Conserved function and regulation of chromosomal replication origins in a -proteobacteria : Caulobacter crescentus and Rickettsia prowazekii

The aquatic Gram-negative organism, Caulobacter crescentus, is a member of the alpha (alpha) subclass of the proteobacteria. C. crescentus possesses an unique cell cycle with a complex developmental program in that the bacterium asymmetrically divides to produce a replication competent stalked cell and replication incompetent swarmer cell. Previously, a chromosomal fragment was shown to support autonomous plasmid replication. In addition, sequence analysis of this autonomous replicating sequence (ARS) 500 by fragment, termed Cori, exhibited some shared features characteristic of a typical replication origin. However, as not all identified ARS elements are chromosomal replication origins, it was therefore shown by two-dimensional (2-D) DNA agarose gel electrophoresis analysis that chromosomal replication is bi-directional and originates within the Cori region (Chapter 2). Tracing of the leading strand path of nascent DNA within the Cori region not only supported the 2-D gel analysis results, but also indicated that replication initiates from within Cori (Chapter 3). Thus, it is confirmed that Cori is the chromosomal replication origin in C. crescentus and that replication proceeds bi-directionally. Nascent replicated DNA is methylated by the methyltransferase CcrM, but transcription of ccrM is indirectly dependent on synthesis of glycerol-3-phosphate, a precursor for cell membrane phospholipid synthesis (Chapter 6). A feature unique to C. crescentus is the global response regulator CtrA that controls key events within the cell cycle such as chromosomal replication. CtrA represses replication in the swarmer cell by binding to five specific sites on Cori. In order to allow replication to initiate, CtrA is removed from the stalked cell by proteolytic degradation. Another interesting feature is that Cori resides within an unusual gene cluster. It has also shown that this unusual gene cluster is not unique to C. crescentus as the alpha-proteobacterium intracellular pa