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Host responses influence on the induction of lambda prophage.

Rokney A, Kobiler O, Amir A, Court DL, Stavans J, Adhya S, Oppenheim AB - Mol. Microbiol. (2008)

Bottom Line: We studied the effects of these two methods of induction on the lytic pathway by monitoring the activation of different lambda promoters, and found that the lambda genetic network co-ordinates information from the host stress response networks.Our results show that the function of the CII transcriptional activator, which facilitates the lysogenic developmental pathway, is not observed following either method of induction.We also show that, despite the common inhibitory effect on CII function, there are significant differences in the heat- and SOS-induced pathways leading to the lytic cascade.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Genetics and Biotechnology, The Hebrew University-Hadassah Medical School, Jerusalem, Israel. assafr@ekmd.huji.ac.il

ABSTRACT
Inactivation of bacteriophage lambda CI repressor leads almost exclusively to lytic development. Prophage induction can be initiated either by DNA damage or by heat treatment of a temperature-sensitive repressor. These two treatments also cause a concurrent activation of either the host SOS or heat-shock stress responses respectively. We studied the effects of these two methods of induction on the lytic pathway by monitoring the activation of different lambda promoters, and found that the lambda genetic network co-ordinates information from the host stress response networks. Our results show that the function of the CII transcriptional activator, which facilitates the lysogenic developmental pathway, is not observed following either method of induction. Mutations in the cro gene restore the CII function irrespective of the induction method. Deletion of the heat-shock protease gene ftsH can also restore CII function following heat induction but not following SOS induction. Our findings highlight the importance of the elimination of CII function during induction as a way to ensure an efficient lytic outcome. We also show that, despite the common inhibitory effect on CII function, there are significant differences in the heat- and SOS-induced pathways leading to the lytic cascade.

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Prophage induction by heat shock at 42°C.A. Strains W3110 (λcI857knR) carrying the pR-gfp (orange), the pE-gfp (blue) or the pR′-tR′-gfp (red) plasmids were grown exponentially in minimal medium, heat induced at 42°C for 5, 10 or 15 min (light to darker colours respectively), and assayed for fluorescence at 32°C for the indicated times. The onset time of the graphs describing fluorescence levels (top) and promoter activity (bottom) is the transfer to 32°C.B. Strains A8926 (λcI857knR) deleted for the ftsH gene (left) and A9855 (λcI857knR) deleted for the hflK gene (right) carrying the pE-gfp plasmid were grown exponentially in minimal medium, heat induced at 42°C for 5, 10 or 15 min (light to darker colours respectively), and transferred to 32°C (time 0 min). Fluorescence activity (top) and promoter activity (bottom) are shown. The activity of pE–GFP was monitored following heat induction.
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fig02: Prophage induction by heat shock at 42°C.A. Strains W3110 (λcI857knR) carrying the pR-gfp (orange), the pE-gfp (blue) or the pR′-tR′-gfp (red) plasmids were grown exponentially in minimal medium, heat induced at 42°C for 5, 10 or 15 min (light to darker colours respectively), and assayed for fluorescence at 32°C for the indicated times. The onset time of the graphs describing fluorescence levels (top) and promoter activity (bottom) is the transfer to 32°C.B. Strains A8926 (λcI857knR) deleted for the ftsH gene (left) and A9855 (λcI857knR) deleted for the hflK gene (right) carrying the pE-gfp plasmid were grown exponentially in minimal medium, heat induced at 42°C for 5, 10 or 15 min (light to darker colours respectively), and transferred to 32°C (time 0 min). Fluorescence activity (top) and promoter activity (bottom) are shown. The activity of pE–GFP was monitored following heat induction.

Mentions: To probe the induction process, we studied cells lysogenic for λ carrying the cI857 allele coding for a temperature-sensitive CI repressor (Sussman and Jacob, 1962). The transfer of these cells from 32°C to 42°C leads to rapid and coherent induction and expression of prophage genes. We introduced into these cells reporter plasmids containing either pR–GFP to monitor the inactivation of the CI repressor, pE–GFP to follow CII activity or pR′-tR′–GFP to monitor the anti-termination activity of Q. We have previously shown that expression from the pE–GFP and pR′-tR′–GFP reporters is absolutely dependent on the presence of CII and Q respectively (Kobiler et al., 2005). Lysogenic cells were transferred for 5, 10 or 15 min to 42°C, and then returned to 32°C for the monitoring of GFP levels. In a control experiment, heat treatment of λcI+ lysogenic cells carrying the wild-type repressor showed no effect on GFP activity from any of the three promoters (data not shown). The results, shown in Fig. 2A, can be summarized as follows:


Host responses influence on the induction of lambda prophage.

Rokney A, Kobiler O, Amir A, Court DL, Stavans J, Adhya S, Oppenheim AB - Mol. Microbiol. (2008)

Prophage induction by heat shock at 42°C.A. Strains W3110 (λcI857knR) carrying the pR-gfp (orange), the pE-gfp (blue) or the pR′-tR′-gfp (red) plasmids were grown exponentially in minimal medium, heat induced at 42°C for 5, 10 or 15 min (light to darker colours respectively), and assayed for fluorescence at 32°C for the indicated times. The onset time of the graphs describing fluorescence levels (top) and promoter activity (bottom) is the transfer to 32°C.B. Strains A8926 (λcI857knR) deleted for the ftsH gene (left) and A9855 (λcI857knR) deleted for the hflK gene (right) carrying the pE-gfp plasmid were grown exponentially in minimal medium, heat induced at 42°C for 5, 10 or 15 min (light to darker colours respectively), and transferred to 32°C (time 0 min). Fluorescence activity (top) and promoter activity (bottom) are shown. The activity of pE–GFP was monitored following heat induction.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2327240&req=5

fig02: Prophage induction by heat shock at 42°C.A. Strains W3110 (λcI857knR) carrying the pR-gfp (orange), the pE-gfp (blue) or the pR′-tR′-gfp (red) plasmids were grown exponentially in minimal medium, heat induced at 42°C for 5, 10 or 15 min (light to darker colours respectively), and assayed for fluorescence at 32°C for the indicated times. The onset time of the graphs describing fluorescence levels (top) and promoter activity (bottom) is the transfer to 32°C.B. Strains A8926 (λcI857knR) deleted for the ftsH gene (left) and A9855 (λcI857knR) deleted for the hflK gene (right) carrying the pE-gfp plasmid were grown exponentially in minimal medium, heat induced at 42°C for 5, 10 or 15 min (light to darker colours respectively), and transferred to 32°C (time 0 min). Fluorescence activity (top) and promoter activity (bottom) are shown. The activity of pE–GFP was monitored following heat induction.
Mentions: To probe the induction process, we studied cells lysogenic for λ carrying the cI857 allele coding for a temperature-sensitive CI repressor (Sussman and Jacob, 1962). The transfer of these cells from 32°C to 42°C leads to rapid and coherent induction and expression of prophage genes. We introduced into these cells reporter plasmids containing either pR–GFP to monitor the inactivation of the CI repressor, pE–GFP to follow CII activity or pR′-tR′–GFP to monitor the anti-termination activity of Q. We have previously shown that expression from the pE–GFP and pR′-tR′–GFP reporters is absolutely dependent on the presence of CII and Q respectively (Kobiler et al., 2005). Lysogenic cells were transferred for 5, 10 or 15 min to 42°C, and then returned to 32°C for the monitoring of GFP levels. In a control experiment, heat treatment of λcI+ lysogenic cells carrying the wild-type repressor showed no effect on GFP activity from any of the three promoters (data not shown). The results, shown in Fig. 2A, can be summarized as follows:

Bottom Line: We studied the effects of these two methods of induction on the lytic pathway by monitoring the activation of different lambda promoters, and found that the lambda genetic network co-ordinates information from the host stress response networks.Our results show that the function of the CII transcriptional activator, which facilitates the lysogenic developmental pathway, is not observed following either method of induction.We also show that, despite the common inhibitory effect on CII function, there are significant differences in the heat- and SOS-induced pathways leading to the lytic cascade.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Genetics and Biotechnology, The Hebrew University-Hadassah Medical School, Jerusalem, Israel. assafr@ekmd.huji.ac.il

ABSTRACT
Inactivation of bacteriophage lambda CI repressor leads almost exclusively to lytic development. Prophage induction can be initiated either by DNA damage or by heat treatment of a temperature-sensitive repressor. These two treatments also cause a concurrent activation of either the host SOS or heat-shock stress responses respectively. We studied the effects of these two methods of induction on the lytic pathway by monitoring the activation of different lambda promoters, and found that the lambda genetic network co-ordinates information from the host stress response networks. Our results show that the function of the CII transcriptional activator, which facilitates the lysogenic developmental pathway, is not observed following either method of induction. Mutations in the cro gene restore the CII function irrespective of the induction method. Deletion of the heat-shock protease gene ftsH can also restore CII function following heat induction but not following SOS induction. Our findings highlight the importance of the elimination of CII function during induction as a way to ensure an efficient lytic outcome. We also show that, despite the common inhibitory effect on CII function, there are significant differences in the heat- and SOS-induced pathways leading to the lytic cascade.

Show MeSH
Related in: MedlinePlus