ROLE DE DEUX ARN DANS LE CONTROLE DE L'EXPRESSION DES GENES: REGULATIONS DE LA REPLICATION DU PLASMIDE R1 PAR UN ARN ANTISENS ET DES GENES DE VIRULENCE DE STAPHYLOCOCCUS AUREUS PAR L'ARN-III

The antisense RNA (CopA) regulates bacterial plasmid R1 replication by controlling the rate of synthesis of the RepA initiator protein. CopA binds its complementary region (CopT) in the 5' untranslated region of repA mRNA. This binding essentially inhibits RepA synthesis and promotes rapid degradation of the mRNA by RNase III. Rapid complex formation is a prerequesite for proper control. We have shown that the two RNAs interact via a loop-loop contact that has to be rapidly converted to form an irreversible and functional complex. This complex is not a full duplex but contains a four-way RNA junction stabilized by a long intermolecular helix. Several binding intermediates leading to the stable complex were characterized, as well as CopA and CopT structural determinants required for this essential conversion step. Thus, we propose a mechanism for stable complex formation, involving several steps in a hierarchical order. This unexpected mode of binding between two complementary RNAs appears to be rather a rule than an exception. Indeed, we have shown that this mechanism is conserved among many R1-related plasmids. RNA-III controls the expression of virulence gene expression in Staphylococcus aureus. The aim of the second part of this thesis was to define the secondary structure of RNA-III in vitro and in vivo, and to define independant functional domains. Combining several in vitro approaches, we established that RNA-III contains 14 stem-loop structures and 3 long-range interactions. We also identified a functional sub-domain involved in the control of protein A expression.