_{Plasmid segregation in Haloferax volcanii is mediated by a hybrid partitioning system}

Proper segregation of the genome, a fundamental process across all domains of life has not been extensively studied in Archaea. Archaea show a high degree of variability in the organisation and copy number of chromosomes and plasmids. The archaeon Haloferax volcanii is a polyploid organism containing two large and two small plasmids as part of its genome. It is generally accepted that genomic segregation is carried out randomly due to its polyploidy.

However, we have identified multiple two-gene operons across its genome; one gene encoding a ParA like protein and the other encoding unknown proteins. One such operons in the large plasmid pHV3 contains the loci hpaB and hpaA (Haloarchaeal partitioning protein A/B). We show phylogenetic and functional similarities between the operon and the ParABS DNA segregation system from bacteria. The predicted structure and conserved domains in the sequence of HpaA suggest that it is a ParA-like protein having a bacterial origin, acquired via horizontal gene transfer. HpaB is a ParB analogue of archaeal origin capable of forming foci in the cytosol. HpaA is necessary for the proper spatial distribution of the HpaB foci. Single particle tracking shows that HpaB regulates the dynamics of HpaA. Comparative qPCR suggests HpaB is essential for maintaining the plasmid to chromosome ratio.

We thus propose a bacterial-archaeal hybrid plasmid segregating system in H. volcanii comprising of a repurposed bacterial protein (HpaA) and a customized archaeal DNA adapter (HpaB); the HpaAB system. We further explore how it could be especially necessary for two-dimensional segregation of its genome as H. volcanii cells form flat disks in middle to late logarithmic phase of growth. Regular spatial distribution of all genome copies ensures proper genome segregation irrespective of the plane of division, which has been shown to not be influenced by the location of DNA in H. volcanii. We suggest that segregation of its genome depends on a two-dimensional distribution of all its copies rather than a directional movement across the plane of division.