Thymineless death in Escherichia coli thyA mutants growing in the absence of thymidine (dT) is preceded by a substantial resistance phase, during which the culture titer remains static, as if the chromosome has to accumulate damage before ultimately failing. Significant chromosomal replication and fragmentation during the resistance phase could provide appropriate sources of this damage. Alternatively, the initial chromosomal replication in thymine (T)-starved cells could reflect a considerable endogenous dT source, making the resistance phase a delay of acute starvation, rather than an integral part of thymineless death. Here we identify such a low-molecular-weight (LMW)-dT source as mostly dTDP-glucose and its derivatives, used to synthesize enterobacterial common antigen (ECA). The thyA mutant, in which dTDP-glucose production is blocked by the rfbA rffH mutations, lacks a LMW-dT pool, the initial DNA synthesis during T-starvation and the resistance phase. Remarkably, the thyA mutant that makes dTDP-glucose and initiates ECA synthesis normally yet cannot complete it due to the rffC defect, maintains a regular LMW-dT pool, but cannot recover dTTP from it, and thus suffers T-hyperstarvation, dying precipitously, completely losing chromosomal DNA and eventually lysing, even without chromosomal replication. At the same time, its ECA+thyA parent does not lyse during T-starvation, while both the dramatic killing and chromosomal DNA loss in the ECA-deficient thyA mutants precede cell lysis. We conclude that: 1) the significant pool of dTDP-hexoses delays acute T-starvation; 2) T-starvation destabilizes even nonreplicating chromosomes, while T-hyperstarvation destroys them; and 3) beyond the chromosome, T-hyperstarvation also destabilizes the cell envelope.
Keywords: cell lysis; chromosome fragmentation; chromosome replication; dTDP-glucose; enterobacterial common antigen.