scholarly journals The RNA-binding protein SUP-12 controls muscle-specific splicing of the ADF/cofilin pre-mRNA in C. elegans

2004 ◽  
Vol 167 (4) ◽  
pp. 639-647 ◽  
Author(s):  
Akwasi Anyanful ◽  
Kanako Ono ◽  
Robert C. Johnsen ◽  
Hinh Ly ◽  
Victor Jensen ◽  
...  
Keyword(s):  
Rna Binding ◽  
Functional Redundancy ◽  
Rna Recognition Motif ◽  
Rna Recognition ◽  
Speckled Pattern ◽  
Tissue Specific ◽  
C Elegans ◽  
Rrm Domain ◽  
New Family ◽  
Specific Alternative

Tissue-specific alternative pre-mRNA splicing is essential for increasing diversity of functionally different gene products. In Caenorhabditis elegans, UNC-60A and UNC-60B, nonmuscle and muscle isoforms of actin depolymerizing factor (ADF)/cofilin, are expressed by alternative splicing of unc-60 and regulate distinct actin-dependent developmental processes. We report that SUP-12, a member of a new family of RNA recognition motif (RRM) proteins, including SEB-4, regulates muscle-specific splicing of unc-60. In sup-12 mutants, expression of UNC-60B is decreased, whereas UNC-60A is up-regulated in muscle. sup-12 mutations strongly suppress muscle defects in unc-60B mutants by allowing expression of UNC-60A in muscle that can substitute for UNC-60B, thus unmasking their functional redundancy. SUP-12 is expressed in muscle and localized to the nuclei in a speckled pattern. The RRM domain of SUP-12 binds to several sites of the unc-60 pre-mRNA including the UG repeats near the 3′-splice site in the first intron. Our results suggest that SUP-12 is a novel tissue-specific splicing factor and regulates functional redundancy among ADF/cofilin isoforms.

scholarly journals Structural basis underlying CAC RNA recognition by the RRM domain of dimeric RNA-binding protein RBPMS

2015 ◽  
Vol 49 ◽  
Author(s):  
Marianna Teplova ◽  
Thalia A. Farazi ◽  
Thomas Tuschl ◽  
Dinshaw J. Patel
Keyword(s):  
Rna Binding ◽  
Binding Protein ◽  
Rna Binding Protein ◽  
Hek293 Cells ◽  
Structural Basis ◽  
Rna Recognition Motif ◽  
Rna Recognition ◽  
Binding Studies ◽  
Muscle Plasticity ◽  
Rrm Domain

AbstractRNA-binding protein with multiple splicing (designated RBPMS) is a higher vertebrate mRNA-binding protein containing a single RNA recognition motif (RRM). RBPMS has been shown to be involved in mRNA transport, localization and stability, with key roles in axon guidance, smooth muscle plasticity, as well as regulation of cancer cell proliferation and migration. We report on structure-function studies of the RRM domain of RBPMS bound to a CAC-containing single-stranded RNA. These results provide insights into potential topologies of complexes formed by the RBPMS RRM domain and the tandem CAC repeat binding sites as detected by photoactivatable-ribonucleoside-enhanced crosslinking and immunoprecipitation. These studies establish that the RRM domain of RBPMS forms a symmetrical dimer in the free state, with each monomer binding sequence-specifically to all three nucleotides of a CAC segment in the RNA bound state. Structure-guided mutations within the dimerization and RNA-binding interfaces of RBPMS RRM on RNA complex formation resulted in both disruption of dimerization and a decrease in RNA-binding affinity as observed by size exclusion chromatography and isothermal titration calorimetry. As anticipated from biochemical binding studies, over-expression of dimerization or RNA-binding mutants of Flag-HA-tagged RBPMS were no longer able to track with stress granules in HEK293 cells, thereby documenting the deleterious effects of such mutations in vivo.

scholarly journals Dose-dependent action of the RNA binding protein FOX-1 to relay X-chromosome number and determine C. elegans sex

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Behnom Farboud ◽  
Catherine S Novak ◽  
Monique Nicoll ◽  
Alyssa Quiogue ◽  
Barbara J Meyer
Keyword(s):  
Chromosome Number ◽  
X Chromosome ◽  
Rna Binding ◽  
Binding Protein ◽  
Intron Retention ◽  
Rna Binding Protein ◽  
Rna Recognition Motif ◽  
Rna Recognition ◽  
C Elegans ◽  
Dose Dependent

We demonstrate how RNA binding protein FOX-1 functions as a dose-dependent X-signal element to communicate X-chromosome number and thereby determine nematode sex. FOX-1, an RNA recognition motif protein, triggers hermaphrodite development in XX embryos by causing non-productive alternative pre-mRNA splicing of xol-1, the master sex-determination switch gene that triggers male development in XO embryos. RNA binding experiments together with genome editing demonstrate that FOX-1 binds to multiple GCAUG and GCACG motifs in a xol-1 intron, causing intron retention or partial exon deletion, thereby eliminating male-determining XOL-1 protein. Transforming all motifs to GCAUG or GCACG permits accurate alternative splicing, demonstrating efficacy of both motifs. Mutating subsets of both motifs partially alleviates non-productive splicing. Mutating all motifs blocks it, as does transforming them to low-affinity GCUUG motifs. Combining multiple high-affinity binding sites with the twofold change in FOX-1 concentration between XX and XO embryos achieves dose-sensitivity in splicing regulation to determine sex.

scholarly journals Structural basis for RNA recognition by a type II poly(A)-binding protein

2008 ◽  
Vol 105 (40) ◽  
pp. 15317-15322 ◽  
Author(s):  
Jikui Song ◽  
Jered V. McGivern ◽  
Karl W. Nichols ◽  
John L. Markley ◽  
Michael D. Sheets
Keyword(s):  
Rna Binding ◽  
Binding Protein ◽  
Recognition Site ◽  
Structural Basis ◽  
Rna Recognition Motif ◽  
Functional Domain ◽  
Rna Recognition ◽  
Type Ii ◽  
Rrm Domain ◽  
Polyproline Motif

We identified a functional domain (XlePABP2-TRP) of Xenopus laevis embryonic type II poly(A)-binding protein (XlePABP2). The NMR structure of XlePABP2-TRP revealed that the protein is a homodimer formed by the antiparallel association of β-strands from the single RNA recognition motif (RRM) domain of each subunit. In each subunit of the homodimer, the canonical RNA recognition site is occluded by a polyproline motif. Upon poly(A) binding, XlePABP2-TRP undergoes a dimer-monomer transition that removes the polyproline motif from the RNA recognition site and allows it to be replaced by the adenosine nucleotides of poly(A). Our results provide high-resolution structural information concerning type II PABPs and an example of a single RRM domain protein that transitions from a homodimer to a monomer upon RNA binding. These findings advance our understanding of RRM domain regulation, poly(A) recognition, and are relevant to understanding how type II PABPs function in mRNA processing and human disease.

scholarly journals Crystal structure of human Acinus RNA recognition motif domain

PeerJ ◽  
2018 ◽  
Vol 6 ◽  
pp. e5163 ◽  
Author(s):  
Humberto Fernandes ◽  
Honorata Czapinska ◽  
Katarzyna Grudziaz ◽  
Janusz M. Bujnicki ◽  
Martyna Nowacka
Keyword(s):  
Crystal Structure ◽  
Rna Binding ◽  
Chromatin Condensation ◽  
Rna Recognition Motif ◽  
Target Sequence ◽  
Rna Recognition ◽  
Recognition Motif ◽  
Rrm Domain ◽  
Α Helix ◽  
Β Sheet

Acinus is an abundant nuclear protein involved in apoptosis and splicing. It has been implicated in inducing apoptotic chromatin condensation and DNA fragmentation during programmed cell death. Acinus undergoes activation by proteolytic cleavage that produces a truncated p17 form that comprises only the RNA recognition motif (RRM) domain. We have determined the crystal structure of the human Acinus RRM domain (AcRRM) at 1.65 Å resolution. It shows a classical four-stranded antiparallel β-sheet fold with two flanking α-helices and an additional, non-classical α-helix at the C-terminus, which harbors the caspase-3 target sequence that is cleaved during Acinus activation. In the structure, the C-terminal α-helix partially occludes the potential ligand binding surface of the β-sheet and hypothetically shields it from non-sequence specific interactions with RNA. Based on the comparison with other RRM-RNA complex structures, it is likely that the C-terminal α-helix changes its conformation with respect to the RRM core in order to enable RNA binding by Acinus.

Identification and expression analysis of Dazl homologue in Cynops cyanurus

Zygote ◽  
2021 ◽  
pp. 1-6
Author(s):  
Yinjiao Zhao ◽  
Ya Du ◽  
Qinglan Ge ◽  
Fang Yan ◽  
Shu Wei
Keyword(s):  
Phylogenetic Analysis ◽  
Comparative Studies ◽  
Rna Binding ◽  
Rna Binding Protein ◽  
Rna Recognition Motif ◽  
Rna Recognition ◽  
Specific Expression ◽  
Recognition Motif ◽  
Deleted In Azoospermia ◽  
Specific Expression Pattern

Summary The Dazl (deleted in azoospermia-like) gene encodes an RNA-binding protein containing an RNA recognition motif (RRM) and a DAZ motif. Dazl is essential for gametogenesis in vertebrates. In this study, we report the cloning of Dazl cDNA from Cynops cyanurus. Ccdazl mRNA showed a germline-specific expression pattern as expected. Ccdazl expression gradually decreased during oogenesis, suggesting that it may be involved in oocyte development. Phylogenetic analysis revealed that the Ccdazl protein shares conserved motifs/domains with Dazl proteins from other species. Cloning of Ccdazl provides a new tool to carry out comparative studies of germ cell development in amphibians.

scholarly journals RNA recognition and self-association of CPEB4 is mediated by its tandem RRM domains

2014 ◽  
Vol 42 (15) ◽  
pp. 10185-10195 ◽  
Author(s):  
Constanze Schelhorn ◽  
James M.B. Gordon ◽  
Lidia Ruiz ◽  
Javier Alguacil ◽  
Enrique Pedroso ◽  
...  
Keyword(s):  
Rna Binding ◽  
Titration Calorimetry ◽  
Rna Recognition Motif ◽  
Rna Recognition ◽  
Mobility Shift ◽  
Cytoplasmic Polyadenylation ◽  
C Terminus ◽  
Self Association ◽  
Mobility Shift Assay ◽  
A Minor

Abstract Cytoplasmic polyadenylation is regulated by the interaction of the cytoplasmic polyadenylation element binding proteins (CPEB) with cytoplasmic polyadenylation element (CPE) containing mRNAs. The CPEB family comprises four paralogs, CPEB1–4, each composed of a variable N-terminal region, two RNA recognition motif (RRM) and a C-terminal ZZ-domain. We have characterized the RRM domains of CPEB4 and their binding properties using a combination of biochemical, biophysical and NMR techniques. Isothermal titration calorimetry, NMR and electrophoretic mobility shift assay experiments demonstrate that both the RRM domains are required for an optimal CPE interaction and the presence of either one or two adenosines in the two most commonly used consensus CPE motifs has little effect on the affinity of the interaction. Both the single RRM1 and the tandem RRM1–RRM2 have the ability to dimerize, although representing a minor population. Self-association does not affect the proteins’ ability to interact with RNA as demonstrated by ion mobility–mass spectrometry. Chemical shift effects measured by NMR of the apo forms of the RRM1–RRM2 samples indicate that the two domains are orientated toward each other. NMR titration experiments show that residues on the β-sheet surface on RRM1 and at the C-terminus of RRM2 are affected upon RNA binding. We propose a model of the CPEB4 RRM1–RRM2–CPE complex that illustrates the experimental data.

TLS (FUS) binds RNA in vivo and engages in nucleo-cytoplasmic shuttling

1997 ◽  
Vol 110 (15) ◽  
pp. 1741-1750 ◽  
Author(s):  
H. Zinszner ◽  
J. Sok ◽  
D. Immanuel ◽  
Y. Yin ◽  
D. Ron
Keyword(s):  
Rna Binding ◽  
Cellular Transformation ◽  
Rna Recognition Motif ◽  
Rna Recognition ◽  
Nucleocytoplasmic Shuttling ◽  
Uv Crosslinking ◽  
Recognition Motif ◽  
Specific Analysis ◽  
Human And Mouse

TLS, the product of a gene commonly translocated in liposarcomas (TLS), is prototypical of a newly identified class of nuclear proteins that contain a C-terminal domain with a distinct RNA recognition motif (RRM) surrounded by Arg-Gly-Gly (RGG) repeats. Its unique N terminus serves as an essential transforming domain for a number of fusion oncoproteins in human sarcomas and leukemias. In this study we use an in vivo UV crosslinking procedure to probe the interactions of TLS with RNA. TLS is found to bind RNA in vivo and the association of TLS with RNA is rapidly diminished by treating cells with transcriptional inhibitors. This suggests that the species bound by TLS turns over rapidly. Surprisingly, the RRM was found to be dispensable for RNA binding by TLS in vivo, suggesting that at any one time most of the interactions between TLS and RNA in the cell are not sequence specific. Analysis of inter specific heterokaryons formed between human and mouse or Xenopus cells revealed that TLS engages in rapid nucleocytoplasmic shuttling, a finding confirmed by the ability of anti-TLS antibodies to trap TLS when injected into the cytoplasm of HeLa cells. Cellular fractionation experiments suggest that TLS binds to RNA in both the nucleus and cytoplasm and support the hypothesis that TLS functions as a heterogeneous ribonuclear protein (hnRNP)-like chaperone of RNA. These findings are discussed in the context of the role altered forms of TLS play in cellular transformation.

scholarly journals Hel-N1: an autoimmune RNA-binding protein with specificity for 3' uridylate-rich untranslated regions of growth factor mRNAs.

1993 ◽  
Vol 13 (6) ◽  
pp. 3494-3504 ◽  
Author(s):  
T D Levine ◽  
F Gao ◽  
P H King ◽  
L G Andrews ◽  
J D Keene
Keyword(s):  
Rna Binding ◽  
Binding Protein ◽  
Rna Binding Protein ◽  
Selection Procedure ◽  
Vital Role ◽  
Untranslated Regions ◽  
Rna Recognition Motif ◽  
Rna Recognition ◽  
Binding Studies ◽  
Recognition Motifs

We have investigated the RNA binding specificity of Hel-N1, a human neuron-specific RNA-binding protein, which contains three RNA recognition motifs. Hel-N1 is a human homolog of Drosophila melanogaster elav, which plays a vital role in the development of neurons. A random RNA selection procedure revealed that Hel-N1 prefers to bind RNAs containing short stretches of uridylates similar to those found in the 3' untranslated regions (3' UTRs) of oncoprotein and cytokine mRNAs such as c-myc, c-fos, and granulocyte macrophage colony-stimulating factor. Direct binding studies demonstrated that Hel-N1 bound and formed multimers with c-myc 3' UTR mRNA and required, as a minimum, a specific 29-nucleotide stretch containing AUUUG, AUUUA, and GUUUUU. Deletion analysis demonstrated that a fragment of Hel-N1 containing 87 amino acids, encompassing the third RNA recognition motif, forms an RNA binding domain for the c-myc 3' UTR. In addition, Hel-N1 was shown to be reactive with autoantibodies from patients with paraneoplastic encephalomyelitis both before and after binding to c-myc mRNA.

scholarly journals Cip1 and Cip2 Are Novel RNA-Recognition-Motif Proteins That Counteract Csx1 Function during Oxidative Stress

2006 ◽  
Vol 17 (3) ◽  
pp. 1176-1183 ◽  
Author(s):  
Victoria Martín ◽  
Miguel A. Rodríguez-Gabriel ◽  
W. Hayes McDonald ◽  
Stephen Watt ◽  
John R. Yates ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Gene Expression ◽  
Stress Response ◽  
Protein Identification ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Bzip Transcription Factor ◽  
Rna Recognition Motif ◽  
Rna Recognition ◽  
Control Of Gene Expression

Eukaryotic cells reprogram their global patterns of gene expression in response to stress. Recent studies in Schizosaccharomyces pombe showed that the RNA-binding protein Csx1 plays a central role in controlling gene expression during oxidative stress. It does so by stabilizing atf1+ mRNA, which encodes a subunit of a bZIP transcription factor required for gene expression during oxidative stress. Here, we describe two related proteins, Cip1 and Cip2, that were identified by multidimensional protein identification technology (MudPIT) as proteins that coprecipitate with Csx1. Cip1 and Cip2 are cytoplasmic proteins that have RNA recognition motifs (RRMs). Neither protein is essential for viability, but a cip1Δ cip2Δ strain grows poorly and has altered cellular morphology. Genetic epistasis studies and whole genome expression profiling show that Cip1 and Cip2 exert posttranscriptional control of gene expression in a manner that is counteracted by Csx1. Notably, the sensitivity of csx1Δ cells to oxidative stress and their inability to induce expression of Atf1-dependent genes are partially rescued by cip1Δ and cip2Δ mutations. This study emphasizes the importance of a modulated mRNA stability in the eukaryotic stress response pathways and adds new information to the role of RNA-binding proteins in the oxidative stress response.

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