Thymineless death (TLD), a phenomenon in which thymine auxotrophy becomes lethal
when cells are starved for thymine, can be prevented by rifampicin addition. In this work,
we have obtained evidence indicating that among the effects of RNA polymerase
inhibition by rifampicin, the inhibition of chromosomal replication initiation is the process
responsible for TLD suppression. We show that diminishing, abolishing or increasing the
transcription level around oriC alleviates TLD by limiting the chromosomal initiation
capacity (ChIC) generated under thymine starvation. TLD suppression was eliminated by
DnaA inactivation or the deletion in the left half of the replication origin or deletion of the
DnaA-boxes located in the right half of the oriC region. In accordance with these data,
two-dimensional DNA electrophoresis gels showed an accumulation of simple-Y and
bubble arc replication intermediates, as well as recombination structures at the oriC
region during thymine starvation. None of these structures were observed under genetic
or physiological conditions that suppress TLD. These results demonstrate that abortive
initiations of chromosome replication occur under thymine starvation, supporting the idea
that these events are a critical target for the lethal damage caused by thymine starvation
and that results in TLD.
