Project description:Previously, we discovered that the global regulator Hha is related to cell death in biofilms and regulates cryptic prophage genes. Here we show Hha induces excision of prophage CP4-57 and DLP12 by inducing excision genes and by reducing SsrA synthesis. SsrA is a tmRNA important for rescuing stalled ribosomes, contains an attachment site for CP4-57, and is shown here to be required for CP4-57 excision. These prophages impact biofilm development as deletion of 35 genes individually of prophage CP4-57 and DLP12 increase biofilm formation up to 17-fold, and 5 genes decrease biofilm formation up to 6-fold. In addition, CP4-57 excises during early biofilm development but not in planktonic cells, whereas DLP12 excision was detected at all the developmental stages for both biofilm and planktonic cells. CP4-57 excision leads to a chromosome region devoid of the prophage and formation of a phage circle (which is lost). These results were corroborated by a whole-transcriptome analysis that showed that complete loss of CP4-57 activated expression of the flg, flh, and fli motility operons and repressed expression of key enzymes in the tricarboxylic acid cycle and enzymes for lactate utilization. Prophage excision also results in the expression of cell lysis genes that reduce cell viability (e.g., alpA, intA, and intD). Hence, defective prophage are involved in host physiology via Hha and in biofilm formation by generating a diversified population with specialized functions in terms of motility and nutrient metabolism. Experiment Overall Design: Strain: CP4-57-deficient vs. wild-type Experiment Overall Design: Medium: LB Experiment Overall Design: Culture type:OD=0.5, planktonic cells
Project description:Previously, we discovered that the global regulator Hha is related to cell death in biofilms and regulates cryptic prophage genes. Here we show Hha induces excision of prophage CP4-57 and DLP12 by inducing excision genes and by reducing SsrA synthesis. SsrA is a tmRNA important for rescuing stalled ribosomes, contains an attachment site for CP4-57, and is shown here to be required for CP4-57 excision. These prophages impact biofilm development as deletion of 35 genes individually of prophage CP4-57 and DLP12 increase biofilm formation up to 17-fold, and 5 genes decrease biofilm formation up to 6-fold. In addition, CP4-57 excises during early biofilm development but not in planktonic cells, whereas DLP12 excision was detected at all the developmental stages for both biofilm and planktonic cells. CP4-57 excision leads to a chromosome region devoid of the prophage and formation of a phage circle (which is lost). These results were corroborated by a whole-transcriptome analysis that showed that complete loss of CP4-57 activated expression of the flg, flh, and fli motility operons and repressed expression of key enzymes in the tricarboxylic acid cycle and enzymes for lactate utilization. Prophage excision also results in the expression of cell lysis genes that reduce cell viability (e.g., alpA, intA, and intD). Hence, defective prophage are involved in host physiology via Hha and in biofilm formation by generating a diversified population with specialized functions in terms of motility and nutrient metabolism.
Project description:Total proteome analysis of CP4-57 prophage- and ssrA-related variants in Escherichia coli. Quantification was performed by SWATH-MS method.
Project description:There are three projects including AE155LQ, RE161LQ, RE235LQ. AE155LQ is about the proteomes of wild type cells (WT) and E. coli RelA* OE strain during exponential growth in glucose cAA medium. E1 & E2 corresponds to the wild type cells (two biological replicate samples) and E3 & E4 corresponds to RelA* OE data (two biological replicate samples). RE161LQ is about the time-course proteome of wild type (four samples including WT_0, WT_20, WT_40 and WT_80) vs relA-deficient strain (relA_0, relA_1.5 h, relA_3 h and relA_4.5 h) during AA downshift. RE235Q is about the time-course proteome of wild type (three samples including WT_0, WT_60, WT_160) vs relA-deficient strain (three samples including relA_0, relA_60 and relA_160) during carbon downshift.