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Matrin3 is an RNA- and DNA-binding nuclear matrix protein found to be associated with neural and muscular degenerative diseases. A number of possible functions of Matrin3 have been suggested, but no widespread role in RNA metabolism has yet been clearly demonstrated. We identified Matrin3 by its interaction with the second RRM domain of the splicing regulator PTB. Using a combination of RNAi knockdown, transcriptome profiling and iCLIP, we find that Matrin3 is a regulator of hundreds of alternative splicing events, principally acting as a splicing repressor with only a small proportion of targeted events being co-regulated by PTB. In contrast to other splicing regulators, Matrin3 binds to an extended region within repressed exons and flanking introns with no sharply defined peaks. The identification of this clear molecular function of Matrin3 should help to clarify the molecular pathology of ALS and other diseases caused by mutations of Matrin3., (© 2015 The Authors. Published under the terms of the CC BY 4.0 license.)

Oncogenic RAS signaling reprograms gene expression through both transcriptional and post‐transcriptional mechanisms. While transcriptional regulation downstream of RAS is relatively well characterized, how RAS post‐transcriptionally modulates gene expression to promote malignancy remains largely unclear. Using quantitative RNA interactome capture analysis, we here reveal that oncogenic RAS signaling reshapes the RNA‐bound proteomic landscape of pancreatic cancer cells, with a network of nuclear proteins centered around nucleolin displaying enhanced RNA‐binding activity. We show that nucleolin is phosphorylated downstream of RAS, which increases its binding to pre‐ribosomal RNA (rRNA), boosts rRNA production, and promotes ribosome biogenesis. This nucleolin‐dependent enhancement of ribosome biogenesis is crucial for RAS‐induced pancreatic cancer cell proliferation and can be targeted therapeutically to inhibit tumor growth. Our results reveal that oncogenic RAS signaling drives ribosome biogenesis by regulating the RNA‐binding activity of nucleolin and highlight a crucial role for this mechanism in RAS‐mediated tumorigenesis. Synopsis: Oncogenic RAS‐induced ERK1/2 activity promotes the phosphorylation and RNA‐binding activity of nucleolin (NCL), boosting ribosome biogenesis, protein synthesis, and pancreatic tumorigenesis. Oncogenic KRAS reshapes the RNA‐bound proteomic landscape of pancreatic ductal adenocarcinoma cells, increasing the activity of RNA‐binding proteins, including NCL.NCL is phosphorylated downstream of oncogenic KRAS, via ERK1/2‐mediated activation of casein kinase 2, enhancing its binding to pre‐rRNA.Increased NCL–pre‐rRNA binding boosts nascent pre‐rRNA levels, leading to increased ribosome biogenesis and protein synthesis.NCL‐promoted ribosome biogenesis is required for KRAS‐driven tumorigenesis and can be therapeutically exploited. [ABSTRACT FROM AUTHOR]

Translational control of mRNAs is a point of convergence for many oncogenic signals through which cancer cells tune protein expression in tumorigenesis. Cancer cells rely on translational control to appropriately adapt to limited resources while maintaining cell growth and survival, which creates a selective therapeutic window compared to non‐transformed cells. In this review, we first discuss how cancer cells modulate the translational machinery to rapidly and selectively synthesize proteins in response to internal oncogenic demands and external factors in the tumor microenvironment. We highlight the clinical potential of compounds that target different translation factors as anti‐cancer therapies. Next, we detail how RNA sequence and structural elements interface with the translational machinery and RNA‐binding proteins to coordinate the translation of specific pro‐survival and pro‐growth programs. Finally, we provide an overview of the current and emerging technologies that can be used to illuminate the mechanisms of selective translational control in cancer cells as well as within the microenvironment. [ABSTRACT FROM AUTHOR]

Post‐translational modifications (PTMs) have emerged as key modulators of protein phase separation and have been linked to protein aggregation in neurodegenerative disorders. The major aggregating protein in amyotrophic lateral sclerosis and frontotemporal dementia, the RNA‐binding protein TAR DNA‐binding protein (TDP‐43), is hyperphosphorylated in disease on several C‐terminal serine residues, a process generally believed to promote TDP‐43 aggregation. Here, we however find that Casein kinase 1δ‐mediated TDP‐43 hyperphosphorylation or C‐terminal phosphomimetic mutations reduce TDP‐43 phase separation and aggregation, and instead render TDP‐43 condensates more liquid‐like and dynamic. Multi‐scale molecular dynamics simulations reveal reduced homotypic interactions of TDP‐43 low‐complexity domains through enhanced solvation of phosphomimetic residues. Cellular experiments show that phosphomimetic substitutions do not affect nuclear import or RNA regulatory functions of TDP‐43, but suppress accumulation of TDP‐43 in membrane‐less organelles and promote its solubility in neurons. We speculate that TDP‐43 hyperphosphorylation may be a protective cellular response to counteract TDP‐43 aggregation. Synopsis: C‐terminal hyperphosphorylation of TDP‐43, a pathological hallmark of several neurodegenerative disorders, enhances the liquidity of TDP‐43 condensates and suppresses TDP‐43 condensation and aggregation, shedding a new light on this TDP‐43 disease‐linked posttranslational modification. TDP‐43 phosphorylation by CK1δ or C‐terminal phosphomimetic mutations suppress TDP‐43 phase separation and aggregation.C‐terminal phosphomimetic mutations enhance liquidity and dynamics of TDP‐43 condensates.Suppression of phase separation is associated with loss of protein‐protein interactions in the C‐terminus and enhanced solvation of negatively charged groups.TDP‐43 bearing C‐terminal phosphomimetic mutations fails to condense into stress‐induced membrane‐less compartments and remains dispersed. [ABSTRACT FROM AUTHOR]

Abstract: TDP‐43 (encoded by the gene TARDBP) is an RNA binding protein central to the pathogenesis of amyotrophic lateral sclerosis (ALS). However, how TARDBP mutations trigger pathogenesis remains unknown. Here, we use novel mouse mutants carrying point mutations in endogenous Tardbp to dissect TDP‐43 function at physiological levels both in vitro and in vivo. Interestingly, we find that mutations within the C‐terminal domain of TDP‐43 lead to a gain of splicing function. Using two different strains, we are able to separate TDP‐43 loss‐ and gain‐of‐function effects. TDP‐43 gain‐of‐function effects in these mice reveal a novel category of splicing events controlled by TDP‐43, referred to as “skiptic” exons, in which skipping of constitutive exons causes changes in gene expression. In vivo, this gain‐of‐function mutation in endogenous Tardbp causes an adult‐onset neuromuscular phenotype accompanied by motor neuron loss and neurodegenerative changes. Furthermore, we have validated the splicing gain‐of‐function and skiptic exons in ALS patient‐derived cells. Our findings provide a novel pathogenic mechanism and highlight how TDP‐43 gain of function and loss of function affect RNA processing differently, suggesting they may act at different disease stages. [ABSTRACT FROM AUTHOR]

FUS is an RNA-binding protein involved in amyotrophic lateral sclerosis ( ALS) and frontotemporal dementia ( FTD). Cytoplasmic FUS-containing aggregates are often associated with concomitant loss of nuclear FUS. Whether loss of nuclear FUS function, gain of a cytoplasmic function, or a combination of both lead to neurodegeneration remains elusive. To address this question, we generated knockin mice expressing mislocalized cytoplasmic FUS and complete FUS knockout mice. Both mouse models display similar perinatal lethality with respiratory insufficiency, reduced body weight and length, and largely similar alterations in gene expression and mRNA splicing patterns, indicating that mislocalized FUS results in loss of its normal function. However, FUS knockin mice, but not FUS knockout mice, display reduced motor neuron numbers at birth, associated with enhanced motor neuron apoptosis, which can be rescued by cell-specific CRE-mediated expression of wild-type FUS within motor neurons. Together, our findings indicate that cytoplasmic FUS mislocalization not only leads to nuclear loss of function, but also triggers motor neuron death through a toxic gain of function within motor neurons. [ABSTRACT FROM AUTHOR]

The RNA‐binding protein ALYREF plays key roles in nuclear export and also 3′‐end processing of polyadenylated mRNAs, but whether such regulation also extends to non‐polyadenylated RNAs is unknown. Replication‐dependent (RD)‐histone mRNAs are not polyadenylated, but instead end in a stem‐loop (SL) structure. Here, we demonstrate that ALYREF prevalently binds a region next to the SL on RD‐histone mRNAs. SL‐binding protein (SLBP) directly interacts with ALYREF and promotes its recruitment. ALYREF promotes histone pre‐mRNA 3′‐end processing by facilitating U7‐snRNP recruitment through physical interaction with the U7‐snRNP‐specific component Lsm11. Furthermore, ALYREF, together with other components of the TREX complex, enhances histone mRNA export. Moreover, we show that 3′‐end processing promotes ALYREF recruitment and histone mRNA export. Together, our results point to an important role of ALYREF in coordinating 3′‐end processing and nuclear export of non‐polyadenylated mRNAs. Synopsis: The RNA‐binding protein ALYREF functions in coupling nuclear export and 3′‐end processing of polyadenylated mRNAs. New data identify a parallel mechanism for ALYREF also linking 3′‐end processing to nuclear export of non‐polyadenylated, replication‐dependent histone mRNAs. ALYREF prevalently binds to a 3′ region of histone mRNAs in an SLBP‐dependent manner.ALYREF promotes histone pre‐mRNA processing by ensuring U7‐snRNP recruitment.The TREX complex facilitates histone mRNA export by recruiting NXF1.3′‐end processing promotes ALYREF recruitment and histone mRNA export. [ABSTRACT FROM AUTHOR]

Elevated expression of RNA binding protein HNRNPC has been reported in cancer cells, while the essentialness and functions of HNRNPC in tumors were not clear. We showed that repression of HNRNPC in the breast cancer cells MCF7 and T47D inhibited cell proliferation and tumor growth. Our computational inference of the key pathways and extensive experimental investigations revealed that the cascade of interferon responses mediated by RIG‐I was responsible for such tumor‐inhibitory effect. Interestingly, repression of HNRNPC resulted in accumulation of endogenous double‐stranded RNA (dsRNA), the binding ligand of RIG‐I. These up‐regulated dsRNA species were highly enriched by Alu sequences and mostly originated from pre‐mRNA introns that harbor the known HNRNPC binding sites. Such source of dsRNA is different than the recently well‐characterized endogenous retroviruses that encode dsRNA. In summary, essentialness of HNRNPC in the breast cancer cells was attributed to its function in controlling the endogenous dsRNA and the down‐stream interferon response. This is a novel extension from the previous understandings about HNRNPC in binding with introns and regulating RNA splicing. Synopsis: Repression of RNA binding protein HNRNPC in breast cancer cells MCF7 and T47D resulted in accumulation of endogenous dsRNA species mostly from Alu introns, which triggered interferon response and tumor growth arrest. HNRNPC is highly expressed in breast cancer tumors and repression of HNRNPC arrests proliferation of MCF7 and T47D cells.Interferon response mediated by RNA sensor RIG‐I is responsible for anti‐proliferation effect of HNRNPC repression.Repression of HNRNPC induces immunostimulatory endogenous dsRNA mostly from introns with Alu.Production of Alu dsRNA can be traced back to RNA quality control machinery such as nonsense‐mediated decay. Repression of the RNA binding protein HNRNPC in breast cancer cells causes accumulation of endogenous dsRNA species leading to interferon response and tumor growth arrest. [ABSTRACT FROM AUTHOR]