scholarly journals Two distinct binding modes provide the RNA binding protein RbFox with extraordinary sequence specificity

2021 ◽  
Author(s):  
Xuan Ye ◽  
Wen Yang ◽  
Soon Yi ◽  
Yanan Zhao ◽  
Fan Yang ◽  
...  
Keyword(s):  
Binding Site ◽  
Protein Interactions ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Rna Recognition Motif ◽  
Rna Recognition ◽  
Sequence Specificity ◽  
Binding Modes ◽  
Nmr Studies ◽  
Second Mode

The specificity of RNA-binding proteins for their target sequences varies considerably. Yet, it is not understood how certain proteins achieve markedly higher sequence specificity than most others. Here we show that the RNA Recognition Motif of RbFox accomplishes extraordinary sequence specificity by employing functionally and structurally distinct binding modes. Affinity measurements of RbFox for all binding site variants reveal the existence of two different binding modes. The first exclusively binds the cognate and a closely related RNA variant with high affinity. The second mode accommodates all other RNAs with greatly reduced affinity, thereby imposing large thermodynamic penalties on even near-cognate sequences. NMR studies indicate marked structural differences between the two binding modes, including large conformational rearrangements distant from the RNA binding site. Distinct binding modes by a single RNA binding module explain extraordinary sequence selectivity and reveal an unknown layer of functional diversity, cross talk and regulation for RNA-protein interactions.

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.

scholarly journals RNA-Binding Domain Proteins in Kinetoplastids: a Comparative Analysis

2005 ◽  
Vol 4 (12) ◽  
pp. 2106-2114 ◽  
Author(s):  
Javier De Gaudenzi ◽  
Alberto C. Frasch ◽  
Christine Clayton
Keyword(s):  
Leishmania Major ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Binding Protein ◽  
Rna Recognition Motif ◽  
Rna Recognition ◽  
Amino Acid Residues ◽  
Regulatory Pathways ◽  
Obligate Intracellular ◽  
Posttranscriptional Processing

ABSTRACT RNA-binding proteins are important in many aspects of RNA processing, function, and destruction. One class of such proteins contains the RNA recognition motif (RRM), which consists of about 90 amino acid residues, including the canonical RNP1 octapeptide: (K/R)G(F/Y)(G/A)FVX(F/Y). We used a variety of homology searches to classify all of the RRM proteins of the three kinetoplastids Trypanosoma brucei, Trypanosoma cruzi, and Leishmania major. All three organisms have similar sets of RRM-containing protein orthologues, suggesting common posttranscriptional processing and regulatory pathways. Of the 75 RRM proteins identified in T. brucei, only 13 had clear homologues in other eukaryotes, although 8 more could be given putative functional assignments. A comparison with the 18 RRM proteins of the obligate intracellular parasite Encephalitozoon cuniculi revealed just 3 RRM proteins which appear to be conserved at the primary sequence level throughout eukaryotic evolution: poly(A) binding protein, the rRNA-processing protein MRD1, and the nuclear cap binding protein.

scholarly journals CUS2, a Yeast Homolog of Human Tat-SF1, Rescues Function of Misfolded U2 through an Unusual RNA Recognition Motif

1998 ◽  
Vol 18 (9) ◽  
pp. 5000-5009 ◽  
Author(s):  
Dong Yan ◽  
Rhonda Perriman ◽  
Haller Igel ◽  
Kenneth J. Howe ◽  
Megan Neville ◽  
...  
Keyword(s):  
Rna Binding ◽  
Rna Binding Proteins ◽  
Normal Activity ◽  
Binding Activity ◽  
Rna Recognition Motif ◽  
Rna Recognition ◽  
Recognition Motif ◽  
Yeast Homolog

ABSTRACT A screen for suppressors of a U2 snRNA mutation identified CUS2, an atypical member of the RNA recognition motif (RRM) family of RNA binding proteins. CUS2 protein is associated with U2 RNA in splicing extracts and interacts with PRP11, a subunit of the conserved splicing factor SF3a. Absence of CUS2 renders certain U2 RNA folding mutants lethal, arguing that a normal activity of CUS2 is to help refold U2 into a structure favorable for its binding to SF3b and SF3a prior to spliceosome assembly. Both CUS2 function in vivo and the in vitro RNA binding activity of CUS2 are disrupted by mutation of the first RRM, suggesting that rescue of misfolded U2 involves the direct binding of CUS2. Human Tat-SF1, reported to stimulate Tat-specific, transactivating region-dependent human immunodeficiency virus transcription in vitro, is structurally similar to CUS2. Anti-Tat-SF1 antibodies coimmunoprecipitate SF3a66 (SAP62), the human homolog of PRP11, suggesting that Tat-SF1 has a parallel function in splicing in human cells.

scholarly journals An autoinhibitory intramolecular interaction proof-reads RNA recognition by the essential splicing factor U2AF2

2020 ◽  
Vol 117 (13) ◽  
pp. 7140-7149 ◽  
Author(s):  
Hyun-Seo Kang ◽  
Carolina Sánchez-Rico ◽  
Stefanie Ebersberger ◽  
F. X. Reymond Sutandy ◽  
Anke Busch ◽  
...  
Keyword(s):  
Rna Binding ◽  
Rna Binding Proteins ◽  
Intramolecular Interaction ◽  
Intramolecular Interactions ◽  
Rna Recognition Motif ◽  
Rna Recognition ◽  
Rna Motifs ◽  
Intrinsically Disordered ◽  
Molecular Features

The recognition of cis-regulatory RNA motifs in human transcripts by RNA binding proteins (RBPs) is essential for gene regulation. The molecular features that determine RBP specificity are often poorly understood. Here, we combined NMR structural biology with high-throughput iCLIP approaches to identify a regulatory mechanism for U2AF2 RNA recognition. We found that the intrinsically disordered linker region connecting the two RNA recognition motif (RRM) domains of U2AF2 mediates autoinhibitory intramolecular interactions to reduce nonproductive binding to weak Py-tract RNAs. This proofreading favors binding of U2AF2 at stronger Py-tracts, as required to define 3′ splice sites at early stages of spliceosome assembly. Mutations that impair the linker autoinhibition enhance the affinity for weak Py-tracts result in promiscuous binding of U2AF2 along mRNAs and impact on splicing fidelity. Our findings highlight an important role of intrinsically disordered linkers to modulate RNA interactions of multidomain RBPs.

scholarly journals Involvement of a tissue-specific RNA recognition motif protein in Drosophila spermatogenesis.

1997 ◽  
Vol 17 (5) ◽  
pp. 2708-2715 ◽  
Author(s):  
S R Haynes ◽  
M T Cooper ◽  
S Pype ◽  
D T Stolow
Keyword(s):  
Posttranscriptional Regulation ◽  
Rna Binding ◽  
Protein Gene ◽  
Germ Line ◽  
Rna Binding Proteins ◽  
Regulation Of Gene Expression ◽  
Rna Recognition Motif ◽  
Rna Recognition ◽  
Two Dimensional Gel Electrophoresis ◽  
Recognition Motif

RNA binding proteins mediate posttranscriptional regulation of gene expression via their roles in nuclear and cytoplasmic mRNA metabolism. Many of the proteins involved in these processes have a common RNA binding domain, the RNA recognition motif (RRM). We have characterized the Testis-specific RRM protein gene (Tsr), which plays an important role in spermatogenesis in Drosophila melanogaster. Disruption of Tsr led to a dramatic reduction in male fertility due to the production of spermatids with abnormalities in mitochondrial morphogenesis. Tsr is located on the third chromosome at 87F, adjacent to the nuclear pre-mRNA binding protein gene Hrb87F. A 1.7-kb Tsr transcript was expressed exclusively in the male germ line. It encoded a protein containing two RRMs similar to those found in HRB87F as well as a unique C-terminal domain. TSR protein was located in the cytoplasm of spermatocytes and young spermatids but was absent from mature sperm. The cellular proteins expressed in premeiotic primary spermatocytes from Tsr mutant and wild-type males were assessed by two-dimensional gel electrophoresis. Lack of TSR resulted in the premature expression of a few proteins prior to meiosis; this was abolished by a transgenic copy of Tsr. These data demonstrate that TSR negatively regulated the expression of some testis proteins and, in combination with its expression pattern and subcellular localization, suggest that TSR regulates the stability or translatability of some mRNAs during spermatogenesis.

scholarly journals A Negative Coregulator for the Human ER

2002 ◽  
Vol 16 (3) ◽  
pp. 459-468 ◽  
Author(s):  
John D. Norris ◽  
Daju Fan ◽  
Andrea Sherk ◽  
Donald P. McDonnell
Keyword(s):  
Transcriptional Activity ◽  
Cellular Proliferation ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Partial Agonist ◽  
Rna Recognition Motif ◽  
Rna Recognition ◽  
Agonist Activity ◽  
Partial Agonist Activity ◽  
Recognition Motif

Abstract ERα is a ligand-activated transcription factor and a key regulator of the processes involved in cellular proliferation and differentiation. In addition, aberrant ERα activity is linked to several pathological conditions including breast cancer. A complex network of coregulatory proteins is largely believed to determine the transcriptional activity of ERα. We report here the isolation of a protein, denoted RTA for repressor of tamoxifen transcriptional activity, which contains an RNA recognition motif and interacts with the receptor N-terminal activation domain. RTA interacts with RNA in vitro, and its overexpression inhibits the partial agonist activity manifest by the antiestrogen tamoxifen while minimally affecting E2-activated transcription. Mutation of the RNA recognition motif alters RNA binding specificity and results in a dominant negative form of RTA that leads to derepression of ERα transcriptional activity, allowing all classes of antiestrogens to manifest partial agonist activity and enhancing agonist efficacy. These findings suggest a role for RNA binding proteins as coregulatory factors of the nuclear receptor family and reveal a novel mechanism by which antiestrogens can manifest agonist activities in some tissues.

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