Probes for INS

The major cardiac cell types composing the adult heart arise from common multipotent precursor cells. Cardiac lineage decisions are guided by extrinsic and cell-autonomous factors, including recently discovered long noncoding RNAs (lncRNAs). The human lncRNA CARMEN, which is known to dictate specification towards the cardiomyocyte (CM) and the smooth muscle cell (SMC) fates, generates a diversity of alternatively spliced isoforms.The CARMEN locus can be manipulated to direct human primary cardiac precursor cells (CPCs) into specific cardiovascular fates. Investigating CARMEN isoform usage in differentiating CPCs represents therefore a unique opportunity to uncover isoform-specific function in lncRNAs. Here, we identify one CARMEN isoform, CARMEN-201, to be crucial for SMC commitment. CARMEN-201 activity is encoded within an alternatively-spliced exon containing a MIRc short interspersed nuclear element. This element binds the transcriptional repressor REST (RE1 Silencing Transcription Factor), targets it to cardiogenic loci, including ISL1, IRX1, IRX5, and SFRP1, and thereby blocks the CM gene program. In turn, genes regulating SMC differentiation are induced.These data show how a critical physiological switch is wired by alternative splicing and functional transposable elements in a long noncoding RNA. They further demonstrated the crucial importance of the lncRNA isoform CARMEN-201 in SMC specification during heart development.

Stable intronic sequence RNAs (sisRNAs) are stable, long noncoding RNAs containing intronic sequences. While sisRNAs have been found across diverse species, their level of conservation remains poorly understood. Here we report that the biogenesis and functions of a sisRNA transcribed from the highly conserved Arglu1 locus are distinct in human and Drosophila melanogaster. The Arglu1 genes in both species show similar exon-intron structures where the intron 2 is orthologous and positionally conserved. In humans, Arglu1 sisRNA retains the entire intron 2 and promotes host gene splicing. Mechanistically, Arglu1 sisRNA represses the splicing-inhibitory activity of ARGLU1 protein by binding to ARGLU1 protein and promoting its localization to nuclear speckles, away from the Arglu1 gene locus. In contrast, Drosophila dArglu1 sisRNA forms via premature cleavage of intron 2 and represses host gene splicing. This repression occurs through a local accumulation of dARGLU1 protein and inhibition of telescripting by U1 snRNPs at the dArglu1 locus. We propose that distinct biogenesis of positionally conserved Arglu1 sisRNAs in both species may have led to functional divergence.

Vascular integrity is essential for organ homeostasis to prevent edema formation and infiltration of inflammatory cells. Long non-coding RNAs (lncRNAs) are important regulators of gene expression and often expressed in a cell type-specific manner. By screening for endothelial-enriched lncRNAs, we identified the undescribed lncRNA NTRAS to control endothelial cell functions. Silencing of NTRAS induces endothelial cell dysfunction in vitro and increases vascular permeability and lethality in mice. Biochemical analysis revealed that NTRAS, through its CA-dinucleotide repeat motif, sequesters the splicing regulator hnRNPL to control alternative splicing of tight junction protein 1 (TJP1; also named zona occludens 1, ZO-1) pre-mRNA. Deletion of the hnRNPL binding motif in mice (Ntras∆CA/∆CA ) significantly repressed TJP1 exon 20 usage, favoring expression of the TJP1α- isoform, which augments permeability of the endothelial monolayer. Ntras∆CA/∆CA mice further showed reduced retinal vessel growth and increased vascular permeability and myocarditis. In summary, this study demonstrates that NTRAS is an essential gatekeeper of vascular integrity.

Metastasis is the predominant cause of mortality in patients with breast cancer. Long noncoding RNAs (lncRNAs) have been shown to drive important phenotypes in tumors, including invasion and metastasis. However, the lncRNAs involved in metastasis and their molecular and cellular mechanisms are still largely unknown.The transcriptional and posttranscriptional processing of LINC00478-associated cytoplasmic RNA (LacRNA) was determined by RT-qPCR, semiquantitative PCR and 5'/3' RACE. Paired-guide CRISPR/cas9 and CRISPR/dead-Cas9 systems was used to knock out or activate the expression of LacRNA. Cell migration and invasion assay was performed to confirm the phenotype of LacRNA. Tail vein model and mammary fat pad model were used for in vivo study. The LacRNA-PHB2-cMyc axis were screened and validated by RNA pulldown, mass spectrometry, RNA immunoprecipitation and RNA-seq assays.Here, we identified a novel cytoplasmic lncRNA, LacRNA (LINC00478-associated cytoplasmic RNA), derived from nucleus-located lncRNA LINC00478. The nascent transcript of LINC00478 full-length (LINC00478_FL) was cleaved and polyadenylated, simultaneously yielding 5' ends stable expressing LacRNA, which is released into the cytoplasm, and long 3' ends of nuclear-retained lncRNA. LINC00478_3'RNA was rapidly degraded. LacRNA significantly inhibited breast cancer invasion and metastasis in vitro and in vivo. Mechanistically, LacRNA physically interacted with the PHB domain of PHB2 through its 61-140-nt region. This specific binding affected the formation of the autophagy degradation complex of PHB2 and LC3, delaying the degradation of the PHB2 protein. Unexpectedly, LacRNA specifically interacted with PHB2, recruited c-Myc and promoted c-Myc ubiquitination and degradation. The negatively regulation of Myc signaling ultimately inhibited breast cancer metastasis. Furthermore, LacRNA and LacRNA-mediated c-Myc signaling downregulation are significantly associated with good clinical outcomes, take advantage of these factors we constructed a prognostic predict model.Therefore, our findings propose LacRNA as a potential prognostic biomarker and a new therapeutic strategy.

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As a common pathology of many ocular disorders such as diabetic retinopathy and glaucoma, retinal ischemia/reperfusion (IR) triggers inflammation and microglia activation that lead to irreversible retinal damage. The detailed molecular mechanism underlying retinal IR injury, however, remains poorly understood at present. Here we report the bioinformatic identification of a lncRNA 1810058I24Rik (181-Rik) that was shown to encode a mitochondrion-located micropeptide Stmp1. Its deficiency in mice protected retinal ganglion cells from retinal IR injury by attenuating the activation of microglia and the Nlrp3 inflammasome pathway. Moreover, its genetic knockout in mice or knockdown in primary microglia promoted mitochondrial fusion, impaired mitochondrial membrane potential, and reactive oxygen species (ROS) production, diminished aerobic glycolysis, and ameliorated inflammation. It appears that 181-Rik may trigger the Nlrp3 inflammasome activation by controlling mitochondrial functions through inhibiting expression of the metabolic sensor uncoupling protein 2 (Ucp2) and activating expression of the Ca2+ sensors S100a8/a9. Together, our findings shed new light on the molecular pathogenesis of retinal IR injury and may provide a fresh therapeutic target for IR-associated neurodegenerative diseases.

The mammalian genome encodes thousands of long non-coding RNAs (lncRNAs), many of which are developmentally regulated and differentially expressed across tissues, suggesting their potential roles in cellular differentiation. Despite this expression pattern, little is known about how lncRNAs influence lineage commitment at the molecular level. Here, we demonstrate that perturbation of an embryonic stem cell/early embryonic lncRNA, pluripotency-associated transcript 4 (Platr4), directly influences the specification of cardiac-mesoderm-lineage differentiation. We show that Platr4 acts as a molecular scaffold or chaperone interacting with the Hippo-signaling pathway molecules Yap and Tead4 to regulate the expression of a downstream target gene, Ctgf, which is crucial to the cardiac-lineage program. Importantly, Platr4 knockout mice exhibit myocardial atrophy and valve mucinous degeneration, which are both associated with reduced cardiac output and sudden heart failure. Together, our findings provide evidence that Platr4 is required in cardiac-lineage specification and adult heart function in mice.

* Alfredo Dueñas Rey Universitair Ziekenhuis Gent Centrum Medische Genetica Gent, Gent, Belgium Department of Biomolecular Medicine, Universiteit Gent Faculteit Geneeskunde en Gezondheidswetenschappen, Gent, Belgium * Víctor López-Soriano Universitair Ziekenhuis Gent Centrum Medische Genetica Gent, Gent, Belgium Department of Biomolecular Medicine, Universiteit Gent Faculteit Geneeskunde en Gezondheidswetenschappen, Gent, Belgium * Zelia Corradi Radboudumc Department of Human Genetics, Nijmegen, Gelderland, Netherlands * Claire-Marie Dhaenens Univ. Lille, Inserm, CHU Lille, U1172 - LilNCog - Lille Neuroscience & Cognition, Lille, France * Manon Bouckaert Universitair Ziekenhuis Gent Centrum Medische Genetica Gent, Gent, Belgium Department of Biomolecular Medicine, Universiteit Gent Faculteit Geneeskunde en Gezondheidswetenschappen, Gent, Belgium * Jasper Verwilt Department of Biomolecular Medicine, Universiteit Gent Faculteit Geneeskunde en Gezondheidswetenschappen, Gent, Belgium OncoRNALab, Cancer Research Institute Ghent, Ghent, Belgium * Avril M Watson Newcastle University Faculty of Medical Sciences, Newcastle upon Tyne, Newcastle upon Tyne, United Kingdom * Majlinda Lako Newcastle University Faculty of Medical Sciences, Newcastle upon Tyne, Newcastle upon Tyne, United Kingdom * Eva D’haene Universitair Ziekenhuis Gent Centrum Medische Genetica Gent, Gent, Belgium Department of Biomolecular Medicine, Universiteit Gent Faculteit Geneeskunde en Gezondheidswetenschappen, Gent, Belgium * Karla Alejandra Ruiz Ceja Telethon Institute of Genetics and Medicine, Napoli, Campania, Italy * Sandro Banfi Telethon Institute of Genetics and Medicine, Napoli, Campania, Italy * Miriam Bauwens Universitair Ziekenhuis Gent Centrum Medische Genetica Gent, Gent, Belgium Department of Biomolecular Medicine, Universiteit Gent Faculteit Geneeskunde en Gezondheidswetenschappen, Gent, Belgium * Frans P Cremers Radboudumc Department of Human Genetics, Nijmegen, Gelderland, Netherlands * Steve Lefever Universitair Ziekenhuis Gent Centrum Medische Genetica Gent, Gent, Belgium Department of Biomolecular Medicine, Universiteit Gent Faculteit Geneeskunde en Gezondheidswetenschappen, Gent, Belgium * Elfride De Baere Universitair Ziekenhuis Gent Centrum Medische Genetica Gent, Gent, Belgium Department of Biomolecular Medicine, Universiteit Gent Faculteit Geneeskunde en Gezondheidswetenschappen, Gent, Belgium * Frauke Coppieters Universitair Ziekenhuis Gent Centrum Medische Genetica Gent, Gent, Belgium Department of Biomolecular Medicine, Universiteit Gent Faculteit Geneeskunde en Gezondheidswetenschappen, Gent, Belgium