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Molecular and Cellular Biology, May 2000, p. 3116-3124, Vol. 20, No. 9
Department of Molecular, Cellular and
Developmental Biology, University of Colorado at Boulder, Boulder,
Colorado 80309
Received 22 November 1999/Returned for modification 5 January
2000/Accepted 8 February 2000
We demonstrate here the first experimental suppression of a
premature termination codon in vivo by using an ochre suppressor tRNA
acting in an intact mouse. Multicopy tRNA expression plasmids were
directly injected into skeletal muscle and into the hearts of
transgenic mice carrying a reporter gene with an ochre mutation. A
strategy for modulation of suppressor efficiency, applicable to diverse
systems and based on tandem multimerization of the tRNA gene, is
developed. The product of suppression (chloramphenicol acetyltransferase) accumulates linearly with increases in suppressor tRNA concentration to the point where the ochre-suppressing
tRNASer is in four- to fivefold excess over the endogenous
tRNASer. The subsequent suppressor activity plateau seems
to be attributable to accumulation of unmodified tRNAs. These results
define many salient variables for suppression in vivo, for example, for
tRNA suppression employed as gene therapy for nonsense defects.
0270-7306/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.
Suppression of Nonsense Mutations in Cell Culture
and Mice by Multimerized Suppressor tRNA Genes
*
Corresponding author. Mailing address: Department of
Molecular, Cellular and Developmental Biology, University of Colorado at Boulder, Campus Box 347, Boulder, CO 80309-0347. Phone: (303) 492-7606. Fax: (303) 492-8907. E-mail:
Leslie.Leinwand{at}Colorado.edu.
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