To let go or not to let go: how ParA can impact the release of the chromosomal anchoring inCaulobacter crescentus

Abstract

ABSTRACTChromosomal maintenance is vital for the survival of bacteria. InCaulobacter crescentus, chromosome replication initiates atoriand segregation is delayed until the nearby centromere-like regionparSis replicated. Our understanding of how this sequence of events is regulated remains limited. The segregation ofparShas been shown to involve multiple steps including polar release from anchoring protein PopZ, slow movement, and fast ParA-dependent movement to opposite cell pole. In this study, we demonstrate that ParA’s competing attractions from PopZ and from DNA are critical for segregation ofparS. Interfering with this balance of attractions – by expressing a variant ParA-R195E unable to bind DNA and thus favoring interactions exclusively between ParA-PopZ – results in cell death. Our data revealed that ParA-R195E’s sole interactions with PopZ obstruct PopZ’s ability to release the polar anchoring ofparSresulting in cells with multipleparSloci fixed at one cell pole. We show that the inability to separate and segregate multipleparSloci from the pole is specifically dependent on the interaction between ParA and PopZ. Interfering with interactions between PopZ and the partitioning protein ParB, which is the interaction that anchorsparSat the cell pole, does not rescue the ability of cells to separate the fixedparSloci when expressingparA-R195E. Thus, ParA and PopZ appear to have a distinct conversation from ParB yet can impact the release of ParB-parSfrom the anchoring at the cell pole. Collectively, our results reveal that the initial steps in chromosome segregation are highly regulated.