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

Sagnol S, Yang Y, Bessin Y, Allemand F, Hapkova I, Notarnicola C, Guichou JF, Faure S, Labesse G, de Santa Barbara P - Nucleic Acids Res. (2014)

Bottom Line: 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.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.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.

View Article: PubMed Central - PubMed

Affiliation: INSERM U1046, Université Montpellier 1, Université Montpellier 2, 34295 Montpellier, France.

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Dimeric structure of RBPMS2 in solution. (A, B) RBPMS2, CPO and MEC-8 proteins harbor evolutionary conserved RNA recognition motif (RRM) domain. (A) Phylogenetic tree using human RBPMS2 protein as query (in red). (B) Phylogenetic tree using the RBPMS2 RRM domain (amino acids from 27 to 117) as query (in red). After each leaf end follow the protein names, the species names (H.sa.: Human; M.mu.: Mus musculus; G.ga.: Gallus gallus; D.me.: Drosophila melanogaster; C.el.: Caenhorabditis elegans), the E-value and the percentage of identity regarding the query. The accession number of each protein is presented on the right column of the figure. Scale bar represents evolution distance of leaf branches (A.U.). The figure was prepared using iTOL (http://itol.embl.de). (C) Sequence-structure alignment of the RRM domain in RBPMS2 homologs. The N-terminus (residues 27–117) of human RBPMS2 was aligned with the corresponding segments in the sequences of RBPMS2, CPO or MEC-8 from different metazoan species (human: Homo sapiens; danre: Dano rerio; galga: Gallus gallus; xenle: Xenopus leavis; drome: Drosophila melanogaster; apime: Apis mellifica; culpi: Culex pipens; Bruma: Brugia malayi; caeel: Caenorhabditis elegans). Arrowheads indicate the predicted RNA binding site and asterisks the newly identified dimerization interface. Residue numbering corresponds to the human RBPMS2 sequence. The figure was made using ESPRIPT (http/espript.ibcp.fr). Overall structure of wild-type human RBMPS2-Nter homodimers (D) and detailed view of the dimerization interface (E) seen in a ribbon diagram. The secondary structure and loops involved in the dimerization are shown as red-to-blue (subunit A) and orange (subunit B) ribbons. Red indicates highly conserved amino acid where blue labels less conserved residues. The side chains that stabilize the dimeric interface are shown as sticks using the CPK color convention. Residues are numbered according to the human RBMPS2 sequence. The figures were prepared using Pymol (http://pymol.sourceforge.net). (F) Experimental small-angle X-ray scattering curve (logarithm of intensity in arbitrary units as a function of the momentum transfer range s in Å−1) for RBPMS2-Nter measured at 1.9 mg/ml (green crosses), with its fitting theoretical curve (red continuous line) back-calculated from the RBPMS2-Nter NMR structure (Supplementary Figure S2). Blue dots represent the relative error bound. The χ2 value of the fit is 1.115.
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Figure 1: Dimeric structure of RBPMS2 in solution. (A, B) RBPMS2, CPO and MEC-8 proteins harbor evolutionary conserved RNA recognition motif (RRM) domain. (A) Phylogenetic tree using human RBPMS2 protein as query (in red). (B) Phylogenetic tree using the RBPMS2 RRM domain (amino acids from 27 to 117) as query (in red). After each leaf end follow the protein names, the species names (H.sa.: Human; M.mu.: Mus musculus; G.ga.: Gallus gallus; D.me.: Drosophila melanogaster; C.el.: Caenhorabditis elegans), the E-value and the percentage of identity regarding the query. The accession number of each protein is presented on the right column of the figure. Scale bar represents evolution distance of leaf branches (A.U.). The figure was prepared using iTOL (http://itol.embl.de). (C) Sequence-structure alignment of the RRM domain in RBPMS2 homologs. The N-terminus (residues 27–117) of human RBPMS2 was aligned with the corresponding segments in the sequences of RBPMS2, CPO or MEC-8 from different metazoan species (human: Homo sapiens; danre: Dano rerio; galga: Gallus gallus; xenle: Xenopus leavis; drome: Drosophila melanogaster; apime: Apis mellifica; culpi: Culex pipens; Bruma: Brugia malayi; caeel: Caenorhabditis elegans). Arrowheads indicate the predicted RNA binding site and asterisks the newly identified dimerization interface. Residue numbering corresponds to the human RBPMS2 sequence. The figure was made using ESPRIPT (http/espript.ibcp.fr). Overall structure of wild-type human RBMPS2-Nter homodimers (D) and detailed view of the dimerization interface (E) seen in a ribbon diagram. The secondary structure and loops involved in the dimerization are shown as red-to-blue (subunit A) and orange (subunit B) ribbons. Red indicates highly conserved amino acid where blue labels less conserved residues. The side chains that stabilize the dimeric interface are shown as sticks using the CPK color convention. Residues are numbered according to the human RBMPS2 sequence. The figures were prepared using Pymol (http://pymol.sourceforge.net). (F) Experimental small-angle X-ray scattering curve (logarithm of intensity in arbitrary units as a function of the momentum transfer range s in Å−1) for RBPMS2-Nter measured at 1.9 mg/ml (green crosses), with its fitting theoretical curve (red continuous line) back-calculated from the RBPMS2-Nter NMR structure (Supplementary Figure S2). Blue dots represent the relative error bound. The χ2 value of the fit is 1.115.

Mentions: To identify the structural features that are responsible for RBPMS2 function, we analyzed RBPMS2 sequence and its predicted structure. Basic Local Alignment Search Tool (BLAST) searches using human RBPMS2 as a query (NP_919248.1) identified its vertebrate orthologs with high sequence conservation (E-values ranging from 10−153 to 10−114) and also its closely related paralog RBPMS1 (E-values from 10−94 to 10−74) (Figure 1A). This analysis also highlighted the strong similarities of human RBPMS2 and insect CPO (E-values from 10−40 to 10−38) and of both RBPMS2 and CPO sequences with C. elegans MEC-8 (∼10−30). These three proteins were the first RRM-containing proteins detected by BLAST, suggesting a common evolutionary history and some functional homology. Indeed, no other human RNA-binding protein showed such a strong similarity with RBPMS2 (see the snRNP proteins with E-values ∼10−3). Similar results were obtained when we used only the sequence of the RRM domain of RBPMS2, suggesting that RBPMS2, CPO and MEC-8 form an original subfamily of RRM-containing proteins (Figure 1B). This family also includes their paralogs (RBPMS1) that share highly similar RRM sequences in vertebrates. Their common N-terminal RRM domains (90-residue long) share 60–69% of sequence identity. The rest of the sequence showed significant amino acid composition bias toward small and/or polar residues (G, A, S, T, P, Q) and rapid divergence among the closely homologous sequences while no significant similarities could be found with known protein structures. In agreement, disordered segments are predicted by various bioinformatic tools implemented in the meta-server MetaDisorder (32) around the RRM domain (data not shown).


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

Sagnol S, Yang Y, Bessin Y, Allemand F, Hapkova I, Notarnicola C, Guichou JF, Faure S, Labesse G, de Santa Barbara P - Nucleic Acids Res. (2014)

Dimeric structure of RBPMS2 in solution. (A, B) RBPMS2, CPO and MEC-8 proteins harbor evolutionary conserved RNA recognition motif (RRM) domain. (A) Phylogenetic tree using human RBPMS2 protein as query (in red). (B) Phylogenetic tree using the RBPMS2 RRM domain (amino acids from 27 to 117) as query (in red). After each leaf end follow the protein names, the species names (H.sa.: Human; M.mu.: Mus musculus; G.ga.: Gallus gallus; D.me.: Drosophila melanogaster; C.el.: Caenhorabditis elegans), the E-value and the percentage of identity regarding the query. The accession number of each protein is presented on the right column of the figure. Scale bar represents evolution distance of leaf branches (A.U.). The figure was prepared using iTOL (http://itol.embl.de). (C) Sequence-structure alignment of the RRM domain in RBPMS2 homologs. The N-terminus (residues 27–117) of human RBPMS2 was aligned with the corresponding segments in the sequences of RBPMS2, CPO or MEC-8 from different metazoan species (human: Homo sapiens; danre: Dano rerio; galga: Gallus gallus; xenle: Xenopus leavis; drome: Drosophila melanogaster; apime: Apis mellifica; culpi: Culex pipens; Bruma: Brugia malayi; caeel: Caenorhabditis elegans). Arrowheads indicate the predicted RNA binding site and asterisks the newly identified dimerization interface. Residue numbering corresponds to the human RBPMS2 sequence. The figure was made using ESPRIPT (http/espript.ibcp.fr). Overall structure of wild-type human RBMPS2-Nter homodimers (D) and detailed view of the dimerization interface (E) seen in a ribbon diagram. The secondary structure and loops involved in the dimerization are shown as red-to-blue (subunit A) and orange (subunit B) ribbons. Red indicates highly conserved amino acid where blue labels less conserved residues. The side chains that stabilize the dimeric interface are shown as sticks using the CPK color convention. Residues are numbered according to the human RBMPS2 sequence. The figures were prepared using Pymol (http://pymol.sourceforge.net). (F) Experimental small-angle X-ray scattering curve (logarithm of intensity in arbitrary units as a function of the momentum transfer range s in Å−1) for RBPMS2-Nter measured at 1.9 mg/ml (green crosses), with its fitting theoretical curve (red continuous line) back-calculated from the RBPMS2-Nter NMR structure (Supplementary Figure S2). Blue dots represent the relative error bound. The χ2 value of the fit is 1.115.
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Related In: Results  -  Collection

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Show All Figures
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Figure 1: Dimeric structure of RBPMS2 in solution. (A, B) RBPMS2, CPO and MEC-8 proteins harbor evolutionary conserved RNA recognition motif (RRM) domain. (A) Phylogenetic tree using human RBPMS2 protein as query (in red). (B) Phylogenetic tree using the RBPMS2 RRM domain (amino acids from 27 to 117) as query (in red). After each leaf end follow the protein names, the species names (H.sa.: Human; M.mu.: Mus musculus; G.ga.: Gallus gallus; D.me.: Drosophila melanogaster; C.el.: Caenhorabditis elegans), the E-value and the percentage of identity regarding the query. The accession number of each protein is presented on the right column of the figure. Scale bar represents evolution distance of leaf branches (A.U.). The figure was prepared using iTOL (http://itol.embl.de). (C) Sequence-structure alignment of the RRM domain in RBPMS2 homologs. The N-terminus (residues 27–117) of human RBPMS2 was aligned with the corresponding segments in the sequences of RBPMS2, CPO or MEC-8 from different metazoan species (human: Homo sapiens; danre: Dano rerio; galga: Gallus gallus; xenle: Xenopus leavis; drome: Drosophila melanogaster; apime: Apis mellifica; culpi: Culex pipens; Bruma: Brugia malayi; caeel: Caenorhabditis elegans). Arrowheads indicate the predicted RNA binding site and asterisks the newly identified dimerization interface. Residue numbering corresponds to the human RBPMS2 sequence. The figure was made using ESPRIPT (http/espript.ibcp.fr). Overall structure of wild-type human RBMPS2-Nter homodimers (D) and detailed view of the dimerization interface (E) seen in a ribbon diagram. The secondary structure and loops involved in the dimerization are shown as red-to-blue (subunit A) and orange (subunit B) ribbons. Red indicates highly conserved amino acid where blue labels less conserved residues. The side chains that stabilize the dimeric interface are shown as sticks using the CPK color convention. Residues are numbered according to the human RBMPS2 sequence. The figures were prepared using Pymol (http://pymol.sourceforge.net). (F) Experimental small-angle X-ray scattering curve (logarithm of intensity in arbitrary units as a function of the momentum transfer range s in Å−1) for RBPMS2-Nter measured at 1.9 mg/ml (green crosses), with its fitting theoretical curve (red continuous line) back-calculated from the RBPMS2-Nter NMR structure (Supplementary Figure S2). Blue dots represent the relative error bound. The χ2 value of the fit is 1.115.
Mentions: To identify the structural features that are responsible for RBPMS2 function, we analyzed RBPMS2 sequence and its predicted structure. Basic Local Alignment Search Tool (BLAST) searches using human RBPMS2 as a query (NP_919248.1) identified its vertebrate orthologs with high sequence conservation (E-values ranging from 10−153 to 10−114) and also its closely related paralog RBPMS1 (E-values from 10−94 to 10−74) (Figure 1A). This analysis also highlighted the strong similarities of human RBPMS2 and insect CPO (E-values from 10−40 to 10−38) and of both RBPMS2 and CPO sequences with C. elegans MEC-8 (∼10−30). These three proteins were the first RRM-containing proteins detected by BLAST, suggesting a common evolutionary history and some functional homology. Indeed, no other human RNA-binding protein showed such a strong similarity with RBPMS2 (see the snRNP proteins with E-values ∼10−3). Similar results were obtained when we used only the sequence of the RRM domain of RBPMS2, suggesting that RBPMS2, CPO and MEC-8 form an original subfamily of RRM-containing proteins (Figure 1B). This family also includes their paralogs (RBPMS1) that share highly similar RRM sequences in vertebrates. Their common N-terminal RRM domains (90-residue long) share 60–69% of sequence identity. The rest of the sequence showed significant amino acid composition bias toward small and/or polar residues (G, A, S, T, P, Q) and rapid divergence among the closely homologous sequences while no significant similarities could be found with known protein structures. In agreement, disordered segments are predicted by various bioinformatic tools implemented in the meta-server MetaDisorder (32) around the RRM domain (data not shown).

Bottom Line: 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.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.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.

View Article: PubMed Central - PubMed

Affiliation: INSERM U1046, Université Montpellier 1, Université Montpellier 2, 34295 Montpellier, France.

Show MeSH
Related in: MedlinePlus