Biology subjects

Francisco-Velilla, R.

Publications and source records attributed to Francisco-Velilla, R..

3 recordsLinked to original sources

bioRxiv2022

Functional and structural deficiencies of Gemin5 variants associated with neurological disease

Dysfunction of RNA-binding proteins are often linked to a wide range of human disease in general, and particularly with neurological conditions. Gemin5 is a member of the survival of motor neuron (SMN) complex, a ribosome-binding protein and a translation reprogramming factor. Recently, pathogenic mutations in Gemin5 protein have been reported but the functional consequences of these variants remain elusive. Here we report functional and structural deficiencies associated with compound heterozygosity variants within the Gemin5 gene found in patients with neurodevelopmental disorders. These clinical variants are located in key domains of Gemin5, the tetratricopeptide repeat (TPR)-like dimerization module and the non-canonical RNA-binding site 1 (RBS1). We show that the TPR-like variants disrupt protein dimerization while the RBS1 variant confers protein instability. All mutants are defective in the interaction with protein networks involved in translation and RNA-driven pathways. Importantly, the TPR-like variants fail to associate with native ribosomes, abolishing the cap-dependent and selective translation control of Gemin5, and establishing a functional difference with the wild type protein. Our study provides a molecular basis of disease associated with malfunction of Gemin5 protein.

molecular biology↗

bioRxiv2019

A novel dimerization module in Gemin5 is critical for protein recruitment and translation control

The versatile multifunctional protein Gemin5 is involved in small nuclear ribonucleoproteins (snRNPs) assembly, ribosome binding, and translation control through distinct domains located at the protein ends. However, the structure and function of the central moiety of Gemin5 remained unknown. Here, we solved the crystal structure of an extended tetratricopeptide (TPR)-like domain in the middle region of Gemin5, demonstrating that it self-assembles into a canoe-shaped dimer. Mass spectrometry analysis shows that this dimerization module is functional in living cells and drives the interaction between p85, a viral-induced Gemin5 cleavage fragment, and the full-length Gemin5. In contrast, disruption of the dimerization surface by a point mutation in the TPR-like domain prevents this interaction and abrogates the translation enhancement induced by p85. The structural characterization of this unprecedented dimerization domain provides the mechanistic basis for a role of the middle region of Gemin5 as a key mediator of protein-protein interactions.\n\nHIGHLIGHTS- The crystal structure of a central region of Gemin5 reveals a novel dimerization domain\n- The proteolytic product of Gemin5 (p85) recruits the endogenous protein through the dimerization module\n- The dimerization capability of Gemin5 determines the factors recruited in human cells\n- Disruption of the dimerization domain impairs p85 ability to stimulate translation\n\n\n\n\nO_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC=\"FIGDIR/small/654111v1_ufig1.gif\" ALT=\"Figure 1\">\nView larger version (78K):\norg.highwire.dtl.DTLVardef@9d8e69org.highwire.dtl.DTLVardef@1f0a453org.highwire.dtl.DTLVardef@1b486c4org.highwire.dtl.DTLVardef@fbb5ea_HPS_FORMAT_FIGEXP M_FIG Graphical abstract\n\nC_FIG

molecular biology↗

bioRxiv2018

Rab1b and ARF5 are novel RNA-binding proteins involved in IRES-driven RNA localization

Internal ribosome entry site (IRES) elements are organized in domains that guide internal initiation of translation. Here we have combined proteomic and imaging analysis to study novel IRES interactors recognizing specific RNA structural subdomains. Besides known IRES-binding proteins, we identified novel factors belonging to networks involved in RNA and protein transport. Among those, Rab1b and ARF5, two components of the ER-Golgi, revealed direct binding to IRES transcripts. However, these proteins exert different effects on translation. While a dominant-negative mutant of Rab1b decreased IRES function, ARF5 silencing stimulated IRES activity. RNA FISH studies revealed novel features of the IRES element. First, IRES-RNA formed clusters within the cell cytoplasm, whereas cap-RNA displayed disperse punctuated distribution. Second, the IRES-driven RNA colocalized with ARF5 and Rab1b, but not with the dominant-negative of Rab1b. Thus, our data suggest a role for domain 3 of the IRES in RNA localization around ER-Golgi, a ribosome-rich cellular compartment.

molecular biology↗