Role of Pre-rRNA Base Pairing and 80S Complex Formation in Subnucleolar Localization of the U3 snoRNP

Abstract

In the nucleolus the U3 snoRNA is recruited to the 80S pre-rRNA processing complex in the dense fibrillar component (DFC). The U3 snoRNA is found throughout the nucleolus and has been proposed to move with the preribosomes to the granular component (GC). In contrast, the localization of other RNAs, such as the U8 snoRNA, is restricted to the DFC. Here we show that the incorporation of the U3 snoRNA into the 80S processing complex is not dependent on pre-rRNA base pairing sequences but requires the B/C motif, a U3-specific protein-binding element. We also show that the binding of Mpp10 to the 80S U3 complex is dependent on sequences within the U3 snoRNA that base pair with the pre-rRNA adjacent to the initial cleavage site. Furthermore, mutations that inhibit 80S complex formation and/or the association of Mpp10 result in retention of the U3 snoRNA in the DFC. From this we propose that the GC localization of the U3 snoRNA is a direct result of its active involvement in the initial steps of ribosome biogenesis.

We thank Reinout Raijmakers and Claudia Schneider for critically reading the manuscript, Sylviane Muller for generating the anti-hU3-55K antibodies, and Larry Gerace for providing the anti-hMpp10 antibodies. We also thank Angus Lamond and Anthony Leung for providing the stably transfected ECFP-fibrillarin strain.

This work was supported by grants from the Deutsche Forschungsgemeinschaft (SFB523/A-8) and Fonds der Chemischen Industrie (to R.L.), the Council for Chemical Sciences of The Netherlands Organization for Scientific Research (NWO-CW; grant 97-033 to W.J.V.V.) and by an EMBO Short Term Fellowship (ASTF no. 9781 to S.G.).