Reciprocal regulation of hnRNP C and CELF2 through translation and transcription tunes splicing activity in T cells

Abstract RNA binding proteins (RBPs) frequently regulate the expression of other RBPs in mammalian cells. Such cross-regulation has been proposed to be important to control networks of coordinated gene expression; however, much remains to be understood about how such networks of cross-regulation are established and what the functional consequence is of coordinated or reciprocal expression of RBPs. Here we demonstrate that the RBPs CELF2 and hnRNP C regulate the expression of each other, such that depletion of one results in reduced expression of the other. Specifically, we show that loss of hnRNP C reduces the transcription of CELF2 mRNA, while loss of CELF2 results in decreased efficiency of hnRNP C translation. We further demonstrate that this reciprocal regulation serves to fine tune the splicing patterns of many downstream target genes. Together, this work reveals new activities of hnRNP C and CELF2, provides insight into a previously unrecognized gene regulatory network, and demonstrates how cross-regulation of RBPs functions to shape the cellular transcriptome.

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RNA-binding proteins and heat-shock protein 90 are constituents of the cytoplasmic capping enzyme interactome

Journal of Biological Chemistry ◽

10.1074/jbc.ra118.004973 ◽

2018 ◽

Vol 293 (43) ◽

pp. 16596-16607 ◽

Cited By ~ 6

Author(s):

Jackson B. Trotman ◽

Bernice A. Agana ◽

Andrew J. Giltmier ◽

Vicki H. Wysocki ◽

Daniel R. Schoenberg

Keyword(s):

Heat Shock ◽

Heat Shock Protein ◽

Mammalian Cells ◽

Binding Proteins ◽

Rna Binding ◽

Rna Binding Proteins ◽

Heat Shock Protein 90 ◽

Hsp90 Inhibitor ◽

Capping Enzyme ◽

Eukaryotic Mrnas

The N7-methylguanosine cap is added in the nucleus early in gene transcription and is a defining feature of eukaryotic mRNAs. Mammalian cells also possess cytoplasmic machinery for restoring the cap at uncapped or partially degraded RNA 5′ ends. Central to both pathways is capping enzyme (CE) (RNA guanylyltransferase and 5′-phosphatase (RNGTT)), a bifunctional, nuclear and cytoplasmic enzyme. CE is recruited to the cytoplasmic capping complex by binding of a C-terminal proline-rich sequence to the third Src homology 3 (SH3) domain of NCK adapter protein 1 (NCK1). To gain broader insight into the cellular context of cytoplasmic recapping, here we identified the protein interactome of cytoplasmic CE in human U2OS cells through two complementary approaches: chemical cross-linking and recovery with cytoplasmic CE and protein screening with proximity-dependent biotin identification (BioID). This strategy unexpectedly identified 66 proteins, 52 of which are RNA-binding proteins. We found that CE interacts with several of these proteins independently of RNA, mediated by sequences within its N-terminal triphosphatase domain, and we present a model describing how CE-binding proteins may function in defining recapping targets. This analysis also revealed that CE is a client protein of heat shock protein 90 (HSP90). Nuclear and cytoplasmic CEs were exquisitely sensitive to inhibition of HSP90, with both forms declining significantly following treatment with each of several HSP90 inhibitors. Importantly, steady-state levels of capped mRNAs decreased in cells treated with the HSP90 inhibitor geldanamycin, raising the possibility that the cytotoxic effect of these drugs may partially be due to a general reduction in translatable mRNAs.

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Premature Translation of oskar in Oocytes Lacking the RNA-Binding Protein Bicaudal-C

Molecular and Cellular Biology ◽

10.1128/mcb.18.8.4855 ◽

1998 ◽

Vol 18 (8) ◽

pp. 4855-4862 ◽

Cited By ~ 60

Author(s):

Emma E. Saffman ◽

Sylvia Styhler ◽

Katherine Rother ◽

Weihua Li ◽

Stéphane Richard ◽

...

Keyword(s):

Drosophila Melanogaster ◽

Point Mutation ◽

Mammalian Cells ◽

Rna Binding ◽

Rna Binding Proteins ◽

Rna Binding Protein ◽

C Protein ◽

Kh Domain ◽

Anterior Posterior

ABSTRACT Bicaudal-C (Bic-C) is required duringDrosophila melanogaster oogenesis for several processes, including anterior-posterior patterning. The gene encodes a protein with five copies of the KH domain, a motif found in a number of RNA-binding proteins. Using antibodies raised against the BIC-C protein, we show that multiple isoforms of the protein exist in ovaries and that the protein, like the RNA, accumulates in the developing oocyte early in oogenesis. BIC-C protein expressed in mammalian cells can bind RNA in vitro, and a point mutation in one of the KH domains that causes a strong Bic-C phenotype weakens this binding. In addition, oskar translation commences prior to posterior localization of oskar RNA inBic-C − oocytes, indicating thatBic-C may regulate oskar translation during oogenesis.

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A Posttranscriptional Mechanism That Controls Ptbp1 Abundance in the Xenopus Epidermis

Molecular and Cellular Biology ◽

10.1128/mcb.01040-14 ◽

2014 ◽

Vol 35 (4) ◽

pp. 758-768 ◽

Cited By ~ 4

Author(s):

Agnès Méreau ◽

Vincent Anquetil ◽

Hubert Lerivray ◽

Justine Viet ◽

Claire Schirmer ◽

...

Keyword(s):

Rna Binding ◽

Rna Binding Proteins ◽

Fine Tuning ◽

Splice Isoforms ◽

Content Type ◽

Splicing Pattern ◽

Exon 11 ◽

Splicing Patterns ◽

Different Tissues

The output of alternative splicing depends on the cooperative or antagonistic activities of several RNA-binding proteins (RBPs), like Ptbp1 and Esrp1 inXenopus. Fine-tuning of the RBP abundance is therefore of prime importance to achieve tissue- or cell-specific splicing patterns. Here, we addressed the mechanisms leading to the high expression of theptbp1gene, which encodes Ptbp1, inXenopusepidermis. Two splice isoforms ofptbp1mRNA differ by the presence of an alternative exon 11, and only the isoform including exon 11 can be translated to a full-length protein.In vivominigene assays revealed that the nonproductive isoform was predominantly produced. Knockdown experiments demonstrated that Esrp1, which is specific to the epidermis, strongly stimulated the expression ofptbp1by favoring the productive isoform. Consequently, knocking downesrp1phenocopiedptbp1inactivation. Conversely, Ptbp1 repressed the expression of its own gene by favoring the nonproductive isoform. Hence, a complex posttranscriptional mechanism controls Ptbp1 abundance inXenopusepidermis: skipping of exon 11 is the default splicing pattern, but Esrp1 stimulatesptbp1expression by favoring the inclusion of exon 11 up to a level that is limited by Ptbp1 itself. These results decipher a posttranscriptional mechanism that achieves various abundances of the ubiquitous RBP Ptbp1 in different tissues.

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Characterization and Functional Analysis of Polyadenylation Sites in Fast and Slow Muscles

BioMed Research International ◽

10.1155/2020/2626584 ◽

2020 ◽

Vol 2020 ◽

pp. 1-13

Author(s):

Lulu Deng ◽

Long Li ◽

Cheng Zou ◽

Chengchi Fang ◽

Changchun Li

Keyword(s):

Single Molecule ◽

Posttranscriptional Regulation ◽

Muscle Development ◽

Target Genes ◽

Rna Binding ◽

De Novo ◽

Rna Binding Proteins ◽

Fast And Slow Muscles ◽

Molecular Features ◽

Polyadenylation Sites

Many increasing documents have proved that alternative polyadenylation (APA) events with different polyadenylation sites (PAS) contribute to posttranscriptional regulation. However, little is known about the detailed molecular features of PASs and its role in porcine fast and slow skeletal muscles through microRNAs (miRNAs) and RNA binding proteins (RBPs). In this study, we combined single-molecule real-time sequencing and Illumina RNA-seq datasets to comprehensively analyze polyadenylation in pigs. We identified a total of 10,334 PASs, of which 8734 were characterized by reference genome annotation. 32.86% of PAS-associated genes were determined to have more than one PAS. Further analysis demonstrated that tissue-specific PASs between fast and slow muscles were enriched in skeletal muscle development pathways. In addition, we obtained 1407 target genes regulated by APA events through potential binding 69 miRNAs and 28 RBPs in variable 3′ UTR regions and some are involved in myofiber transformation. Furthermore, the de novo motif search confirmed that the most common usage of canonical motif AAUAAA and three types of PASs may be related to the strength of motifs. In summary, our results provide a useful annotation of PASs for pig transcriptome and suggest that APA may serve as a role in fast and slow muscle development under the regulation of miRNAs and RBPs.

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Vimentin protects differentiating stem cells from stress

Scientific Reports ◽

10.1038/s41598-020-76076-4 ◽

2020 ◽

Vol 10 (1) ◽

Cited By ~ 1

Author(s):

Sundararaghavan Pattabiraman ◽

Gajendra Kumar Azad ◽

Triana Amen ◽

Shlomi Brielle ◽

Jung Eun Park ◽

...

Keyword(s):

Stem Cells ◽

Stress Response ◽

Mammalian Cells ◽

Rna Binding ◽

Rna Binding Proteins ◽

Embryonic Stem ◽

Cellular Stress Response ◽

Cell Stiffness ◽

Asymmetric Partitioning ◽

Organismal Development

Abstract Vimentin is one of the first cytoplasmic intermediate filaments to be expressed in mammalian cells during embryogenesis, but its role in cellular fitness has long been a mystery. Vimentin is acknowledged to play a role in cell stiffness, cell motility, and cytoplasmic organization, yet it is widely considered to be dispensable for cellular function and organismal development. Here, we show that Vimentin plays a role in cellular stress response in differentiating cells, by recruiting aggregates, stress granules, and RNA-binding proteins, directing their elimination and asymmetric partitioning. In the absence of Vimentin, pluripotent embryonic stem cells fail to differentiate properly, with a pronounced deficiency in neuronal differentiation. Our results uncover a novel function for Vimentin, with important implications for development, tissue homeostasis, and in particular, stress response.

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Concurrent binding to DNA and RNA facilitates the pluripotency reprogramming activity of Sox2

Nucleic Acids Research ◽

10.1093/nar/gkaa067 ◽

2020 ◽

Vol 48 (7) ◽

pp. 3869-3887 ◽

Cited By ~ 1

Author(s):

Linlin Hou ◽

Yuanjie Wei ◽

Yingying Lin ◽

Xiwei Wang ◽

Yiwei Lai ◽

...

Keyword(s):

Dna Sequences ◽

Target Genes ◽

Rna Binding ◽

Rna Binding Proteins ◽

High Mobility ◽

Binding Motif ◽

Dna And Rna ◽

Somatic Cell Reprogramming ◽

Double Stranded Dna

Abstract Some transcription factors that specifically bind double-stranded DNA appear to also function as RNA-binding proteins. Here, we demonstrate that the transcription factor Sox2 is able to directly bind RNA in vitro as well as in mouse and human cells. Sox2 targets RNA via a 60-amino-acid RNA binding motif (RBM) positioned C-terminally of the DNA binding high mobility group (HMG) box. Sox2 can associate with RNA and DNA simultaneously to form ternary RNA/Sox2/DNA complexes. Deletion of the RBM does not affect selection of target genes but mitigates binding to pluripotency related transcripts, switches exon usage and impairs the reprogramming of somatic cells to a pluripotent state. Our findings designate Sox2 as a multi-functional factor that associates with RNA whilst binding to cognate DNA sequences, suggesting that it may co-transcriptionally regulate RNA metabolism during somatic cell reprogramming.

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Design and application of circular RNAs with protein-sponge function

Nucleic Acids Research ◽

10.1093/nar/gkaa1085 ◽

2020 ◽

Vol 48 (21) ◽

pp. 12326-12335

Author(s):

Silke Schreiner ◽

Anna Didio ◽

Lee-Hsueh Hung ◽

Albrecht Bindereif

Keyword(s):

Alternative Splicing ◽

Mammalian Cells ◽

Regulatory Networks ◽

Rna Binding ◽

Molecular Medicine ◽

Circular Rnas ◽

Ca Repeat ◽

New Strategy ◽

Nuclear Ribonucleoprotein ◽

Splicing Patterns

Abstract Circular RNAs (circRNAs) are a class of noncoding RNAs, generated from pre-mRNAs by circular splicing of exons and functionally largely uncharacterized. Here we report on the design, expression, and characterization of artificial circRNAs that act as protein sponges, specifically binding and functionally inactivating hnRNP (heterogeneous nuclear ribonucleoprotein) L. HnRNP L regulates alternative splicing, depending on short CA-rich RNA elements. We demonstrate that designer hnRNP L-sponge circRNAs with CA-repeat or CA-rich sequence clusters can efficiently and specifically modulate splicing-regulatory networks in mammalian cells, including alternative splicing patterns and the cellular distribution of a splicing factor. This new strategy can in principle be applied to any RNA-binding protein, opening up new therapeutic strategies in molecular medicine.

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Posttranscriptional gene regulation by RNA-binding proteins during oxidative stress: implications for cellular senescence

Biological Chemistry ◽

10.1515/bc.2008.022 ◽

2008 ◽

Vol 389 (3) ◽

pp. 243-255 ◽

Cited By ~ 156

Author(s):

Kotb Abdelmohsen ◽

Yuki Kuwano ◽

Hyeon Ho Kim ◽

Myriam Gorospe

Keyword(s):

Oxidative Stress ◽

Gene Expression ◽

Cellular Senescence ◽

Mammalian Cells ◽

Rna Binding ◽

Rna Binding Proteins ◽

Expression Profiles ◽

Expression Patterns ◽

Oxidant Damage ◽

Regulated Gene Expression

AbstractTo respond adequately to oxidative stress, mammalian cells elicit rapid and tightly controlled changes in gene expression patterns. Besides alterations in the subsets of transcribed genes, two posttranscriptional processes prominently influence the oxidant-triggered gene expression programs: mRNA turnover and translation. Here, we review recent progress in our knowledge of theturnover andtranslationregulatory (TTR) mRNA-bindingproteins (RBPs) that influence gene expression in response to oxidative damage. Specifically, we identify oxidant damage-regulated mRNAs that are targets of TTR-RBPs, we review the oxidant-triggered signaling pathways that govern TTR-RBP function, and we examine emerging evidence that TTR-RBP activity is altered with senescence and aging. Given the potent influence of TTR-RBPs upon oxidant-regulated gene expression profiles, we propose that the senescence-associated changes in TTR-RBPs directly contribute to the impaired responses to oxidant damage that characterize cellular senescence and advancing age.

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M1 and M2 Functional Imprinting of Primary Microglia: Role of P2X7 Activation and miR-125b

Mediators of Inflammation ◽

10.1155/2016/2989548 ◽

2016 ◽

Vol 2016 ◽

pp. 1-9 ◽

Cited By ~ 20

Author(s):

Chiara Parisi ◽

Giulia Napoli ◽

Pablo Pelegrin ◽

Cinzia Volonté

Keyword(s):

Motor Neuron ◽

Target Genes ◽

Rna Binding ◽

Rna Binding Proteins ◽

Short Review ◽

Severe Form ◽

Primary Microglia ◽

Neuron Death ◽

Fused In Sarcoma ◽

Neuroprotective Actions

Amyotrophic lateral sclerosis (ALS) is a most frequently occurring and severe form of motor neuron disease, causing death within 3–5 years from diagnosis and with a worldwide incidence of about 2 per 100,000 person-years. Mutations in over twenty genes associated with familial forms of ALS have provided insights into the mechanisms leading to motor neuron death. Moreover, mutations in two RNA binding proteins, TAR DNA binding protein 43 and fused in sarcoma, have raised the intriguing possibility that perturbations of RNA metabolism, including that of the small endogenous RNA molecules that repress target genes at the posttranscriptional level, that is, microRNAs, may contribute to disease pathogenesis. At present, the mechanisms by which microglia actively participate to both toxic and neuroprotective actions in ALS constitute an important matter of research. Among the pathways involved in ALS-altered microglia responses, in previous works we have uncovered the hyperactivation of P2X7 receptor by extracellular ATP and the overexpression of miR-125b, both leading to uncontrolled toxic M1 reactions. In order to shed further light on the complexity of these processes, in this short review we will describe the M1/M2 functional imprinting of primary microglia and a role played by P2X7 and miR-125b in ALS microglia activation.

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Dr. Jekyll and Mr. Hyde: The Two Faces of the FUS/EWS/TAF15 Protein Family

Sarcoma ◽

10.1155/2011/837474 ◽

2011 ◽

Vol 2011 ◽

pp. 1-13 ◽

Cited By ~ 64

Author(s):

Heinrich Kovar

Keyword(s):

Protein Function ◽

Target Genes ◽

Rna Binding ◽

Rna Binding Proteins ◽

Fusion Gene ◽

Tumor Biology ◽

Central Function ◽

Epithelial Cancers ◽

Transcription Factor Genes

FUS, EWS, and TAF15 form the FET family of RNA-binding proteins whose genes are found rearranged with various transcription factor genes predominantly in sarcomas and in rare hematopoietic and epithelial cancers. The resulting fusion gene products have attracted considerable interest as diagnostic and promising therapeutic targets. So far, oncogenic FET fusion proteins have been regarded as strong transcription factors that aberrantly activate or repress target genes of their DNA-binding fusion partners. However, the role of the transactivating domain in the context of the normal FET proteins is poorly defined, and, therefore, our knowledge on how FET aberrations impact on tumor biology is incomplete. Since we believe that a full understanding of aberrant FET protein function can only arise from looking at both sides of the coin, the good and the evil, this paper summarizes evidence for the central function of FET proteins in bridging RNA transcription, processing, transport, and DNA repair.

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