Academic journal article Genetics

Static and Dynamic Factors Limit Chromosomal Replication Complexity in Escherichia Coli, Avoiding Dangers of Runaway Overreplication

Academic journal article Genetics

Static and Dynamic Factors Limit Chromosomal Replication Complexity in Escherichia Coli, Avoiding Dangers of Runaway Overreplication

Article excerpt

EUKARYOTIC and prokaryotic chromosomes differ in many important aspects (Kuzminov 2014), and one key difference lies in the spatio-temporal organization of chromosomal replication. In contrast to eukaryotes, which perform multibubble replication (Masai et al. 2010), most bacteria replicate their singular chromosome in the unibubble format by initiating bidirectional replication from a designated replication origin (oriC) (Sernova and Gelfand 2008; Leonard and Méchali 2013) and finishing replication within a broad termination zone (ter) (Mirkin and Mirkin 2007; Duggin et al. 2008). Eukaryotes always perform a single replication round in their chromosomes (Masai et al. 2010; Diffley 2011), keeping the ratio of maximally replicated to unreplicated DNA in the same chromosome (the "replication complexity index") strictly at 2 (Figure 1A). Due to the defined origin and terminus of prokaryotic chromosomes, chromosomal replication complexity in bacteria can be simply expressed as the ori/ter ratio (Figure 1B). Even though unique cell cycles in some bacteria, such as Caulobacter, also maintain a strict CRC = 2 (Collier 2012), bacterial cells are generally recognized for their ability to support multiple concurrent replication rounds within the same chromosome (Morigen et al. 2009).

In reality, under typical growth conditions the ori/ter ratio in exponentially growing bacterial cells is still ~2 (Bird et al. 1972; Bipatnath et al. 1998; Wang et al. 2007; Murray and Errington 2008; Rotman et al. 2009; Stokke et al. 2011), showing that bacterial cells, like eukaryotes, also prefer to deal with a single replication round in their chromosomes. However, due to the peculiarity of the prokaryotic chromosome cycle (Kuzminov 2013), when the rate of cell mass duplication surpasses the genome duplication rate [Escherichia coli cannot replicate its chromosome in less than ~40 min (Chandler et al. 1975; Bremer and Dennis 1996)], E. coli and some other bacteria are capable of initiating additional replication rounds in the same chromosome. For example, E. coli cells dividing every 20 min double their chromosomal replication complexity (ori/ter) to ~4 (Morigen et al. 2009), a state referred to as "multifork replication" (Cooper and Helmstetter 1968; Quinn and Sueoka 1970) (Figure 1B).

Is multifork replication observed only in the fastestgrowing bacterial cells? In fact, even with a modest rate of cell mass increase, inhibition of replication fork progressionwill cause bacterial cells to employ multi-fork replication. A classic condition in bacteria when the DNA replication rate lags behind that of cell mass accumulation is thymine limitation (when the limited availability of the DNA precursors dTTP reduces the replication rate) (Ahmad et al. 1998). Thyminelimited E. coli cells have 100% viability, shownormal growth rates, and divide on time, but their CRC increases to compensate for the slower-moving replication forks (Zaritsky et al. 2006). The system that regulates these extra initiations is proposed to be the one that determines the "eclipse period," a cell cycle phase of enforced origin inactivity following each replication initiation, lasting ~60% of the generation time (von Freiesleben et al. 2000; Olsson et al. 2002). By preventing closely spaced origin firing events, the eclipse phenomenon defines a minimal allowed distance between codirectional replication forks in the E. coli chromosome.

Theoretical considerations of the eclipse phenomenon predicted a natural upper limit for increased chromosomal replication complexity in E. coli replicating under conditions of thymine limitation of two replication rounds per chromosome (CRC=4, Figure 1B) before accumulation of inhibitory chromosomal problems of unknown nature (Zaritsky et al. 2006, 2007). For example, initiation from multiple origins in the same cell becomes asynchronous when the eclipse period is reduced below half-a-generation time (Olsson et al. 2003). Indeed, maximal reported ori/ter values in thymine-limited cells generally do not go over 4 (Bird et al. …

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