Quantitative analysis of C. elegans transcripts by Nanopore direct-cDNA sequencing reveals terminal hairpins in non trans-spliced mRNAs

Bernard, Florian; Dargère, Delphine; Rechavi, Oded; Dupuy, Denis

doi:10.1038/s41467-023-36915-0

Cited by 10 publications

(5 citation statements)

References 30 publications

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“…Of the 4144 (SNA-3) and 2709 (SUT-1) peaks that were no more than 1000 nucleotides away from a TSS, 91% and 86% were upstream of the 5′ end of the CDS, whereas 9% and 14% were downstream of the 5′ end of the CDS (Figure 5F ). These numbers are roughly consistent with the proportion of trans -spliced genes (85 – 87%) determined from genome-wide studies ( 8 , 46 ). Out of 3764 and 2340 genes where we predicted trans -splicing because the respective SNA-3 or SUT-1 peak was upstream of the 5′ end of the CDS (Figure 5F ), 75.0% (SNA-3) and 63.0% (SUT-1) had a bona fide trans -splice acceptor site upstream of the 5′ end of the CDS.…”

Section: Resultssupporting

confidence: 86%

A nematode-specific ribonucleoprotein complex mediates interactions between the major nematode spliced leader snRNP and its target pre-mRNAs

Eijlers,

Al-Khafaji,

Soto-Martin

et al. 2024

Nucleic Acids Research

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Spliced leader trans-splicing of pre-mRNAs is a critical step in the gene expression of many eukaryotes. How the spliced leader RNA and its target transcripts are brought together to form the trans-spliceosome remains an important unanswered question. Using immunoprecipitation followed by protein analysis via mass spectrometry and RIP-Seq, we show that the nematode-specific proteins, SNA-3 and SUT-1, form a complex with a set of enigmatic non-coding RNAs, the SmY RNAs. Our work redefines the SmY snRNP and shows for the first time that it is essential for nematode viability and is involved in spliced leader trans-splicing. SNA-3 and SUT-1 are associated with the 5′ ends of most, if not all, nascent capped RNA polymerase II transcripts, and they also interact with components of the major nematode spliced leader (SL1) snRNP. We show that depletion of SNA-3 impairs the co-immunoprecipitation between one of the SL1 snRNP components, SNA-2, and several core spliceosomal proteins. We thus propose that the SmY snRNP recruits the SL1 snRNP to the 5′ ends of nascent pre-mRNAs, an instrumental step in the assembly of the trans-spliceosome.

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Section: Resultssupporting

confidence: 86%

A nematode-specific ribonucleoprotein complex mediates interactions between the major nematode spliced leader snRNP and its target pre-mRNAs

Eijlers,

Al-Khafaji,

Soto-Martin

et al. 2024

Nucleic Acids Research

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“…Therefore, the trans-spliceosome needs to bring two separate RNA molecules together. The majority of the trans-spliced RNAs use SL1 non-coding RNA (80-90% of all trans-spliced RNAs), and the SL2 non-coding RNA is mainly used by transcripts in operons (7% of all trans-spliced RNAs) (31)(32)(33). The 5′SS on the SL1 RNA is an invariant AG//GUAAA, and most of the C. elegans 3′ trans-splice sites have the same UUUCAG/R sequence found in cis-spliced 3′SSs.…”

Section: Introductionmentioning

confidence: 99%

U6 snRNA m6A modification is required for accurate and efficient cis- and trans-splicing ofC. elegansmRNAs

Shen,

Hencel,

Parker

et al. 2023

Preprint

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pre-mRNA splicing is a critical feature of eukaryotic gene expression. Many eukaryotes use cis-splicing to remove intronic sequences from pre-mRNAs. In addition to cis-splicing, many organisms use trans-splicing to replace the 5′ ends of mRNAs with a non-coding spliced-leader RNA. Both cis- and trans-splicing rely on accurately recognising splice site sequences by spliceosomal U snRNAs and associated proteins. Spliceosomal snRNAs carry multiple RNA modifications with the potential to affect different stages of pre-mRNA splicing. Here, we show that m6A modification of U6 snRNA A43 by the RNA methyltransferase METT-10 is required for accurate and efficient cis- and trans-splicing of C. elegans pre-mRNAs. The absence of U6 snRNA m6A modification primarily leads to alternative splicing at 5′ splice sites. Furthermore, weaker 5′ splice site recognition by the unmodified U6 snRNA A43 affects splicing at 3′ splice sites. U6 snRNA m6A43 and the splicing factor SNRNP27K function to recognise an overlapping set of 5′ splice sites with an adenosine at +4 position. Finally, we show that U6 snRNA m6A43 is required for efficient SL trans-splicing at weak 3′ trans-splice sites. We conclude that the U6 snRNA m6A modification is important for accurate and efficient cis- and trans-splicing in C. elegans.

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“…Despite the enigmatic nature of the precise mechanism of SLTS, it has been demonstrated that the process impacts translation efficiency 47 – 49 . For instance, a study on A. lumbricoides showed that the SL sequence and its hypermethylated cap enhance translational efficiency, especially at the initiation of protein synthesis 38 .…”

Section: Discussionmentioning

confidence: 99%

Molecular insights and antibody response to Dr20/22 in dogs naturally infected with Dirofilaria repens

Pękacz,

Basałaj,

Młocicki

et al. 2024

Sci Rep

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Subcutaneous dirofilariasis, caused by the parasitic nematode Dirofilaria repens, is a growing concern in Europe, affecting both dogs and humans. This study focused on D. repens Dr20/22, a protein encoded by an alt (abundant larval transcript) gene family. While well-documented in L3 larvae of other filariae species, this gene family had not been explored in dirofilariasis. The research involved cloning Dr20/22 cDNA, molecular characterization, and evaluating its potential application in the diagnosis of dirofilariasis. Although Real-Time analysis revealed mRNA expression in both adult worms and microfilariae, the native protein remained undetected in lysates from both developmental stages. This suggests the protein’s specificity for L3 larvae and may be related to a process called SLTS (spliced leader trans-splicing), contributing to stage-specific gene expression. The specificity of the antigen for invasive larvae positions it as a promising early marker for dirofilariasis. However, ELISA tests using sera from infected and uninfected dogs indicated limited diagnostic utility. While further research is required, our findings contribute to a deeper understanding of the molecular and immunological aspects of host-parasite interactions and could offer insights into the parasite's strategies for evading the immune system.

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Quantitative analysis of C. elegans transcripts by Nanopore direct-cDNA sequencing reveals terminal hairpins in non trans-spliced mRNAs

Cited by 10 publications

References 30 publications

A nematode-specific ribonucleoprotein complex mediates interactions between the major nematode spliced leader snRNP and its target pre-mRNAs

A nematode-specific ribonucleoprotein complex mediates interactions between the major nematode spliced leader snRNP and its target pre-mRNAs

U6 snRNA m6A modification is required for accurate and efficient cis- and trans-splicing ofC. elegansmRNAs

Molecular insights and antibody response to Dr20/22 in dogs naturally infected with Dirofilaria repens

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