Cellular Variability of RpoS Expression Underlies Subpopulation Activation of an Integrative and Conjugative Element
Conjugative transfer of the integrative and conjugative element ICEclc in the bacterium Pseudomonas knackmussii is the consequence of a bistable decision taken in some 3% of cells in a population during stationary phase. Here we study the possible control exerted by the stationary phase sigma factor RpoS on the bistability decision. The gene for RpoS in P. knackmussii B13 was characterized, and a loss-of-function mutant was produced and complemented. We found that, in absence of RpoS, ICEclc transfer rates and activation of two key ICEclc promoters (Pint and PinR) decrease significantly in cells during stationary phase. Microarray and gene reporter analysis indicated that the most direct effect of RpoS is on PinR, whereas one of the gene products from the PinR-controlled operon (InrR) transmits activation to Pint and other ICEclc core genes. Addition of a second rpoS copy under control of its native promoter resulted in an increase of the proportion of cells expressing the Pint and PinR promoters to 18%. Strains in which rpoS was replaced by an rpoS-mcherry fusion showed high mCherry fluorescence of individual cells that had activated Pint and PinR, whereas a double-copy rpoS-mcherry–containing strain displayed twice as much mCherry fluorescence. This suggested that high RpoS levels are a prerequisite for an individual cell to activate PinR and thus ICEclc transfer. Double promoter–reporter fusions confirmed that expression of PinR is dominated by extrinsic noise, such as being the result of cellular variability in RpoS. In contrast, expression from Pint is dominated by intrinsic noise, indicating it is specific to the ICEclc transmission cascade. Our results demonstrate how stochastic noise levels of global transcription factors can be transduced to a precise signaling cascade in a subpopulation of cells leading to ICE activation.
Vyšlo v časopise:
Cellular Variability of RpoS Expression Underlies Subpopulation Activation of an Integrative and Conjugative Element. PLoS Genet 8(7): e32767. doi:10.1371/journal.pgen.1002818
Kategorie:
Research Article
prolekare.web.journal.doi_sk:
https://doi.org/10.1371/journal.pgen.1002818
Souhrn
Conjugative transfer of the integrative and conjugative element ICEclc in the bacterium Pseudomonas knackmussii is the consequence of a bistable decision taken in some 3% of cells in a population during stationary phase. Here we study the possible control exerted by the stationary phase sigma factor RpoS on the bistability decision. The gene for RpoS in P. knackmussii B13 was characterized, and a loss-of-function mutant was produced and complemented. We found that, in absence of RpoS, ICEclc transfer rates and activation of two key ICEclc promoters (Pint and PinR) decrease significantly in cells during stationary phase. Microarray and gene reporter analysis indicated that the most direct effect of RpoS is on PinR, whereas one of the gene products from the PinR-controlled operon (InrR) transmits activation to Pint and other ICEclc core genes. Addition of a second rpoS copy under control of its native promoter resulted in an increase of the proportion of cells expressing the Pint and PinR promoters to 18%. Strains in which rpoS was replaced by an rpoS-mcherry fusion showed high mCherry fluorescence of individual cells that had activated Pint and PinR, whereas a double-copy rpoS-mcherry–containing strain displayed twice as much mCherry fluorescence. This suggested that high RpoS levels are a prerequisite for an individual cell to activate PinR and thus ICEclc transfer. Double promoter–reporter fusions confirmed that expression of PinR is dominated by extrinsic noise, such as being the result of cellular variability in RpoS. In contrast, expression from Pint is dominated by intrinsic noise, indicating it is specific to the ICEclc transmission cascade. Our results demonstrate how stochastic noise levels of global transcription factors can be transduced to a precise signaling cascade in a subpopulation of cells leading to ICE activation.
Zdroje
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Štítky
Genetika Reprodukčná medicínaČlánok vyšiel v časopise
PLOS Genetics
2012 Číslo 7
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