The Evolutionarily Conserved Protein LAS1 Is Required for Pre-rRNA Processing at Both Ends of ITS2

Abstract

Ribosome synthesis entails the formation of mature rRNAs from long precursor molecules, following a complex pre-rRNA processing pathway. Why the generation of mature rRNA ends is so complicated is unclear. Nor is it understood how pre-rRNA processing is coordinated at distant sites on pre-rRNA molecules. Here we characterized, in budding yeast and human cells, the evolutionarily conserved protein Las1. We found that, in both species, Las1 is required to process ITS2, which separates the 5.8S and 25S/28S rRNAs. In yeast, Las1 is required for pre-rRNA processing at both ends of ITS2. It is required for Rrp6-dependent formation of the 5.8S rRNA 3′ end and for Rat1-dependent formation of the 25S rRNA 5′ end. We further show that the Rat1-Rai1 5′-3′ exoribonuclease (exoRNase) complex functionally connects processing at both ends of the 5.8S rRNA. We suggest that pre-rRNA processing is coordinated at both ends of 5.8S rRNA and both ends of ITS2, which are brought together by pre-rRNA folding, by an RNA processing complex. Consistently, we note the conspicuous presence of ∼7- or 8-nucleotide extensions on both ends of 5.8S rRNA precursors and at the 5′ end of pre-25S RNAs suggestive of a protected spacer fragment of similar length.

SUPPLEMENTAL MATERIAL

Supplemental material for this article may be found at http://dx.doi.org/10.1128/MCB.06019-11.

ACKNOWLEDGMENTS

We thank the following colleagues for providing reagents: Ambro van Hoof (University of Texas Health Science Center-Houston) for the rex mutants, Giorgio Dieci (Università di Parma) for the anti-Rps8 antibody, Micheline Fromont-Racine (Institut Pasteur, Paris) for the anti-Nog1 antibody, Jonathan Warner (Albert Einstein College, NYC) for the anti-Rpl3 antibody, and Aziz El Hage and David Tollervey (Wellcome Trust Centre for Cell Biology, Edinburgh) for the pMET::rat1 xrn1Δ strain. We are deeply indebted to Christiane Zorbas, Taylor Mullineux, and Lionel Tafforeau for help with the RNA interference depletions.

This work was funded by the FRIA, the FRS-F.N.R.S., the Communauté Française de Belgique (ARC), and the Région Wallone (Cibles).