Thesis

Caulobacter Crescentus SpotT is essential for preventing the over-initiation of DNA replication during fatty acid starvation

Caulobacter crescentus FabH is a cell cycle regulated -ketoacyl-acyl carrier protein synthase that initiates fatty acid elongation and is essential in rich PYE media. Depletion of FabH induces fatty acid starvation and a bacteriostatic condition in cells. Therefore, fatty acid synthesis serves as a checkpoint for cells that are about to begin cell division. Also, the accumulation of the master regulators DnaA, GcrA, and CcrM, is drastically reduced while CtrA accumulation is only slightly decreased. Although no significant changes are observed for gcrA transcription, dnaA transcription increases 2.5-fold and ctrA transcription increases 1.7-fold when FabH is depleted. Fatty acid starvation triggers the SpoT-dependent production of ppGpp, which presumably helps RNA polymerase to activate the transcription of genes that are beneficial under starvation conditions. Increased dnaA and ctrA transcription under fatty acid starvation is SpoT-dependent. In contrast, ccrM transcription decreases 2-fold under FabH-depletion conditions and is SpoT-independent. CtrA normally blocks the initiation of DNA replication, however, when FabH is depleted, and in the absence of SpoT, CtrA fails to accumulate; resulting in the buildup of multiple chromosomes. In sum, the miscoordination of cell cycle events, such as DNA replication, during FabH depletion, results from the absence of SpoT.

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