Une étape clé dans la maturation des pré-ARNms est le clivage et la polyadénylation que ceux-ci subissent sur leur extrémité 3’. Chez les métazoaires, le complexe CstF (Cleavage stimulation Factor) reconnaît une région de l’ARNm riche en U et U/G et stabilise le complexe CPSF (Cleavage Polyadenylation Stimulating Factor) sur le site de polyadénylation. Nous avons déterminé la structure cristallographique du domaine N-terminal d’une des trois sous-unités de CstF, CstF-50. Ce domaine forme un homodimère compact et présente deux surfaces identiques conservées dérivées de la formation du dimère. La structure dimérique de CstF-50 est en accord avec le modèle hexamèrique du complexe. L’homologue de CstF chez la levure, CF IA (Cleavage/polyadenylation Factor IA), est impliqué dans les réactions de clivage et polyadénylation de la maturation 3’. Nous avons reconstitué le complexe entier ‘in vitro’ et résolu la structure en solution par RMN des régions minimales impliquées dans l’interaction des sous-unités Rna14p et Rna15p. Pour la formation de l’hétérodimère, la région C-terminale de Rna14p, que nous avons appelé domaine « monkeytail », s’entrelace intimement avec la région « hinge » de Rna15p. La présence de ces deux domaines chez leurs homologues de mammifères, CstF-77 et CstF-64, suggère la conservation de ce type d’organisation entre ces deux sous-unités à travers les espèces. / The removal of the 3’ region of pre-mRNA followed by polyadenylation is a key step in mRNA maturation. In metazoa, Cleavage stimulation Factor (CstF) recognizes U and G/U rich cis-acting RNA sequence elements through its 64kDa subunit and helps stabilize the Cleavage Polyadenylation Stimulating Factor (CPSF) complex at the polyadenylation site. We describe the crystal structure of the N-terminal domain of the CstF-50 subunit. Through highly conserved residues, CstF-50 forms a compact homodimer that exposes two geometrically opposite and identical conserved surfaces. Together with prior data, the structure of the CstF-50 homodimerization domain supports a hexameric model of CstF. The yeast homologue of CstF is the Cleavage/polyadenylation Factor IA (CF IA) complex and is involved in both the cleavage and polyadenylation of pre-mRNA. We have reconstituted ‘in vitro’ the overall complex and also solved the solution structure of one of the inter-subunit regions, specifically the heterodimer involving peptides from Rna14p and Rna15p. Upon binding, a short C-terminal region from Rna14p wraps intimately within the central hinge domain from Rna15p. Conservation of residues reveals that the structural tethering is preserved in the homologous mammalian proteins. / La maduración 3’ del pre-ARNm es un proceso clave de la expresión génica que incluye el corte y la poliadenilación del extremo 3’ libre del pre-ARNm. En metazoos, el complejo CstF (Cleavage stimulation Factor) reconoce una secuencia del pre-ARNm rica en U y G/U y permite la estabilización del complejo CPSF (Cleavage Polyadenylation Stimulating Factor) en el sitio de poliadenilación. Hemos descrito la estructura cristalina del dominio N-terminal de una de las tres subunidades de CstF, CstF-50. La estructura ha revelado la organización de la proteína en un dímero compacto y conservado entre las especies. Dos zonas idénticas conservadas se encuentran expuestas a ambos lados de la superficie estructural. Nuestros resultados corroboran así la hipótesis sobre el modelo hexamérico del complejo CstF. CF IA (Cleavage/ polyadenylation Factor IA), el homólogo de CstF en levaduras, interviene en las dos etapas de la maduración 3’. Las bases para la reconstitución del factor CF IA ‘in vitro’ han sido establecidas. Al mismo tiempo, hemos resuelto la estructura del subcomplejo formado por las regiones de interacción de Rna14p y de Rna15p en solución mediante RMN. En el heterodímero, las dos proteínas forman una entidad única a través de la región C-terminal de Rna14p, dominio “monkeytail”, y el dominio “hinge” de Rna15p, quedando las hélices de la dos proteínas entrelazadas. La localización de estos dominios en sus homólogos mamíferos, CstF-77 et CstF-64, sugiere que este tipo de organización está conservada entre las especies.

3’-end modification by polyadenylation is a ubiquitous feature of almost all eukaryotic mRNA species and is catalyzed by a consortium of enzymes, the polyadenylation factors. Poly(A) polymerase (PAP), the enzyme catalyzing the addition of adenosine residues during the polyadenylation stage, exists in four isoforms within Arabidopsis. In silico and yeast two-hybrid studies showed that PAP1 has unique expression and interaction pattern in Arabidopsis, suggesting non-canonical functions of PAP1. Its exclusive interaction with PAP4 has not been reported in other living systems until now and hints at a difference in polyadenylation in plants with respect to mammals and yeast. Cleavage Stimulation Factor (CstF), a heterotrimeric complex of the polyadenylation factors CstF50, CstF64 and CstF77, plays a role largely in cleavage of pre-mRNA. This study highlights some aspects of the Arabidopsis homologs of CstF64 and CstF77, central to various cellular processes other than nuclear polyadenylation. In silico studies showed an elevated expression of CstF64 in the pollen while that of CstF77 remained fairly low. Yeast two-hybrid assays indicated a novel kind of interaction of CstF64 with Fip1(V). It is also speculated from sub-cellular localization techniques by agroinfiltration in tobacco leaves that CstF64 localizes in the cytoplasm and CstF77 in the nucleus, as found for the orthologs of CstF77 in other systems.