Timing and coordination of DNA replication in Escherichia coli
Appears in the following Collection
AbstractInitiation of DNA replication in the bacterium Escherichia coli (E. coli) occurs once per cell cycle at a unique origin site named oriC. The initiator, DnaA, is essential for initiation from oriC. Two control systems have been discovered that prevent re-initiation. One of the control systems involves inactivation of the initiator protein, DnaA, by active replication forks. The other system relies on specific inactivation (sequestration) of newly replicated origins. The SeqA protein plays an important role in sequestration. In the work presented in this thesis we study the role of the SeqA protein in control of replication initiation and in origin and replication fork organization. We show that SeqA binds cooperatively to newly replicated origins and propose a model in which SeqA forms a left-handed helical multimer upon DNA binding. The replication forks originated from the same origin has been suggested to stay in close proximity into a structure called the replication factory. We find support for this model in cells that grow rapidly with multifork DNA replication. We also find that sister origins stay colocalized, increasingly so with increasing growth rate. We show that the SeqA protein is involved in colocalization of sister replication forks and origins in vivo. In agreement, we show that SeqA is capable of paring newly replicated sister DNA molecules in vitro. We also report the development of a bacterial screening assay for discovery of novel antibacterial agents that target the initiator protein, DnaA. So far no drugs that target the essential DNA replication machinery have been discovered. Compounds discovered by this screen will therefore constitute a new group of antibacterial drugs.
List of papers
|1. Fossum S, Crooke E and Skarstad K (2007). Organization of sister origins and replisomes during multifork DNA replication in Escherichia coli. Submitted. The paper is not available in DUO.|
|2. Fossum S, De Pascale G, Weigel C, Messer W, Donadio S and Skarstad K (2007). A robust screen for novel antibiotics: Specific knockout of the initiator of bacterial DNA replication. Submitted. The paper is not available in DUO.|
|3. Odsbu I, Klungsøyr HK, Fossum S, Skarstad K (2005). Specific N-terminal interactions of the E.coli SeqA protein are required to form multimers that restrain negative supercoils and form foci. Genes Cells, 10:1039-1049 The paper is not available in DUO. The published version is available at: http://dx.doi.org/10.1111/j.1365-2443.2005.00898.x|
|4. Fossum S, Søreide S, and Skarstad K. (2003). Lack of SeqA focus formation, DNA binding and proper protein multimerization in the Escherichia coli sequestration mutant seqA2. Mol Microbiol, 47(3):619-32 The paper is not available in DUO. The published version is available at: http://dx.doi.org/10.1046/j.1365-2958.2003.t01-1-03329.x|