La traduction représente un processus central au sein de la cellule, elle assure le transfert de l’information génétique de l’ARNm vers les protéines. De nombreux acteurs y sont impliqués directement ou indirectement et parmi eux, chez les eucaryotes, la petite protéine Trm112. Celle-ci participe à la modification de plusieurs acteurs directs en interagissant et en activant quatre MTases. Le facteur de terminaison eRF1 est méthylé par le complexe Mtq2-Trm112, l’ARNr 18S par Bud23-Trm112 et certains ARNt par les complexes Trm9-Trm112 et Trm11-Trm112. Au cours de ce travail, les structures cristallographiques de Trm9-Trm112 et de Bud23-Trm112 de levure ont été résolues. L’étude comparative structurale de ces complexes et de la structure connue de Mtq2-Trm112, a permis de mettre en évidence que dans un même organisme, les séquences des trois protéines ont évolué de manière à conserver l’interaction avec Trm112. Même si les quatre partenaires présentent moins de 20% d’identité de séquence, les résidus clés pour l’interaction avec la petite protéine activatrice sont conservés ou partagent des caractéristiques identiques. En plus de l’analyse structurale, le complexe Trm9-Trm112 a fait l’objet d’une étude fonctionnelle chez S. cerevisiae ce qui a permis de cartographier le site actif de l’enzyme et de proposer un modèle de mécanisme d’action. Enfin, les premières études in vivo réalisées chez Haloferax volcanii suggèrent que cette plateforme serait également présente chez certains organismes procaryotes. / Protein synthesis is a central process in the cell; it ensures the transfer of genetic information from mRNA in to protein. A lot of actors are involved directly or indirectly in translation. In Eukaryotes, Trm112, a small protein, interacts with and activates four methyltransferases modifying direct actors of translation. The termination factor eRF1 is methylated by the Mtq2-Trm112 complex, the 18S rRNA by Bud23-Trm112 and some tRNA by the Trm9-Trm112 and Trm11-Trm112 complexes. During this work, the crystal structures of Trm9-Trm112 and Bud23-Trm112 complexes from yeast were solved. The comparative analysis of these two new structures with Mtq2-Trm112 structure highlights the structural plasticity allowing Trm112 to interact through a very similar mode with its partners although those share less than 20% sequence identity. In the same organism, the key residues for the interaction with Trm112 are conserved or share similar characteristics. In addition to the structural analysis, the function of the Trm9-Trm112 complex was studied in S. cerevisiae. This analysis allowed to map the active site of the enzyme and to propose a model of its mechanism of action. Finally, the first data obtained in vivo, with the Archaea Haloferax volcanii suggest that the Trm112 platform might also be present in some prokaryotic organisms.

Ribosomes are the cellular structures responsible for the synthesis of protein in all branches of life. All ribosomes are made from a large and small subunit that in turn are composed of protein and RNA. The synthesis of eukaryotic ribosomes is a complex process involving more than 200 factors and spans three cellular compartments: the nucleolus, the nucloplasm, and the cytoplasm. The precise function of most of these ribosome biogenesis factors remains unknown. The RNA component of ribosomes is in part processed from a large RNA transcript that yields most of the RNA present in mature ribosomes. Part of the maturation process involves modification of this ribosomal RNA as processing is carried out. Recent work constructing protein interaction networks in Saccharomyces cerevisiae suggested the methyltransferase Bud23 was involved in ribosome biogenesis (1). This thesis describes my work to characterize Bud23 and place it within the ribosome biogenesis pathway. Bud23 is a SAM methyltransferase important for the proper biogenesis of the small ribosomal subunit. Here I will demonstrate that Bud23 methylates G1575 of the small subunit ribosomal RNA (SSU rRNA), and its absence delays export of the SSU rRNA from the nucleolas, and the nucleus, and results in the delayed maturation of the SSU rRNA. Finally, I will show that Bud23 function is connected to small subunit processome factor Utp14 through identification of a Utp14 mutant that suppresses the bud23[Delta] deletion phenotype. / text

京都大学 / 新制・課程博士 / 博士(生命科学) / 甲第23555号 / 生博第466号 / 新制||生||62(附属図書館) / 京都大学大学院生命科学研究科高次生命科学専攻 / (主査)教授 朝長 啓造, 教授 野田 岳志, 教授 千坂 修 / 学位規則第4条第1項該当 / Doctor of Philosophy in Life Sciences / Kyoto University / DFAM