NMR Structural Study of TcUBP1, a Single RRM Domain Protein fromTrypanosoma cruzi:  Contribution of a β Hairpin to RNA Binding†,‡

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.

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A novel RRM domain protein is involved in the assembly of the tubulin cytoskeleton in Marsilea sperm cells

Developmental Biology ◽

10.1016/j.ydbio.2009.05.111 ◽

2009 ◽

Vol 331 (2) ◽

pp. 418

Author(s):

Corine M. Van der Weele ◽

Stephen M. Wolniak

Keyword(s):

Sperm Cells ◽

Rrm Domain ◽

Domain Protein

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A New Method for Determining Structure Ensemble: Application to a RNA Binding Di-Domain Protein

Biophysical Journal ◽

10.1016/j.bpj.2016.04.009 ◽

2016 ◽

Vol 110 (9) ◽

pp. 1943-1956 ◽

Cited By ~ 7

Author(s):

Wei Liu ◽

Jingfeng Zhang ◽

Jing-Song Fan ◽

Giancarlo Tria ◽

Gerhard Grüber ◽

...

Keyword(s):

Rna Binding ◽

New Method ◽

Domain Protein ◽

Structure Ensemble

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Homodimerization of RBPMS2 through a new RRM-interaction motif is necessary to control smooth muscle plasticity

Nucleic Acids Research ◽

10.1093/nar/gku692 ◽

2014 ◽

Vol 42 (15) ◽

pp. 10173-10184 ◽

Cited By ~ 16

Author(s):

Sébastien Sagnol ◽

Yinshan Yang ◽

Yannick Bessin ◽

Fréderic Allemand ◽

Ilona Hapkova ◽

...

Keyword(s):

Smooth Muscle ◽

Molecular Mechanisms ◽

Rna Binding ◽

Sequence Motif ◽

Rna Recognition Motif ◽

Muscle Plasticity ◽

Rrm Domain ◽

Bone Morphogenetic Protein Pathway ◽

Morphogenetic Protein

Abstract In vertebrates, smooth muscle cells (SMCs) can reversibly switch between contractile and proliferative phenotypes. This involves various molecular mechanisms to reactivate developmental signaling pathways and induce cell dedifferentiation. The protein RBPMS2 regulates early development and plasticity of digestive SMCs by inhibiting the bone morphogenetic protein pathway through its interaction with NOGGIN mRNA. RBPMS2 contains only one RNA recognition motif (RRM) while this motif is often repeated in tandem or associated with other functional domains in RRM-containing proteins. Herein, we show using an extensive combination of structure/function analyses that RBPMS2 homodimerizes through a particular sequence motif (D-x-K-x-R-E-L-Y-L-L-F: residues 39–51) located in its RRM domain. We also show that this specific motif is conserved among its homologs and paralogs in vertebrates and in its insect and worm orthologs (CPO and MEC-8, respectively) suggesting a conserved molecular mechanism of action. Inhibition of the dimerization process through targeting a conserved leucine inside of this motif abolishes the capacity of RBPMS2 to interact with the translational elongation eEF2 protein, to upregulate NOGGIN mRNA in vivo and to drive SMC dedifferentiation. Our study demonstrates that RBPMS2 possesses an RRM domain harboring both RNA-binding and protein-binding properties and that the newly identified RRM-homodimerization motif is crucial for the function of RBPMS2 at the cell and tissue levels.

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Over-expression of a conserved RNA-binding motif (RRM) domain (csRRM2) improves components ofBrassica napusyield by regulating cell size

Plant Breeding ◽

10.1111/j.1439-0523.2012.01998.x ◽

2012 ◽

Vol 131 (5) ◽

pp. 614-619 ◽

Cited By ~ 4

Author(s):

Weiwei Qi ◽

Fengqi Zhang ◽

Fan Sun ◽

Yongjuan Huang ◽

Rongzhan Guan ◽

...

Keyword(s):

Cell Size ◽

Rna Binding ◽

Binding Motif ◽

Over Expression ◽

Rrm Domain

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The key protein of endosomal mRNP transport Rrm4 binds translational landmark sites of cargo mRNAs

10.1101/418079 ◽

2018 ◽

Cited By ~ 1

Author(s):

Lilli Olgeiser ◽

Carl Haag ◽

Susan Boerner ◽

Jernej Ule ◽

Anke Busch ◽

...

Keyword(s):

Rna Binding ◽

Rna Binding Proteins ◽

Specific Binding ◽

Hyphal Growth ◽

Binding Motif ◽

Uv Crosslinking ◽

Rrm Domain ◽

Messenger Ribonucleoprotein ◽

Close Proximity ◽

Nucleotide Resolution

AbstractRNA-binding proteins (RBPs) determine spatiotemporal gene expression by mediating active transport and local translation of cargo mRNAs. Here, we cast a transcriptome-wide view on the transported mRNAs and cognate RBP binding sites during endosomal messenger ribonucleoprotein (mRNP) transport in Ustilago maydis. Using individual-nucleotide resolution UV crosslinking and immunoprecipitation (iCLIP), we compare the key transport RBP Rrm4 and the newly identified endosomal mRNP component Grp1 that is crucial to coordinate hyphal growth. Both RBPs bind predominantly in the 3’ untranslated region of thousands of shared cargo mRNAs, often in close proximity. Intriguingly, Rrm4 precisely binds at stop codons, which constitute landmark sites of translation, suggesting an intimate connection of mRNA transport and translation. Towards uncovering the code of recognition, we identify UAUG as specific binding motif of Rrm4 that is bound by its third RRM domain. Altogether, we provide first insights into the positional organisation of co-localising RBPs on individual cargo mRNAs.

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Structural basis for mRNA recognition by human RBM38

Biochemical Journal ◽

10.1042/bcj20190652 ◽

2020 ◽

Vol 477 (1) ◽

pp. 161-172 ◽

Cited By ~ 3

Author(s):

Kaiyue Qian ◽

Mengyu Li ◽

Junchao Wang ◽

Min Zhang ◽

Mingzhu Wang

Keyword(s):

Rna Binding ◽

Structural Information ◽

Protein Translation ◽

Effector Proteins ◽

Structural Basis ◽

Binding Property ◽

Rna Recognition ◽

Recognition Mechanism ◽

Rrm Domain ◽

Regulate Protein

RNA-binding protein RBM38 was reported to bind the mRNA of several p53-related genes through its RRM domain and to up-regulate or down-regulate protein translation by increasing mRNA stability or recruitment of other effector proteins. The recognition mechanism, however, for RNA-binding of RBM38 remains unclear. Here, we report the crystal structure of the RRM domain of human RBM38 in complex with a single-stranded RNA. Our structural and biological results revealed that RBM38 recognizes G(U/C/A)GUG sequence single-stranded RNA in a sequence-specific and structure-specific manner. Two phenylalanine stacked with bases of RNA were crucial for RNA binding, and a series of hydrogen bonds between the base atoms of RNA and main-chain or side-chain atoms of RBM38 determine the sequence-specific recognition. Our results revealed the RNA-recognition mechanism of human RBM38 and provided structural information for understanding the RNA-binding property of RBM38.

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UHM–ULM interactions in the RBM39–U2AF65 splicing-factor complex

Acta Crystallographica Section D Structural Biology ◽

10.1107/s2059798316001248 ◽

2016 ◽

Vol 72 (4) ◽

pp. 497-511 ◽

Cited By ~ 17

Author(s):

Galina A. Stepanyuk ◽

Pedro Serrano ◽

Eigen Peralta ◽

Carol L. Farr ◽

Herbert L. Axelrod ◽

...

Keyword(s):

Protein Interactions ◽

Rna Binding ◽

Splicing Factor ◽

Structural Basis ◽

Titration Calorimetry ◽

Protein Protein Interactions ◽

Rrm Domain ◽

Nmr Structures ◽

Binding Interface ◽

Cell Extracts

RNA-binding protein 39 (RBM39) is a splicing factor and a transcriptional co-activator of estrogen receptors and Jun/AP-1, and its function has been associated with malignant progression in a number of cancers. The C-terminal RRM domain of RBM39 belongs to the U2AF homology motif family (UHM), which mediate protein–protein interactions through a short tryptophan-containing peptide known as the UHM-ligand motif (ULM). Here, crystal and solution NMR structures of the RBM39-UHM domain, and the crystal structure of its complex with U2AF65-ULM, are reported. The RBM39–U2AF65 interaction was confirmed by co-immunoprecipitation from human cell extracts, by isothermal titration calorimetry and by NMR chemical shift perturbation experiments with the purified proteins. When compared with related complexes, such as U2AF35–U2AF65 and RBM39–SF3b155, the RBM39-UHM–U2AF65-ULM complex reveals both common and discriminating recognition elements in the UHM–ULM binding interface, providing a rationale for the known specificity of UHM–ULM interactions. This study therefore establishes a structural basis for specific UHM–ULM interactions by splicing factors such as U2AF35, U2AF65, RBM39 and SF3b155, and a platform for continued studies of intermolecular interactions governing disease-related alternative splicing in eukaryotic cells.

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A crucial RNA-binding lysine residue in the Nab3 RRM domain undergoes SET1 and SET3-responsive methylation

Nucleic Acids Research ◽

10.1093/nar/gkaa029 ◽

2020 ◽

Vol 48 (6) ◽

pp. 2897-2911 ◽

Cited By ~ 1

Author(s):

Kwan Yin Lee ◽

Anand Chopra ◽

Giovanni L Burke ◽

Ziyan Chen ◽

Jack F Greenblatt ◽

...

Keyword(s):

Rna Binding ◽

Rna Recognition ◽

Rna Sequences ◽

Rna Recognition Motifs ◽

Rrm Domain ◽

Lysine Residues ◽

Recognition Motifs ◽

Nascent Transcripts ◽

Target Rna

Abstract The Nrd1–Nab3–Sen1 (NNS) complex integrates molecular cues to direct termination of noncoding transcription in budding yeast. NNS is positively regulated by histone methylation as well as through Nrd1 binding to the initiating form of RNA PolII. These cues collaborate with Nrd1 and Nab3 binding to target RNA sequences in nascent transcripts through their RRM RNA recognition motifs. In this study, we identify nine lysine residues distributed amongst Nrd1, Nab3 and Sen1 that are methylated, suggesting novel molecular inputs for NNS regulation. We identify mono-methylation of one these residues (Nab3-K363me1) as being partly dependent on the H3K4 methyltransferase, Set1, a known regulator of NNS function. Moreover, the accumulation of Nab3-K363me1 is essentially abolished in strains lacking SET3, a SET domain containing protein that is positively regulated by H3K4 methylation. Nab3-K363 resides within its RRM and physically contacts target RNA. Mutation of Nab3-K363 to arginine (Nab3-K363R) decreases RNA binding of the Nab3 RRM in vitro and causes transcription termination defects and slow growth. These findings identify SET3 as a potential contextual regulator of Nab3 function through its role in methylation of Nab3-K363. Consistent with this hypothesis, we report that SET3 exhibits genetic activation of NAB3 that is observed in a sensitized context.

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