## 2 Problems with Current Explanations of Initiation Control  
During state-state growth, a bacterium like _E. coli_[31] must initiate chromosome replication at the same rate as its mass increases since failure to do so would lead to cells either filling up with DNA or lacking it. Over forty years ago, two classes of models began to vie with one another to explain the initiation of chromosome replication (a very different type of model received less attention [32]). The first class, which had been used successfully for plasmids, invoked a repressor acting at the origin of replication, _oriC_, that was diluted by growth to allow initiation every mass doubling [33]. The second class invoked an activator of replication that accumulated to a critical level to trigger initiation during the mass doubling; this class of model benefited in particular from an analysis of data [34,35] that suggested that the bacterium initiated replication at a constant mass--the initiation mass--regardless of growth rate [36]. The DnaA protein became identified with this activator [37]. Unfortunately for simple models based on the dilution of a repressor or the accumulation of an activator with respect to an origin of replication, it was found possible to create minichromosomes (small plasmids that replicated using _oriC_) [38]. Worse, the _E. coli_ cell cycle was found to be unperturbed by the presence of scores of these minichromosomes that even replicated in synchrony with the chromosome itself [39]. An adapted version of the model of DnaA as the activator was proposed in which DnaA first binds to many high affinity sites throughout the chromosome before finally binding to lower affinity sites in _oriC_ to trigger initiation [40]; hence, the chromosome itself became part of the timing mechanism. The original idea of the initiation mass came into question when it was found that the initiation mass varies at different growth rates and it was concluded that \"the timing of initiation is not governedby a direct connection between mass accumulation and the molecule(s) determining initiation of replication\" [41; 42; 43]; more recent studies using cell size mutants have led to the suggestion that the initiation mass in _E. coli_ depends on the amount of DnaA rather than on its concentration and, moreover, that the initiation mass model must be different for _B. subtilis_[44]. Models for explaining initiation mass also have to contend with the decrease in the mass of DNA (but unaltered rates of growth and division) when _dnaAts_ mutants are grown at semi-permissive temperatures, which gave rise to the idea that DnaA controls the number of initiations but not their timing [45]. Worse still, the initiation of replication of minichromosomes is apparently normal in a strain in which the chromosome itself replicates at random using the origin of replication of a plasmid [46]; this effectively disqualifies the chromosome itself from being the essential element in the timer. Most seriously of all, perhaps, the presence of an extra _oriC_ in the _E. coli_ chromosome, which results in two origins that function at the same time, has relatively little effect on the cell cycle [47; 48]. Recent models for initiation have been based on the proportion of the DnaA 'initiator' protein in the ATP-DnaA form, which is active in initiation, or the ADP-DnaA form, which is not. There are several hypotheses to explain how the level of active DnaA might increase cyclically to trigger initiation [49]. Lipids have long been known to be involved in initiation [50] and one possibility is that anionic phospholipids might promote conversion from ADP-DnaA to ATP-DnaA _in vivo_ as _in vitro_[51]. Another, complementary, possibility is that specific DNA sequences promote this conversion via the formation of homomultimers of the DnaA protein [52]. Following initiation, removal of DnaA may also be achieved by DnaA binding to sequences in the _datA_ locus [53].