicative of disrupted miRNA processing, miRISC formation, and/or target repression. We show that the alg-1 NDD mutations are antimorphic as they cause developmental and molecular phenotypes stronger than those exhibited by the alg-1 null mutants, likely by sequestrating functional miRNA silencing complex (miRISC) components into non-functional complexes that fail to confer robust gene repression. The alg-1 NDD mutations cause allele-specific disruptions in mature miRNA profiles, both in overall abundances and ALG-1 NDD association, accompanied by perturbation of downstream gene expression, including altered translational efficiency and/or mRNA abundance. The perturbed genes include those with human orthologs whose dysfunction is associated with NDD. These cross-clade genetic studies illuminate fundamental Argonaute functions and provide insights into the conservation of miRNA-mediated post-transcriptional regulatory mechanisms.

2023-12-30 | GSE252065 | GEO

Modeling neurodevelopmental disorder-associated human AGO1 mutations in C. elegans Argonaute alg-1. [smallRNA-Seq]

2023-12-30 | GSE252064 | GEO