Biology subjects

Vandevenne, M.

Publications and source records attributed to Vandevenne, M..

3 recordsLinked to original sources

bioRxiv2024

The Arabidopsis RS2Z32 and RS2Z33 proteins are dynamic splicing factors whose RNA recognition motif (RRM) domain contributes to protein-protein and protein-RNA interactions

The Arabidopsis splicing factors arginine/serine-rich zinc knuckle-containing proteins 32 and 33 (RS2Z32 and RS2Z33) are plant-specific members of the SR family whose molecular functions received little attention. Here, we characterized both RS2Z32 and RS2Z33 by examining their expression profile at different stages of development and their spatial cellular distribution, as well as the contribution of their domains in the establishment of protein-protein interactions and RNA binding specificity. We report that the RS2Z32 and RS2Z33 promoters are ubiquitously active during vegetative and reproductive growth, and that both RS2Z splicing factors localize in the nucleus (except the nucleolus). We show that the C-terminal arginine/serine-rich (RS) domain, but not the serine/proline-rich (SP) extension, is a determinant of nuclear localization, which likely requires phosphoresidues putatively phosphorylated by kinases of the SRPK family. We demonstrate that their RNA recognition motif (RRM) domain specifically binds pyrimidine-rich RNA motifs via three residues (Y14, Y46, F48), and is also involved in protein-protein interactions with at least three SR proteins, namely SR45, SCL30, and SR34. Finally, we show that mutations in RNA-binding domains (i.e. RRM and zinc knuckles, ZnKs) affect the nucleocytoplasmic dynamics of both RS2Z proteins. Our findings provide molecular evidence for the involvement of plant-specific SR splicing factors into the regulation of the splicing process. HighlightSpecific domains of the Arabidopsis RS2Z splicing factors contribute to their nuclear localization, nucleocytoplasmic dynamics, and ability to contact protein partners and specific pyrimidine-rich RNA motifs.

plant biology↗

bioRxiv2023

The Arabidopsis SR45 splicing factor bridges the splicing machinery and the exon-exon junction complex

The Arabidopsis splicing factor serine/arginine-rich 45 (SR45) contributes to several biological processes. The sr45-1 loss-of-function mutant exhibits delayed root development, late flowering, unusual numbers of floral organs, shorter siliques with decreased seed sets, narrower leaves and petals, and altered metal distribution. SR45 bears a unique RNA recognition motif (RRM) flanked by one serine/arginine-rich (RS) domain on both sides. Here, we studied the function of each of SR45 domains by examining their involvement in: (i) the spatial distribution of SR45, (ii) the establishment of a protein-protein interaction network including spliceosomal and exon-exon junction complex (EJC) components, and (iii) the RNA binding specificity. We report that the endogenous SR45 promoter is active during vegetative and reproductive growth, and that the SR45 protein localizes in the nucleus. We demonstrate that the C-terminal arginine/serine-rich domain is a determinant of nuclear localization. We show that the SR45 RNA recognition motif (RRM) domain specifically binds purine-rich RNA motifs via three residues (H101, H141, Y143), and is also involved in protein-protein interactions. We further show that SR45 bridges both mRNA splicing and surveillance machineries as a partner of EJC core components and peripheral factors, which requires phosphoresidues likely phosphorylated by kinases from both CLK and SRPK families. Our findings provide insights into the contribution of each SR45 domain to both spliceosome and EJC assemblies. HighlightThe contribution of the Arabidopsis SR45 splicing factor individual domains to its nuclear localization, ability to contact in planta novel protein partners and specifically bind RNA motifs was examined.

plant biology↗

bioRxiv2022

Development of nanobodies as theranostic agents against CMY-2-like class C β-lactamases

Soluble single-domain fragments derived from the unique variable region of camelid heavy-chain antibodies (VHHs) against enzymes may behave as potent inhibitors. The immunization of alpacas with the CMY-2 {beta}-lactamase led to the isolation of three VHHs that specifically recognized and inhibited CMY-2. The structure of the complex VHH cAbCMY-2(254)/CMY-2 was determined by X-ray crystallography. We showed that the epitope is close to the active site and that the CDR3 of the VHH protrudes in the catalytic site. The {beta}-lactamase inhibition was found to follow a mixed profile with a predominant non-competitive component. The three isolated VHHs recognized overlapping epitopes since they behaved as competitive binder. Our study identified a binding site that can be targeted by a new class of {beta}-lactamases inhibitors designed with the help of a peptidomimetic approach. Furthermore, the use of mono or bivalent VHH and rabbit polyclonal anti-CMY-2 antibodies enable the development of the first generation of ELISA test for the detection of CMY-2 produced by resistant bacteria. IMPORTANCEThe still increasing antimicrobial resistance in human clinic or veterinary medicine is a major threat for modern chemotherapy. Beside the major caution in the use of current antibiotics, it is important to develop new classes of antibiotics. This work was focused on {beta}-lactamases that are the enzymes involved in the hydrolysis of the major class of antibiotics, the {beta}-lactam compounds. We selected camelid antibodies that inhibit CMY-2, a class C {beta}-lactamase produced by bacteria isolated from the veterinary and human settings. We characterized the conformational epitope present in CMY-2 in order to create a new family of inhibitors based on the paratope of the antibody. Finally, we designed a primary version of a detection system based on an ELISA using VHH and polyclonal antibodies.

biochemistry↗