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Molecular and Cellular Biology, October 2002, p. 7053-7065, Vol. 22, No. 20
0270-7306/02/$04.00+0     DOI: 10.1128/MCB.22.20.7053-7065.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Naf1p, an Essential Nucleoplasmic Factor Specifically Required for Accumulation of Box H/ACA Small Nucleolar RNPs{dagger}

Christophe Dez, Jacqueline Noaillac-Depeyre, Michèle Caizergues-Ferrer, and Yves Henry*

Laboratoire de Biologie Moléculaire Eucaryote du CNRS, Université Paul Sabatier, 31062 Toulouse Cedex 04, France

Received 8 May 2002/ Returned for modification 7 June 2002/ Accepted 19 July 2002

Box H/ACA small nucleolar ribonucleoprotein particles (H/ACA snoRNPs) play key roles in the synthesis of eukaryotic ribosomes. The ways in which these particles are assembled and correctly localized in the dense fibrillar component of the nucleolus remain largely unknown. Recently, the essential Saccharomyces cerevisiae Naf1p protein (encoded by the YNL124W open reading frame) was found to interact in a two-hybrid assay with two core protein components of mature H/ACA snoRNPs, Cbf5p and Nhp2p (T. Ito, T. Chiba, R. Ozawa, M. Yoshida, M. Hattori, and Y. Sakaki, Proc. Natl. Acad. Sci. USA 98:4569-4574, 2001). Here we show that several H/ACA snoRNP components are weakly but specifically immunoprecipitated with epitope-tagged Naf1p, suggesting that the latter protein is involved in H/ACA snoRNP biogenesis, trafficking, and/or function. Consistent with this, we find that depletion of Naf1p leads to a defect in 18S rRNA accumulation. Naf1p is unlikely to directly assist H/ACA snoRNPs during pre-rRNA processing in the dense fibrillar component of the nucleolus for two reasons. Firstly, Naf1p accumulates predominantly in the nucleoplasm. Secondly, Naf1p sediments in a sucrose gradient chiefly as a free protein or associated in a complex of the size of free snoRNPs, whereas extremely little Naf1p is found in fractions containing preribosomes. These results are more consistent with a role for Naf1p in H/ACA snoRNP biogenesis and/or intranuclear trafficking. Indeed, depletion of Naf1p leads to a specific and dramatic decrease in the steady-state accumulation of all box H/ACA snoRNAs tested and of Cbf5p, Gar1p, and Nop10p. Naf1p is unlikely to be directly required for the synthesis of H/ACA snoRNP components. Naf1p could participate in H/ACA snoRNP assembly and/or transport.

* Corresponding author. Mailing address: Laboratoire de Biologie Moléculaire Eucaryote du CNRS, Université Paul Sabatier, 118 Route de Narbonne, 31062 Toulouse Cedex 04, France. Phone: 33 5 61 33 59 53. Fax: 33 5 61 33 58 86. E-mail: henry{at}ibcg.biotoul.fr.

{dagger} We dedicate this work to the memory of Albert Henry (1910-2002), Université Libre de Bruxelles et Académie Royale de Belgique.

Molecular and Cellular Biology, October 2002, p. 7053-7065, Vol. 22, No. 20
0022-538X/02/$04.00+0     DOI: 10.1128/MCB.22.20.7053-7065.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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