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Molecular and Cellular Biology, August 2004, p. 7284-7297, Vol. 24, No. 16
0270-7306/04/$08.00+0     DOI: 10.1128/MCB.24.16.7284-7297.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

A Plant snoRNP Complex Containing snoRNAs, Fibrillarin, and Nucleolin-Like Proteins Is Competent for both rRNA Gene Binding and Pre-rRNA Processing In Vitro

Julio Sáez-Vasquez, David Caparros-Ruiz, Fredy Barneche, and Manuel Echeverría^*

Laboratoire Génome et Développement des Plantes, UMR CNRS-IRD 5096, Université de Perpignan, 66860 Perpignan Cedex, France

Received 26 February 2004/ Returned for modification 31 March 2004/ Accepted 24 May 2004

In eukaryotes the primary cleavage of the precursor rRNA (pre-rRNA) occurs in the 5' external transcribed spacer (5'ETS). In Saccharomyces cerevisiae and animals this cleavage depends on a conserved U3 small nucleolar ribonucleoprotein particle (snoRNP), including fibrillarin, and on other transiently associated proteins such as nucleolin. This large complex can be visualized by electron microscopy bound to the nascent pre-rRNA soon after initiation of transcription. Our group previously described a radish rRNA gene binding activity, NF D, that specifically binds to a cluster of conserved motifs preceding the primary cleavage site in the 5'ETS of crucifer plants including radish, cauliflower, and Arabidopsis thaliana (D. Caparros-Ruiz, S. Lahmy, S. Piersanti, and M. Echeverria, Eur. J. Biochem. 247:981-989, 1997). Here we report the purification and functional characterization of NF D from cauliflower inflorescences. Remarkably NF D also binds to 5'ETS RNA and accurately cleaves it at the primary cleavage site mapped in vivo. NF D is a multiprotein factor of 600 kDa that dissociates into smaller complexes. Two polypeptides of NF D identified by microsequencing are homologues of nucleolin and fibrillarin. The conserved U3 and U14 snoRNAs associated with fibrillarin and required for early pre-rRNA cleavages are also found in NF D. Based on this it is proposed that NF D is a processing complex that assembles on the rDNA prior to its interaction with the nascent pre-rRNA.

Present address: IBMB-CSIC, 08034 Barcelona, Spain.

Present address: Département de Biologie Moléculaire, Université de Genève-Sciences III, 1211 Genève, Switzerland.

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