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Published online 1 June 2004

Nucleic Acids Research, 2004, Vol. 32, No. 10 2987-2994
© 2004 Oxford University Press

3'-Box-dependent processing of human pre-U1 snRNA requires a combination of RNA and protein co-factors

Patricia Uguen and Shona Murphy*

Sir William Dunn School of Pathology, University of Oxford, South Parks Road, Oxford OX1 3RE, UK

*To whom correspondence should be addressed. Tel: +44 1865 275616; Fax: +44 1865 275556; Email: shona.murphy{at}pathology.ox.ac.uk
Correspondence may also be addressed to Patricia Uguen at present address: Signalisation, Développement et Cancer, Bâtiment 442 bis, Université Paris XI, 91405 Orsay cédex, France. Tel: +33 1 69 15 65 93; Fax: +33 1 69 15 68 02; Email: patricia.uguen{at}ibaic.u-psud.fr

Received February 26, 2004; Revised and Accepted May 4, 2004

Using an in vitro system we have recently shown that the 3' ends of human pre-snRNAs synthesized by RNA polymerase II are produced by RNA processing directed by the snRNA gene-specific 3' box. Towards a complete characterization of this processing reaction we have further investigated the in vitro requirements for proper 3' end formation of pre-U1 snRNA. Here we show that the 5' cap plays a stimulatory role and processing requires creatine phosphate. Our results also indicate that the pre-U1 processing activity is heat sensitive and that an RNA component is required. In addition, the exact sequence adjacent to the 3' box influences the position of the pre-U1 3' end produced in vitro. Interestingly, the processing extract active for 3'-box-dependent processing also contains an activity that converts the 3' end of RNA containing the U1 Sm protein binding site and the 3' terminal stem–loop into the mature form.


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