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Abstract

Although several proteins have been identified that facilitate
chromosome segregation in bacteria, no clear analogue of the mitotic
machinery in eukaryotic cells has been identified. In order to
investigate if recognizable patterns of segregation exist during the
cell cycle, we tracked the segregation of duplicated origin regions in
Bacillus subtilis for 60 min in the fastest practically achievable
resolution, achieving 10-s intervals. We found that while separation
occurred in random patterns, often including backwards movement,
overall, segregation of loci near the origins of replication was linear
for the entire cell cycle. Thus, the process of partitioning can be best
described as directed motion. Simulations with entropy-driven separation
of polymers synthesized by two polymerases show sudden bursts of
movement and segregation patterns compatible with the observed in vivo
patterns, showing that for Bacillus, segregation patterns can be modeled
based on entropic forces. To test if obstacles for replication forks
lead to an alteration of the partitioning pattern, we challenged cells
with chemicals inducing DNA damage or blocking of topoisomerase
activity. Both treatments led to a moderate slowing down of separation,
but linear segregation was retained, showing that chromosome segregation
is highly robust against cell cycle perturbation.
IMPORTANCE We have followed the segregation of origin regions on the
Bacillus subtilis chromosome in the fastest practically achievable
temporal manner, for a large fraction of the cell cycle. We show that
segregation occurred in highly variable patterns but overall in an
almost linear manner throughout the cell cycle. Segregation was slowed
down, but not arrested, by treatment of cells that led to transient
blocks in DNA replication, showing that segregation is highly robust
against cell cycle perturbation. Computer simulations based on
entropy-driven separation of newly synthesized DNA polymers can
recapitulate sudden bursts of movement and segregation patterns
compatible with the observed in vivo patterns, indicating that for
Bacillus, segregation patterns may include entropic forces helping to
separate chromosomes during the cell cycle.