PIWI‐interacting RNAs (piRNAs) are genome‐encoded small RNAs that regulate germ cell development and maintain germline integrity in many animals. Mature piRNAs engage Piwi Argonaute proteins to ...silence complementary transcripts, including transposable elements and endogenous genes. piRNA biogenesis mechanisms are diverse and remain poorly understood. Here, we identify the RNA polymerase II (RNA Pol II) core subunit RPB‐9 as required for piRNA‐mediated silencing in the nematode Caenorhabditis elegans. We show that rpb‐9 initiates heritable piRNA‐mediated gene silencing at two DNA transposon families and at a subset of somatic genes in the germline. We provide genetic and biochemical evidence that RPB‐9 is required for piRNA biogenesis by recruiting the Integrator complex at piRNA genes, hence promoting transcriptional termination. We conclude that, as a part of its rapid evolution, the piRNA pathway has co‐opted an ancient machinery for high‐fidelity transcription.
Synopsis
The piRNA pathway regulates germ cell development and promotes fertility in animals. The finding that RNA Polymerase II subunit RPB‐9 recruits the Integrator complex to terminate transcription at piRNA genes in C. elegans provides new insight into the mechanisms of piRNA biogenesis.
rpb‐9 is required for the generation of mature piRNAs and the initiation of piRNA‐mediated silencing.
rpb‐9 is required to repress two DNA transposon families and a subset of somatic genes in the germline.
RPB‐9 recruits the Integrator complex at motif‐dependent piRNA loci to terminate transcription of piRNA precursors.
piRNA biogenesis requires RBP‐9 mediated transcriptional termination at complex piRNA genes.
Methylation of carbon‐5 of cytosines (m5C) is a post‐transcriptional nucleotide modification of RNA found in all kingdoms of life. While individual m5C‐methyltransferases have been studied, the ...impact of the global cytosine‐5 methylome on development, homeostasis and stress remains unknown. Here, using Caenorhabditis elegans, we generated the first organism devoid of m5C in RNA, demonstrating that this modification is non‐essential. Using this genetic tool, we determine the localisation and enzymatic specificity of m5C sites in the RNome in vivo. We find that NSUN‐4 acts as a dual rRNA and tRNA methyltransferase in C. elegans mitochondria. In agreement with leucine and proline being the most frequently methylated tRNA isoacceptors, loss of m5C impacts the decoding of some triplets of these two amino acids, leading to reduced translation efficiency. Upon heat stress, m5C loss leads to ribosome stalling at UUG triplets, the only codon translated by an m5C34‐modified tRNA. This leads to reduced translation efficiency of UUG‐rich transcripts and impaired fertility, suggesting a role of m5C tRNA wobble methylation in the adaptation to higher temperatures.
SYNOPSIS
Cytosines carbon‐5 methylation (m5C) is a conserved RNA modification affected in various diseases, yet its functions and molecular interactions are not well characterized. A mutant C. elegans strain devoid of m5C RNA modification reveals its contribution to physiological translation and importance for translational adaptation to heat stress.
m5C and its derivatives are non‐essential RNA modifications in Caenorhabditis elegans.
m5C is reliably detected in tRNA, rRNA, and some small non‐coding RNAs, but not in coding transcripts in the nematode.
NSUN‐4 acts as a dual rRNA and tRNA methyltransferase in C. elegans mitochondria.
m5C controls translational efficiency of leucine and proline codons.
m5C loss reduces C. elegans fertility and Leu‐UUG decoding at elevated temperatures.
Analysis of worm mutants devoid of m5C RNA modifications uncovers their contribution to efficient Leu/Pro codon translation, and requirement for fertility‐supporting Leu‐UUG decoding at higher temperatures.
Chagas disease is caused by the parasite Trypanosoma cruzi, and it begins with a short acute phase characterized by high parasitemia followed by a life-long chronic phase with scarce parasitism. ...Cardiac involvement is the most prominent manifestation, as 30% of infected subjects will develop abnormal ventricular repolarization with myocarditis, fibrosis and cardiomyocyte hypertrophy by undefined mechanisms. Nevertheless, follow-up studies in chagasic patients, as well as studies with murine models, suggest that the intensity of clinical symptoms and pathophysiological events that occur during the acute phase of disease are associated with the severity of cardiac disease observed during the chronic phase. In the present study we investigated the role of microRNAs (miRNAs) in the disease progression in response to T. cruzi infection, as alterations in miRNA levels are known to be associated with many cardiovascular disorders. We screened 641 rodent miRNAs in heart samples of mice during an acute infection with the Colombiana T.cruzi strain and identified multiple miRNAs significantly altered upon infection. Seventeen miRNAs were found significantly deregulated in all three analyzed time points post infection. Among these, six miRNAs had their expression correlated with clinical parameters relevant to the disease, such as parasitemia and maximal heart rate-corrected QT (QTc) interval. Computational analyses identified that the gene targets for these six miRNAs were involved in networks and signaling pathways related to increased ventricular depolarization and repolarization times, important factors for QTc interval prolongation. The data presented here will guide further studies about the contribution of microRNAs to Chagas heart disease pathogenesis.
RNA flow between organisms has been documented within and among different kingdoms of life. Recently, we demonstrated horizontal RNA transfer between honeybees involving secretion and ingestion of ...worker and royal jellies. However, how the jelly facilitates transfer of RNA is still unknown. Here, we show that worker and royal jellies harbor robust RNA-binding activity. We report that a highly abundant jelly component, major royal jelly protein 3 (MRJP-3), acts as an extracellular non-sequence-specific RNA-aggregating factor. Multivalent RNA binding stimulates higher-order assembly of MRJP-3 into extracellular ribonucleoprotein granules that protect RNA from degradation and enhance RNA bioavailability. These findings reveal that honeybees have evolved a secreted dietary RNA-binding factor to concentrate, stabilize, and share RNA among individuals. Our work identifies high-order ribonucleoprotein assemblies with functions outside cells and organisms.
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•Worker and royal jellies harbor robust RNA-binding activity•Major royal jelly protein 3 (MRJP-3) is the dietary RNA-binding factor•RNA stimulates higher-order assembly of MRJP-3 into extracellular RNP granules•MRJP-3 granules concentrate, stabilize, and enhance environmental RNA bioavailability
Honeybees share RNA between individuals through secretion and ingestion of worker and royal jellies. Maori et al. identify a jelly factor, MRJP-3, that forms extracellular ribonucleoprotein granules that concentrate, protect, and enhance RNA uptake. They propose that MRJP-3 facilitates RNA transfer that drives social immunity and signaling among bees.
Functional characterisation of the compact genome of the model organism Caenorhabditis elegans remains incomplete despite its sequencing 20 years ago. The last decade of research has seen a ...tremendous increase in the number of non-coding RNAs identified in various organisms. While we have mechanistic understandings of small non-coding RNA pathways, long non-coding RNAs represent a diverse class of active transcripts whose function remains less well characterised.
By analysing hundreds of published transcriptome datasets, we annotated 3392 potential lncRNAs including 143 multi-exonic loci that showed increased nucleotide conservation and GC content relative to other non-coding regions. Using CRISPR/Cas9 genome editing, we generated deletion mutants for ten long non-coding RNA loci. Using automated microscopy for in-depth phenotyping, we show that six of the long non-coding RNA loci are required for normal development and fertility. Using RNA interference-mediated gene knock-down, we provide evidence that for two of the long non-coding RNA loci, the observed phenotypes are dependent on the corresponding RNA transcripts.
Our results highlight that a large section of the non-coding regions of the C. elegans genome remains unexplored. Based on our in vivo analysis of a selection of high-confidence lncRNA loci, we expect that a significant proportion of these high-confidence regions is likely to have a biological function at either the genomic or the transcript level.
Chagas disease, caused by the parasite Trypanosoma cruzi, is endemic in Latin America. Its acute phase is associated with high parasitism, myocarditis and profound myocardial gene expression changes. ...A chronic phase ensues where 30% develop severe heart lesions. Mouse models of T. cruzi infection have been used to study heart damage in Chagas disease. The aim of this study was to provide an interactome between miRNAs and their targetome in Chagas heart disease by integrating gene and microRNA expression profiling data from hearts of T. cruzi infected mice. Gene expression profiling revealed enrichment in biological processes and pathways associated with immune response and metabolism. Pathways, functional and upstream regulator analysis of the intersections between predicted targets of differentially expressed microRNAs and differentially expressed mRNAs revealed enrichment in biological processes and pathways such as IFNγ, TNFα, NF-kB signaling signatures, CTL-mediated apoptosis, mitochondrial dysfunction, and Nrf2-modulated antioxidative responses. We also observed enrichment in other key heart disease-related processes like myocarditis, fibrosis, hypertrophy and arrhythmia. Our correlation study suggests that miRNAs may be implicated in the pathophysiological processes taking place the hearts of acutely T. cruzi-infected mice.
Long noncoding RNAs (lncRNAs) modulate gene expression at the epigenetic, transcriptional, and posttranscriptional levels. Dysregulation of the lncRNA known as myocardial infarction–associated ...transcript (MIAT) has been associated with myocardial infarction. Chagas disease causes a severe inflammatory dilated chronic cardiomyopathy (CCC). We investigated the role of MIAT in CCC. A whole-transcriptome analysis of heart biopsy specimens and formalin-fixed, paraffin-embedded samples revealed that MIAT was overexpressed in patients with CCC, compared with subjects with noninflammatory dilated cardiomyopathy and controls. These results were confirmed in a mouse model. Results suggest that MIAT is a specific biomarker of CCC.
Chagas disease, caused by the protozoan parasite Trypanosoma cruzi (T. cruzi), is characterized by immunopathology driven by IFN-γ secreting Th1-like T cells. T. cruzi has a thick coat of mucin-like ...glycoproteins covering its surface, which plays an important role in parasite invasion and host immunomodulation. It has been extensively described that T. cruzi or its products—like GPI anchors isolated from GPI-anchored mucins from the trypomastigote life cycle stage (tGPI-mucins)—are potent inducers of proinflammatory responses (i.e., cytokines and NO production) by IFN-γ primed murine macrophages. However, little is known about whether T. cruzi or GPI-mucins exert a similar action in human cells. We therefore decided to further investigate the in vitro cytokine production profile from human mononuclear cells from uninfected donors exposed to T. cruzi as well as tGPI-mucins. We observed that both living T. cruzi trypomastigotes and tGPI-mucins are potent inducers of IL-12 by human peripheral blood monocytes and this effect depends on CD40-CD40L interaction and IFN-γ. Our findings suggest that the polarized T1-type cytokine profile seen in T. cruzi infected patients might be a long-term effect of IL-12 production induced by lifelong exposure to T. cruzi tGPI-mucins.
Chagas disease cardiomyopathy(CCC), the main consequence of Trypanosoma cruzi(T.cruzi) infection, is an inflammatory cardiomyopathy that develops in up to 30% of infected individuals. The heart ...inflammation in CCC patients is characterized by a Th1 T cell-rich myocarditis with increased production of interferon(IFN)-γ, produced by the CCC myocardial infiltrate and detected at high levels in the periphery. IFN-γ has a central role in the cardiomyocyte signaling during both acute and chronic phases of T.cruzi infection. In this review, we have chosen to focus in its pleiotropic mode of action during CCC, which may ultimately be the strongest driver towards pathological remodeling and heart failure. We describe here the antiparasitic protective and pathogenic dual role of IFN-γ in Chagas disease.
Abstract Background/methods Chagas disease is caused by an intracellular parasite, Trypanosoma cruzi , and it is a leading cause of heart failure in Latin America. The main clinical consequence of ...the infection is the development of a Chronic Chagas disease Cardiomyopathy (CCC), which is characterized by myocarditis, hypertrophy and fibrosis and affects about 30% of infected patients. CCC has a worse prognosis than other cardiomyopathies, like idiopathic dilated cardiomyopathy (DCM). It is well established that myocardial gene expression patterns are altered in CCC, but the molecular mechanisms underlying these differences are not clear. MicroRNAs are recently discovered regulators of gene expression, and are recognized as important factors in heart development and cardiovascular disorders (CD). We analyzed the expression of nine different miRNAs in myocardial tissue samples of CCC patients in comparison to DCM patients and samples from heart transplant donors. Using the results of a cDNA microarray database on CCC and DCM myocardium, signaling networks were built and nodal molecules were identified. Results We observed that five miRNAs were significantly altered in CCC and three in DCM; importantly, three miRNAs were significantly reduced in CCC as compared to DCM. We observed that multiple gene targets of the differentially expressed miRNAs showed a concordant inverse expression in CCC. Significantly, most gene targets and involved networks belong to crucial disease-related signaling pathways. Conclusion These results suggest that miRNAs may play a major role in the regulation of gene expression in CCC pathogenesis, with potential implication as diagnostic and prognostic tools.
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