Nucleic Acids Research, 2002, Vol. 30, No. 7 1606-1612
© 2002 Oxford University Press
Efficient trans-cleavage by the Schistosoma mansoni SM
1 hammerhead ribozyme in the extreme thermophile Thermus thermophilus
Département de Biochimie, Université de Montréal, CP 6128, Succ. Centre-Ville, Montréal, Quebec H3C 3J7, Canada, 1Centro de Biología Molecular ‘Severo Ochoa’, CSIC-UAM, Universidad Autónoma de Madrid, Madrid 28049, Spain and 2Institute of Parasitology, Macdonald Campus of McGill University, Ste-Anne-de Bellevue, Quebec H9X 3V9, Canada
The catalytic hammerhead structure has been found in association with repetitive DNA from several animals, including salamanders, crickets and schistosomes, and functions to process in cis the long multimer transcripts into monomer RNA in vivo. The cellular role of these repetitive elements and their transcripts is unknown. Moreover, none of these natural hammerheads have been shown to trans-cleave a host mRNA in vivo. We analyzed the cis- and trans-cleavage properties of the hammerhead ribozyme associated with the SM
DNA family from the human parasite Schistosoma mansoni. The efficiency of trans-cleavage of a target RNA in vitro was affected mainly by both the temperature-dependent chemical step and the ribozyme–product dissociation step. The optimal temperature for trans-cleavage was 70°C. This result was confirmed when both the SM1 ribozyme and the target RNA were expressed in the extreme thermophile Thermus thermophilus. Moreover, SM1 RNA showed a remarkable thermostability, equal or superior to that of the most stable RNAs in this species, suggesting that SM1 RNA has been selected for stability. Computer analysis predicts that the monomer and multimer transcripts fold into highly compact secondary structures, which may explain their exceptional stability in vivo.
* To whom correspondence should be addressed at present address: Hôpital Sainte-Justine, Centre de Recherche, 3175 Cote Sainte-Catherine Road, Montréal, Quebec H3T 1C5, Canada. Tel: +1 514 345 4931; Fax: +1 514 345 4931; Email: vazqueza{at}magellan.umontreal.ca This work is dedicated to the memory of Professor Robert Cedergren
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