Human H/ACA Small Nucleolar RNPs and Telomerase Share Evolutionarily Conserved Proteins NHP2 and NOP10

doi:10.1128/MCB.20.23.9028-9040.2000

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Molecular and Cellular Biology, December 2000, p. 9028-9040, Vol. 20, No. 23
0270-7306/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Human H/ACA Small Nucleolar RNPs and Telomerase Share Evolutionarily Conserved Proteins NHP2 and NOP10

Vanda Poga $c$ i, François Dragon, and Witold Filipowicz^*

Friedrich-Miescher Institut, CH-4058 Basel, Switzerland

Received 27 July 2000/Returned for modification 30 August 2000/Accepted 12 September 2000

The H/ACA small nucleolar RNAs (snoRNAs) are involved inpseudouridylation of pre-rRNAs. In the yeast Saccharomyces cerevisiae, four commonproteins are associated with H/ACA snoRNAs: Gar1p, Cbf5p, Nhp2p, and Nop10p.In vitro reconstitution studies showed that four proteins also specificallyinteract with H/ACA snoRNAs in mammalian cell extracts. Two mammalian proteins,NAP57/dyskerin (the ortholog of Cbf5p) and hGAR1, have been characterized.In this work we describe properties of hNOP10 and hNHP2, human orthologs ofyeast Nop10p and Nhp2p, respectively, and further characterize hGAR1. hNOP10 and hNHP2complement yeast cells depleted of Nhp2p and Nop10p, respectively. Immunoprecipitationexperiments with extracts from transfected HeLa cells indicated that epitope-tagged hNOP10and hNHP2 specifically associate with hGAR1 and H/ACA RNAs; they alsointeract with the RNA subunit of telomerase, which contains an H/ACA-like domain inits 3' moiety. Immunofluorescence microscopy experiments showed that hGAR1, hNOP10,and hNHP2 are localized in the dense fibrillar component of the nucleolus and in Cajal (coiled)bodies. Deletion analysis of hGAR1 indicated that its evolutionarily conserved core domaincontains all the signals required for localization, but progressive deletions from eitherthe N or the C terminus of the core domain abolish localization in the nucleolus and/orthe Cajal bodies.

^* Corresponding author. Mailing address: Friedrich Miescher Institut, Maulbeerstrasse 66, CH-4058 Basel, Switzerland. Phone: 41 61 697 4128.Fax: 41 61 697 3976. E-mail: Witold.Filipowicz{at}fmi.ch.

Present address: Department of Therapeutic Radiology, Yale University School of Medicine, New Haven, CT 06520-8040.

Molecular and Cellular Biology, December 2000, p. 9028-9040, Vol. 20, No. 23
0270-7306/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

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