Ntc90 is required for recruiting first step factor Yju2 but not for spliceosome activation

Abstract

The Prp19-associated complex (NineTeen Complex [NTC]) is required for spliceosome activation by specifying interactions of U5 and U6 with pre-mRNA on the spliceosome after the release of U4. The NTC consists of at least eight protein components, including two tetratricopeptide repeat (TPR)-containing proteins, Ntc90 and Ntc77. Ntc90 has nine copies of the TPR with seven clustered in the carboxy-terminal half of the protein, and interacts with all identified NTC components except for Prp19 and Ntc25. It forms a stable complex with Ntc31, Ntc30, and Ntc20 in the absence of Ntc25, when other interactions between NTC components are disrupted. In this study, we used both biochemical and genetic methods to analyze the structure of Ntc90, and its function in maintaining the integrity of the NTC and in NTC-mediated spliceosome activation. Our results show that Ntc90 interacts with Ntc31, Ntc30, and other NTC components through different regions of the protein, and that its function may be regulated by Ntc31 and Ntc30. Ntc90 is not required for the association of Prp19, Ntc85, Ntc77, Ntc25, and Ntc20, or for their binding to the spliceosome. It is also not required for NTC-mediated spliceosome activation, but is required for the recruitment of Yju2, which is involved in the first catalytic reaction after the function of Prp2. Our results demonstrate a novel role of the NTC in recruiting splicing factors to the spliceosome after its activation.