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Journal of Bacteriology, February 2007, p. 980-990, Vol. 189, No. 3
0021-9193/07/$08.00+0     doi:10.1128/JB.01287-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Self-Splicing of the Bacteriophage T4 Group I Introns Requires Efficient Translation of the Pre-mRNA In Vivo and Correlates with the Growth State of the Infected Bacterium

Linus Sandegren and Britt-Marie Sjöberg^*

Department of Molecular Biology and Functional Genomics, Stockholm University, Stockholm, Sweden

Received 15 August 2006/ Accepted 13 November 2006

Bacteriophage T4 contains three self-splicing group I introns in genes in de novo deoxyribonucleotide biosynthesis (in td, coding for thymidylate synthase and in nrdB and nrdD, coding for ribonucleotide reductase). Their presence in these genes has fueled speculations that the introns are retained within the phage genome due to a possible regulatory role in the control of de novo deoxyribonucleotide synthesis. To study whether sequences in the upstream exon interfere with proper intron folding and splicing, we inhibited translation in T4-infected bacteria as well as in bacteria containing recombinant plasmids carrying the nrdB intron. Splicing was strongly reduced for all three T4 introns after the addition of chloramphenicol during phage infection, suggesting that the need for translating ribosomes is a general trait for unperturbed splicing. The splicing of the cloned nrdB intron was markedly reduced in the presence of chloramphenicol or when translation was hindered by stop codons inserted in the upstream exon. Several exon regions capable of forming putative interactions with nrdB intron sequences were identified, and the removal or mutation of these exon regions restored splicing efficiency in the absence of translation. Interestingly, splicing of the cloned nrdB intron was also reduced as cells entered stationary phase and splicing of all three introns was reduced upon the T4 infection of stationary-phase bacteria. Our results imply that conditions likely to be frequently encountered by natural phage populations may limit the self-splicing efficiency of group I introns. This is the first time that environmental effects on bacterial growth have been linked to the regulation of splicing of phage introns.

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* Corresponding author. Mailing address: Department of Molecular Biology and Functional Genomics, Stockholm University, Svante Arrhenius väg 16 F3, SE-10691 Stockholm, Sweden. Phone: 46-8-164150. Fax: 46-8-166488. E-mail: britt-marie.sjoberg{at}molbio.su.se.

Published ahead of print on 22 November 2006.

Present address: Department of Medical Biochemistry and Microbiology, Uppsala University, Uppsala, Sweden.

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Journal of Bacteriology, February 2007, p. 980-990, Vol. 189, No. 3
0021-9193/07/$08.00+0     doi:10.1128/JB.01287-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

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