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

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.

Both trust in, and polarization of trust in, relevant sciences have increased through the COVID-19 pandemic.

While attempts to promote acceptance of well-evidenced science have historically focused on increasing scientific knowledge, it is now thought that for acceptance of science, trust in, rather than simply knowledge of, science is foundational. Here we employ the COVID-19 pandemic as a natural experiment on trust modulation as it has enabled unprecedented exposure of science. We ask whether trust in science has on the average altered, whether trust has changed the same way for all and, if people have responded differently, what predicts these differences? We 1) categorize the nature of self-reported change in trust in "scientists" in a random sample of over 2000 UK adults after the introduction of the first COVID vaccines, 2) ask whether any reported change is likely to be real through consideration of both a negative control and through experiment, and 3) address what predicts change in trust considering sex, educational attainment, religiosity, political attitude, age and pre-pandemic reported trust. We find that many more (33%) report increased trust towards "scientists" than report decreased trust (7%), effects of this magnitude not being seen in negative controls. Only age and prior degree of trust predict change in trust, the older population increasing trust more. The prior degree of trust effect is such that those who say they did not trust science prior to the pandemic are more likely to report becoming less trusting, indicative of both trust polarization and a backfire effect. Since change in trust is predictive of willingness to have a COVID-19 vaccine, it is likely that these changes have public health consequences.

People with more extreme attitudes towards science have self-confidence in their understanding of science, even if this is not justified.

People differ greatly in their attitudes towards well-evidenced science. What characterises this variation? Here, we consider this issue in the context of genetics and allied sciences. While most prior research has focused on the relationship between attitude to science and what people know about it, recent evidence suggests that individuals with strongly negative attitudes towards specific genetic technologies (genetic modification (GM) technology and vaccines) commonly do not objectively understand the science, but, importantly, believe that they do. Here, using data from a probability survey of United Kingdom adults, we extend this prior work in 2 regards. First, we ask whether people with more extreme attitudes, be they positive or negative, are more likely to believe that they understand the science. Second, as negativity to genetics is commonly framed around issues particular to specific technologies, we ask whether attitudinal trends are contingent on specification of technology. We find (1) that individuals with strongly positive or negative attitudes towards genetics more strongly believe that they well understand the science; but (2) only for those most positive to the science is this self-confidence warranted; and (3) these effects are not contingent on specification of any particular technologies. These results suggest a potentially general model to explain why people differ in their degree of acceptance or rejection of science, this being that the more someone believes they understand the science, the more confident they will be in their acceptance or rejection of it. While there are more technology nonspecific opponents who also oppose GM technology than expected by chance, most GM opponents fit a different demographic. For the most part, opposition to GM appears not to reflect a smokescreen concealing a broader underlying negativity.

TES-1/Tes and ZYX-1/Zyxin protect junctional actin networks under tension during epidermal morphogenesis in the C. elegans embryo.

LIM-domain-containing repeat (LCR) proteins are recruited to strained actin filaments within stress fibers in cultured cells,1,2,3 but their roles at cell-cell junctions in living organisms have not been extensively studied. Here, we show that the Caenorhabditis elegans LCR proteins TES-1/Tes and ZYX-1/Zyxin are recruited to apical junctions during embryonic elongation when junctions are under tension. In genetic backgrounds in which embryonic elongation fails, junctional recruitment is severely compromised. The two proteins display complementary patterns of expression: TES-1 is expressed in lateral (seam) epidermal cells, whereas ZYX-1 is expressed in dorsal and ventral epidermal cells. tes-1 and zyx-1 mutant embryos display junctional F-actin defects. The loss of either protein strongly enhances morphogenetic defects in hypomorphic mutant backgrounds for cadherin/catenin complex (CCC) components. The LCR regions of TES-1 and ZYX-1 are recruited to stress fiber strain sites (SFSSs) in cultured vertebrate cells. Together, these data establish TES-1 and ZYX-1 as components of a multicellular, tension-sensitive system that stabilizes the junctional actin cytoskeleton during embryonic morphogenesis.

CMTr mediated 2'-O-ribose methylation status of cap-adjacent nucleotides across animals.

Cap methyltransferases (CMTrs) O methylate the 2' position of the ribose (cOMe) of cap-adjacent nucleotides of animal, protist, and viral mRNAs. Animals generally have two CMTrs, whereas trypanosomes have three, and many viruses encode one in their genome. In the splice leader of mRNAs in trypanosomes, the first four nucleotides contain cOMe, but little is known about the status of cOMe in animals. Here, we show that cOMe is prominently present on the first two cap-adjacent nucleotides with species- and tissue-specific variations in Caenorhabditis elegans, honeybees, zebrafish, mouse, and human cell lines. In contrast, Drosophila contains cOMe primarily on the first cap-adjacent nucleotide. De novo RoseTTA modeling of CMTrs reveals close similarities of the overall structure and near identity for the catalytic tetrad, and for cap and cofactor binding for human, Drosophila and C. elegans CMTrs. Although viral CMTrs maintain the overall structure and catalytic tetrad, they have diverged in cap and cofactor binding. Consistent with the structural similarity, both CMTrs from Drosophila and humans methylate the first cap-adjacent nucleotide of an AGU consensus start. Because the second nucleotide is also methylated upon heat stress in Drosophila, these findings argue for regulated cOMe important for gene expression regulation.

A novel, essential trans-splicing protein connects the nematode SL1 snRNP to the CBC-ARS2 complex.

Spliced leader trans-splicing is essential for gene expression in many eukaryotes. To elucidate the molecular mechanism of this process, we characterise the molecules associated with the Caenorhabditis elegans major spliced leader snRNP (SL1 snRNP), which donates the spliced leader that replaces the 5' untranslated region of most pre-mRNAs. Using a GFP-tagged version of the SL1 snRNP protein SNA-1 created by CRISPR-mediated genome engineering, we immunoprecipitate and identify RNAs and protein components by RIP-Seq and mass spectrometry. This reveals the composition of the SL1 snRNP and identifies associations with spliceosome components PRP-8 and PRP-19. Significantly, we identify a novel, nematode-specific protein required for SL1 trans-splicing, which we designate SNA-3. SNA-3 is an essential, nuclear protein with three NADAR domains whose function is unknown. Mutation of key residues in NADAR domains inactivates the protein, indicating that domain function is required for activity. SNA-3 interacts with the CBC-ARS2 complex and other factors involved in RNA metabolism, including SUT-1 protein, through RNA or protein-mediated contacts revealed by yeast two-hybrid assays, localisation studies and immunoprecipitations. Our data are compatible with a role for SNA-3 in coordinating trans-splicing with target pre-mRNA transcription or in the processing of the Y-branch product of the trans-splicing reaction.

SLIDR and SLOPPR: flexible identification of spliced leader trans-splicing and prediction of eukaryotic operons from RNA-Seq data.

BACKGROUND: Spliced leader (SL) trans-splicing replaces the 5' end of pre-mRNAs with the spliced leader, an exon derived from a specialised non-coding RNA originating from elsewhere in the genome. This process is essential for resolving polycistronic pre-mRNAs produced by eukaryotic operons into monocistronic transcripts. SL trans-splicing and operons may have independently evolved multiple times throughout Eukarya, yet our understanding of these phenomena is limited to only a few well-characterised organisms, most notably C. elegans and trypanosomes. The primary barrier to systematic discovery and characterisation of SL trans-splicing and operons is the lack of computational tools for exploiting the surge of transcriptomic and genomic resources for a wide range of eukaryotes. RESULTS: Here we present two novel pipelines that automate the discovery of SLs and the prediction of operons in eukaryotic genomes from RNA-Seq data. SLIDR assembles putative SLs from 5' read tails present after read alignment to a reference genome or transcriptome, which are then verified by interrogating corresponding SL RNA genes for sequence motifs expected in bona fide SL RNA molecules. SLOPPR identifies RNA-Seq reads that contain a given 5' SL sequence, quantifies genome-wide SL trans-splicing events and predicts operons via distinct patterns of SL trans-splicing events across adjacent genes. We tested both pipelines with organisms known to carry out SL trans-splicing and organise their genes into operons, and demonstrate that (1) SLIDR correctly detects expected SLs and often discovers novel SL variants; (2) SLOPPR correctly identifies functionally specialised SLs, correctly predicts known operons and detects plausible novel operons. CONCLUSIONS: SLIDR and SLOPPR are flexible tools that will accelerate research into the evolutionary dynamics of SL trans-splicing and operons throughout Eukarya and improve gene discovery and annotation for a wide range of eukaryotic genomes. Both pipelines are implemented in Bash and R and are built upon readily available software commonly installed on most bioinformatics servers. Biological insight can be gleaned even from sparse, low-coverage datasets, implying that an untapped wealth of information can be retrieved from existing RNA-Seq datasets as well as from novel full-isoform sequencing protocols as they become more widely available.

Resolution of polycistronic RNA by SL2 trans-splicing is a widely conserved nematode trait.

Spliced leader trans-splicing is essential for the processing and translation of polycistronic RNAs generated by eukaryotic operons. In C. elegans, a specialized spliced leader, SL2, provides the 5' end for uncapped pre-mRNAs derived from polycistronic RNAs. Studies of other nematodes suggested that SL2-type trans-splicing is a relatively recent innovation, confined to Rhabditina, the clade containing C. elegans and its close relatives. Here we conduct a survey of transcriptome-wide spliced leader trans-splicing in Trichinella spiralis, a distant relative of C. elegans with a particularly diverse repertoire of 15 spliced leaders. By systematically comparing the genomic context of trans-splicing events for each spliced leader, we identified a subset of T. spiralis spliced leaders that are specifically used to process polycistronic RNAs-the first examples of SL2-type spliced leaders outside of Rhabditina. These T. spiralis spliced leader RNAs possess a perfectly conserved stem-loop motif previously shown to be essential for SL2-type trans-splicing in C. elegans We show that genes trans-spliced to these SL2-type spliced leaders are organized in operonic fashion, with short intercistronic distances. A subset of T. spiralis operons show conservation of synteny with C. elegans operons. Our work substantially revises our understanding of nematode spliced leader trans-splicing, showing that SL2 trans-splicing is a major mechanism for nematode polycistronic RNA processing, which may have evolved prior to the radiation of the Nematoda. This work has important implications for the improvement of genome annotation pipelines in nematodes and other eukaryotes with operonic gene organization.

A high-throughput screen for the identification of compounds that inhibit nematode gene expression by targeting spliced leader trans-splicing.

Infections with parasitic nematodes are among the most significant of the neglected tropical diseases affecting about a billion people living mainly in tropical regions with low economic activity. The most effective current strategy to control nematode infections involves large scale treatment programs with anthelmintic drugs. This strategy is at risk from the emergence of drug resistant parasites. Parasitic nematodes also affect livestock, which are treated with the same limited group of anthelmintic drugs. Livestock parasites resistant to single drugs, and even multi-drug resistant parasites, are appearing in many areas. There is therefore a pressing need for new anthelmintic drugs. Here we use the nematode Caenorhabditis elegans as a model for parasitic nematodes and demonstrate that sinefungin, a competitive inhibitor of methyltransferases, causes a delay in development and reduced fecundity, and inhibits spliced leader trans-splicing. Spliced leader trans-splicing is an essential step in gene expression that does not occur in the hosts of parasitic nematodes, and is therefore a potential target for new anthelmintic drugs. We have exploited the ability of sinefungin to inhibit spliced leader trans-splicing to adapt a green fluorescent protein based reporter gene assay that monitors spliced leader trans-splicing for high-throughput screening for new anthelmintic compounds. We have established a protocol for robust high-throughput screening, combining mechanical dispensing of living C. elegans into 384- or 1536- well plates with addition of compounds using an acoustic liquid dispenser, and the detection of the inhibition of SL trans-splicing using a microplate reader. We have tested this protocol in a first pilot screen and envisage that this assay will be a valuable tool in the search for new anthelmintic drugs.

An in vivo genetic screen for genes involved in spliced leader trans-splicing indicates a crucial role for continuous de novo spliced leader RNP assembly.

Spliced leader (SL) trans-splicing is a critical element of gene expression in a number of eukaryotic groups. This process is arguably best understood in nematodes, where biochemical and molecular studies in Caenorhabditis elegans and Ascaris suum have identified key steps and factors involved. Despite this, the precise details of SL trans-splicing have yet to be elucidated. In part, this is because the systematic identification of the molecules involved has not previously been possible due to the lack of a specific phenotype associated with defects in this process. We present here a novel GFP-based reporter assay that can monitor SL1 trans-splicing in living C. elegans. Using this assay, we have identified mutants in sna-1 that are defective in SL trans-splicing, and demonstrate that reducing function of SNA-1, SNA-2 and SUT-1, proteins that associate with SL1 RNA and related SmY RNAs, impairs SL trans-splicing. We further demonstrate that the Sm proteins and pICln, SMN and Gemin5, which are involved in small nuclear ribonucleoprotein assembly, have an important role in SL trans-splicing. Taken together these results provide the first in vivo evidence for proteins involved in SL trans-splicing, and indicate that continuous replacement of SL ribonucleoproteins consumed during trans-splicing reactions is essential for effective trans-splicing.

Operons are a conserved feature of nematode genomes.

The organization of genes into operons, clusters of genes that are co-transcribed to produce polycistronic pre-mRNAs, is a trait found in a wide range of eukaryotic groups, including multiple animal phyla. Operons are present in the class Chromadorea, one of the two main nematode classes, but their distribution in the other class, the Enoplea, is not known. We have surveyed the genomes of Trichinella spiralis, Trichuris muris, and Romanomermis culicivorax and identified the first putative operons in members of the Enoplea. Consistent with the mechanism of polycistronic RNA resolution in other nematodes, the mRNAs produced by genes downstream of the first gene in the T. spiralis and T. muris operons are trans-spliced to spliced leader RNAs, and we are able to detect polycistronic RNAs derived from these operons. Importantly, a putative intercistronic region from one of these potential enoplean operons confers polycistronic processing activity when expressed as part of a chimeric operon in Caenorhabditis elegans. We find that T. spiralis genes located in operons have an increased likelihood of having operonic C. elegans homologs. However, operon structure in terms of synteny and gene content is not tightly conserved between the two taxa, consistent with models of operon evolution. We have nevertheless identified putative operons conserved between Enoplea and Chromadorea. Our data suggest that operons and "spliced leader" (SL) trans-splicing predate the radiation of the nematode phylum, an inference which is supported by the phylogenetic profile of proteins known to be involved in nematode SL trans-splicing.

Cadherins and their partners in the nematode worm Caenorhabditis elegans.

The extreme simplicity of Caenorhabditis elegans makes it an ideal system to study the basic principles of cadherin function at the level of single cells within the physiologically relevant context of a developing animal. The genetic tractability of C. elegans also means that components of cadherin complexes can be identified through genetic modifier screens, allowing a comprehensive in vivo characterization of the macromolecular assemblies involved in cadherin function during tissue formation and maintenance in C. elegans. This work shows that a single cadherin system, the classical cadherin-catenin complex, is essential for diverse morphogenetic events during embryogenesis through its interactions with a range of mostly conserved proteins that act to modulate its function. The role of other members of the cadherin family in C. elegans, including members of the Fat-like, Flamingo/CELSR and calsyntenin families is less well characterized, but they have clear roles in neuronal development and function.

Specific conserved C-terminal amino acids of Caenorhabditis elegans HMP-1/α-catenin modulate F-actin binding independently of vinculin.

Stable intercellular adhesions formed through the cadherin-catenin complex are important determinants of proper tissue architecture and help maintain tissue integrity during morphogenetic movements in developing embryos. A key regulator of this stability is α-catenin, which connects the cadherin-catenin complex to the actin cytoskeleton. Although the C-terminal F-actin-binding domain of α-catenin has been shown to be crucial for its function, a more detailed in vivo analysis of discrete regions and residues required for actin binding has not been performed. Using Caenorhabditis elegans as a model system, we have characterized mutations in hmp-1/α-catenin that identify HMP-1 residues 687-742 and 826-927, as well as amino acid 802, as critical to the localization of junctional proximal actin during epidermal morphogenesis. We also find that the S823F transition in a hypomorphic allele, hmp-1(fe4), decreases actin binding in vitro. Using hmp-1(fe4) animals in a mutagenesis screen, we were then able to identify 11 intragenic suppressors of hmp-1(fe4) that revert actin binding to wild-type levels. Using homology modeling, we show that these amino acids are positioned at key conserved sites within predicted α-helices in the C terminus. Through the use of transgenic animals, we also demonstrate that HMP-1 residues 315-494, which correspond to a putative mechanotransduction domain that binds vinculin in vertebrate αE-catenin, are not required during epidermal morphogenesis but may aid efficient recruitment of HMP-1 to the junction. Our studies are the first to identify key conserved amino acids in the C terminus of α-catenin that modulate F-actin binding in living embryos of a simple metazoan.

A genome-wide functional screen shows MAGI-1 is an L1CAM-dependent stabilizer of apical junctions in C. elegans.

BACKGROUND: In multicellular organisms, cell-cell junctions are involved in many aspects of tissue morphogenesis. α-catenin links the cadherin-catenin complex (CCC) to the actin cytoskeleton, stabilizing cadherin-dependent adhesions. RESULTS: To identify modulators of cadherin-based cell adhesion, we conducted a genome-wide RNAi screen in C. elegans and uncovered MAGI-1, a highly conserved protein scaffold. Loss of magi-1 function in wild-type embryos results in disorganized epithelial migration and occasional morphogenetic failure. MAGI-1 physically interacts with the putative actin regulator AFD-1/afadin; knocking down magi-1 or afd-1 function in a hypomorphic α-catenin background leads to complete morphogenetic failure and actin disorganization in the embryonic epidermis. MAGI-1 and AFD-1 localize to a unique domain in the apical junction and normal accumulation of MAGI-1 at junctions requires SAX-7/L1CAM, which can bind MAGI-1 via its C terminus. Depletion of MAGI-1 leads to loss of spatial segregation and expansion of apical junctional domains and greater mobility of junctional proteins. CONCLUSIONS: Our screen is the first genome-wide approach to identify proteins that function synergistically with the CCC during epidermal morphogenesis in a living embryo. We demonstrate novel physical interactions between MAGI-1, AFD-1/afadin, and SAX-7/L1CAM, which are part of a functional interactome that includes components of the core CCC. Our results further suggest that MAGI-1 helps to partition and maintain a stable, spatially ordered apical junction during morphogenesis.

The evolution of spliced leader trans-splicing in nematodes.

Spliced leader trans-splicing occurs in many primitive eukaryotes including nematodes. Most of our knowledge of trans-splicing in nematodes stems from the model organism Caenorhabditis elegans and relatives, and from work with Ascaris. Our investigation of spliced leader trans-splicing in distantly related Dorylaimia nematodes indicates that spliced-leader trans-splicing arose before the nematode phylum and suggests that the spliced leader RNA gene complements in extant nematodes have evolved from a common ancestor with a diverse set of spliced leader RNA genes.

SL2-like spliced leader RNAs in the basal nematode Prionchulus punctatus: New insight into the evolution of nematode SL2 RNAs.

Spliced-leader (SL) trans-splicing has been found in all molecularly characterized nematode species to date, and it is likely to be a nematode synapomorphy. Most information regarding SL trans-splicing has come from the study of nematodes from a single monophyletic group, the Rhabditida, all of which employ SL RNAs that are identical to, or variants of, the SL1 RNA first characterized in Caenorhabditis elegans. In contrast, the more distantly related Trichinella spiralis, belonging to the subclass Dorylaimia, utilizes a distinct set of SL RNAs that display considerable sequence diversity. To investigate whether this is true of other members of the Dorylaimia, we have characterized SL RNAs from Prionchulus punctatus. Surprisingly, this revealed the presence of a set of SLs that show clear sequence similarity to the SL2 family of spliced leaders, which have previously only been found within the rhabditine group (which includes C. elegans). Expression of one of the P. punctatus SL RNAs in C. elegans reveals that it can compete specifically with the endogenous C. elegans SL2 spliced leaders, being spliced to the pre-mRNAs derived from downstream genes in operons, but does not compete with the SL1 spliced leaders. This discovery raises the possibility that SL2-like spliced leaders were present in the last common ancestor of the nematode phylum.

Rapid sublethal toxicity assessment using bioluminescent Caenorhabditis elegans, a novel whole-animal metabolic biosensor.

Sublethal metabolic effects are informative toxicological end points. We used a rapid quantitative metabolic end point, bioluminescence of firefly luciferase expressing Caenorhabditis elegans, to assess effects of sublethal chronic exposure (19 h) to the oxidative stress agent and environmental pollutant cadmium (provided as chloride salt). Bioluminescence declined in a concentration-dependent manner in the concentration range tested (0-30 microM Cd), with comparable sensitivity to reproduction and developmental assay end points (after 67 and 72 h, respectively). Cd concentrations that resulted in 20% reduction in bioluminescence (EC(20)) were 11.8-13.0 microM, whereas the reproduction EC(20) (67 h exposure) was 10.2 microM. At low concentrations of Cd (< or = 15 microM), the decline in bioluminescence reflected a drop in ATP levels. At Cd concentrations of 15-30 microM, decreased bioluminescence was attributable both to effects of Cd on ATP levels and decreased production of luciferase proteins, concomitant with a decline in protein levels. We show that whole-animal bioluminescence is a valid toxicological end point and a rapid and sensitive predictor of effects of Cd exposure on development and reproduction. This provides a platform for high-throughput sublethal screening and will potentially contribute to reduction of testing in higher animals.

Bridging the phenotypic gap: real-time assessment of mitochondrial function and metabolism of the nematode Caenorhabditis elegans.

BACKGROUND: The ATP levels of an organism are an important physiological parameter that is affected by genetic make up, ageing, stress and disease. RESULTS: We have generated luminescent C. elegans through ubiquitous, constitutive expression of firefly luciferase, widely used for in vitro ATP determination. We hypothesise that whole animal luminescence reflects its intracellular ATP levels in vivo. To test this, we characterised the bioluminescence response of C. elegans during sublethal exposure to, and recovery from azide, a treatment that inhibits mitochondrial respiration reversibly, and causes ATP depletion. Consistent with our expectations, in vivo luminescence decreased with increasing sublethal azide levels, and recovered fully when worms were removed from azide. Firefly luciferase expression levels, stability and activity did not influence the final luminescence. Bioluminescence also reflected the lowered activity of the electron transport chain achieved with RNA interference (RNAi) of genes encoding respiratory chain components. CONCLUSION: Results indicated that C. elegans luminescence reports on ATP levels in real-time. For the first time, we are able to directly assess the metabolism of a whole, living, multicellular organism by determination of the relative ATP levels. This will enable genetic analysis based on a readily quantifiable metabolic phenotype and will provide novel insights into mechanisms of fitness and disease that are likely to be of relevance for other organisms, as well as the worm.

Spliced leader trans-splicing in the nematode Trichinella spiralis uses highly polymorphic, noncanonical spliced leaders.

The trans-splicing of short spliced leader (SL) RNAs onto the 5' ends of mRNAs occurs in a diverse range of taxa. In nematodes, all species so far characterized utilize a characteristic, conserved spliced leader, SL1, as well as variants that are employed in the resolution of operons. Here we report the identification of spliced leader trans-splicing in the basal nematode Trichinella spiralis, and show that this nematode does not possess a canonical SL1, but rather has at least 15 distinct spliced leaders, encoded by at least 19 SL RNA genes. The individual spliced leaders vary in both size and primary sequence, showing a much higher degree of diversity compared to other known trans-spliced leaders. In a survey of T. spiralis mRNAs, individual mRNAs were found to be trans-spliced to a number of different spliced leader sequences. These data provide the first indication that the last common ancestor of the phylum Nematoda utilized spliced leader trans-splicing and that the canonical spliced leader, SL1, found in Caenorhabditis elegans, evolved after the divergence of the major nematode clades. This discovery sheds important light on the nature and evolution of mRNA processing in the Nematoda.