TY - JOUR

T1 - Suppressors of DnaAATP imposed overinitiation in Escherichia coli

AU - Charbon, Godefroid

AU - Riber, Leise

AU - Cohen, Malene

AU - Skovgaard, Ole

AU - Fujimitsu, Kazuyuki

AU - Katayama, Tsutomu

AU - Løbner-Olesen, Anders

N1 - © 2010 Blackwell Publishing Ltd.

PY - 2011

Y1 - 2011

N2 - Chromosome replication in Escherichia coli is limited by the supply of DnaA associated with ATP. Cells deficient in RIDA (Regulatory Inactivation of DnaA) due to a deletion of the hda gene accumulate suppressor mutations (hsm) to counteract the overinitiation caused by an elevated DnaA(ATP) level. Eight spontaneous hda suppressor mutations were identified by whole-genome sequencing, and three of these were analysed further. Two mutations (hsm-2 and hsm-4) mapped in the dnaA gene and led to a reduced ability to initiate replication from oriC. One mutation (hsm-1) mapped to the seqA promoter and increased the SeqA protein level in the cell. hsm-1 cells had prolonged origin sequestration, reduced DnaA protein level and reduced DnaA-Reactivating Sequence (DARS)-mediated rejuvenation of DnaA(ADP) to DnaA(ATP) , all of which could contribute to the suppression of RIDA deficiency. Despite of these defects hsm-1 cells were quite similar to wild type with respect to cell cycle parameters. We speculate that since SeqA binding sites might overlap with DnaA binding sites spread throughout the chromosome, excess SeqA could interfere with DnaA titration and thereby increase free DnaA level. Thus, in spite of reduction in total DnaA, the amount of DnaA molecules available for initiation may not be reduced.

AB - Chromosome replication in Escherichia coli is limited by the supply of DnaA associated with ATP. Cells deficient in RIDA (Regulatory Inactivation of DnaA) due to a deletion of the hda gene accumulate suppressor mutations (hsm) to counteract the overinitiation caused by an elevated DnaA(ATP) level. Eight spontaneous hda suppressor mutations were identified by whole-genome sequencing, and three of these were analysed further. Two mutations (hsm-2 and hsm-4) mapped in the dnaA gene and led to a reduced ability to initiate replication from oriC. One mutation (hsm-1) mapped to the seqA promoter and increased the SeqA protein level in the cell. hsm-1 cells had prolonged origin sequestration, reduced DnaA protein level and reduced DnaA-Reactivating Sequence (DARS)-mediated rejuvenation of DnaA(ADP) to DnaA(ATP) , all of which could contribute to the suppression of RIDA deficiency. Despite of these defects hsm-1 cells were quite similar to wild type with respect to cell cycle parameters. We speculate that since SeqA binding sites might overlap with DnaA binding sites spread throughout the chromosome, excess SeqA could interfere with DnaA titration and thereby increase free DnaA level. Thus, in spite of reduction in total DnaA, the amount of DnaA molecules available for initiation may not be reduced.

KW - Adenosine Diphosphate

KW - Adenosine Triphosphate

KW - Bacterial Outer Membrane Proteins

KW - Bacterial Proteins

KW - Chromosomes, Bacterial

KW - DNA Replication

KW - DNA, Bacterial

KW - DNA-Binding Proteins

KW - Escherichia coli

KW - Escherichia coli Proteins

KW - Gene Expression Regulation, Bacterial

KW - Mutation

KW - Suppression, Genetic

U2 - 10.1111/j.1365-2958.2010.07493.x

DO - 10.1111/j.1365-2958.2010.07493.x

M3 - Journal article

C2 - 21299647

VL - 79

SP - 914

EP - 928

JO - Molecular Microbiology

JF - Molecular Microbiology

SN - 0950-382X

IS - 4

ER -