Replication of Vibrio cholerae chromosome I in Escherichia coli: dependence on dam methylation.

Birgit Koch, Xiaofang Ma, Anders Løbner-Olesen

    Publikation: Bidrag til tidsskriftTidsskriftartikelForskningpeer review

    Resumé

    We successfully substituted Escherichia coli's origin of replication oriC with the origin region of Vibrio cholerae chromosome I (oriCIVc). Replication from oriCIVc initiated at a similar or slightly reduced cell mass compared to that of normal E. coli oriC. With respect to sequestration-dependent synchrony of initiation and stimulation of initiation by the loss of Hda activity, replication initiation from oriC and oriCIVc were similar. Since Hda is involved in the conversion of DnaAATP (DnaA bound to ATP) to DnaAADP (DnaA bound to ADP), this indicates that DnaA associated with ATP is limiting for V. cholerae chromosome I replication, which similar to what is observed for E. coli. No hda homologue has been identified in V. cholerae yet. In V. cholerae, dam is essential for viability, whereas in E. coli, dam mutants are viable. Replacement of E. coli oriC with oriCIVc allowed us to specifically address the role of the Dam methyltransferase and SeqA in replication initiation from oriCIVc. We show that when E. coli's origin of replication is substituted by oriCIVc, dam, but not seqA, becomes important for growth, arguing that Dam methylation exerts a critical function at the origin of replication itself. We propose that Dam methylation promotes DnaA-assisted successful duplex opening and replisome assembly at oriCIVc in E. coli. In this model, methylation at oriCIVc would ease DNA melting. This is supported by the fact that the requirement for dam can be alleviated by increasing negative supercoiling of the chromosome through oversupply of the DNA gyrase or loss of SeqA activity.
    OriginalsprogEngelsk
    TidsskriftJournal of Bacteriology
    Vol/bind192
    Udgave nummer15
    Sider (fra-til)3903-3914
    Antal sider11
    ISSN0021-9193
    DOI
    StatusUdgivet - 2010

    Citer dette

    Koch, Birgit ; Ma, Xiaofang ; Løbner-Olesen, Anders. / Replication of Vibrio cholerae chromosome I in Escherichia coli: dependence on dam methylation. I: Journal of Bacteriology. 2010 ; Bind 192, Nr. 15. s. 3903-3914.
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    title = "Replication of Vibrio cholerae chromosome I in Escherichia coli: dependence on dam methylation.",
    abstract = "We successfully substituted Escherichia coli's origin of replication oriC with the origin region of Vibrio cholerae chromosome I (oriCIVc). Replication from oriCIVc initiated at a similar or slightly reduced cell mass compared to that of normal E. coli oriC. With respect to sequestration-dependent synchrony of initiation and stimulation of initiation by the loss of Hda activity, replication initiation from oriC and oriCIVc were similar. Since Hda is involved in the conversion of DnaAATP (DnaA bound to ATP) to DnaAADP (DnaA bound to ADP), this indicates that DnaA associated with ATP is limiting for V. cholerae chromosome I replication, which similar to what is observed for E. coli. No hda homologue has been identified in V. cholerae yet. In V. cholerae, dam is essential for viability, whereas in E. coli, dam mutants are viable. Replacement of E. coli oriC with oriCIVc allowed us to specifically address the role of the Dam methyltransferase and SeqA in replication initiation from oriCIVc. We show that when E. coli's origin of replication is substituted by oriCIVc, dam, but not seqA, becomes important for growth, arguing that Dam methylation exerts a critical function at the origin of replication itself. We propose that Dam methylation promotes DnaA-assisted successful duplex opening and replisome assembly at oriCIVc in E. coli. In this model, methylation at oriCIVc would ease DNA melting. This is supported by the fact that the requirement for dam can be alleviated by increasing negative supercoiling of the chromosome through oversupply of the DNA gyrase or loss of SeqA activity.",
    author = "Birgit Koch and Xiaofang Ma and Anders L{\o}bner-Olesen",
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    Replication of Vibrio cholerae chromosome I in Escherichia coli: dependence on dam methylation. / Koch, Birgit; Ma, Xiaofang; Løbner-Olesen, Anders.

    I: Journal of Bacteriology, Bind 192, Nr. 15, 2010, s. 3903-3914.

    Publikation: Bidrag til tidsskriftTidsskriftartikelForskningpeer review

    TY - JOUR

    T1 - Replication of Vibrio cholerae chromosome I in Escherichia coli: dependence on dam methylation.

    AU - Koch, Birgit

    AU - Ma, Xiaofang

    AU - Løbner-Olesen, Anders

    PY - 2010

    Y1 - 2010

    N2 - We successfully substituted Escherichia coli's origin of replication oriC with the origin region of Vibrio cholerae chromosome I (oriCIVc). Replication from oriCIVc initiated at a similar or slightly reduced cell mass compared to that of normal E. coli oriC. With respect to sequestration-dependent synchrony of initiation and stimulation of initiation by the loss of Hda activity, replication initiation from oriC and oriCIVc were similar. Since Hda is involved in the conversion of DnaAATP (DnaA bound to ATP) to DnaAADP (DnaA bound to ADP), this indicates that DnaA associated with ATP is limiting for V. cholerae chromosome I replication, which similar to what is observed for E. coli. No hda homologue has been identified in V. cholerae yet. In V. cholerae, dam is essential for viability, whereas in E. coli, dam mutants are viable. Replacement of E. coli oriC with oriCIVc allowed us to specifically address the role of the Dam methyltransferase and SeqA in replication initiation from oriCIVc. We show that when E. coli's origin of replication is substituted by oriCIVc, dam, but not seqA, becomes important for growth, arguing that Dam methylation exerts a critical function at the origin of replication itself. We propose that Dam methylation promotes DnaA-assisted successful duplex opening and replisome assembly at oriCIVc in E. coli. In this model, methylation at oriCIVc would ease DNA melting. This is supported by the fact that the requirement for dam can be alleviated by increasing negative supercoiling of the chromosome through oversupply of the DNA gyrase or loss of SeqA activity.

    AB - We successfully substituted Escherichia coli's origin of replication oriC with the origin region of Vibrio cholerae chromosome I (oriCIVc). Replication from oriCIVc initiated at a similar or slightly reduced cell mass compared to that of normal E. coli oriC. With respect to sequestration-dependent synchrony of initiation and stimulation of initiation by the loss of Hda activity, replication initiation from oriC and oriCIVc were similar. Since Hda is involved in the conversion of DnaAATP (DnaA bound to ATP) to DnaAADP (DnaA bound to ADP), this indicates that DnaA associated with ATP is limiting for V. cholerae chromosome I replication, which similar to what is observed for E. coli. No hda homologue has been identified in V. cholerae yet. In V. cholerae, dam is essential for viability, whereas in E. coli, dam mutants are viable. Replacement of E. coli oriC with oriCIVc allowed us to specifically address the role of the Dam methyltransferase and SeqA in replication initiation from oriCIVc. We show that when E. coli's origin of replication is substituted by oriCIVc, dam, but not seqA, becomes important for growth, arguing that Dam methylation exerts a critical function at the origin of replication itself. We propose that Dam methylation promotes DnaA-assisted successful duplex opening and replisome assembly at oriCIVc in E. coli. In this model, methylation at oriCIVc would ease DNA melting. This is supported by the fact that the requirement for dam can be alleviated by increasing negative supercoiling of the chromosome through oversupply of the DNA gyrase or loss of SeqA activity.

    U2 - 10.1128/JB.00311-10

    DO - 10.1128/JB.00311-10

    M3 - Journal article

    VL - 192

    SP - 3903

    EP - 3914

    JO - Journal of Bacteriology

    JF - Journal of Bacteriology

    SN - 0021-9193

    IS - 15

    ER -