Role of Ori in Thermococcus barophilus

Abstract

SummaryThe mechanisms underpinning replication of genomic DNA in Archaea have recently been challenged. Species belonging to two different taxonomic orders grow well in the absence of an origin of replication, challenging the role of the replication origin in these organisms. Here, we pursue the investigation of the particular way some archaea manage their DNA replication with Thermococcus barophilus and the role of Ori in this Archaea. Surprisingly we discovered that T. barophilus uses its Ori all along the growth curve with marked increase at the end of exponential phase. Through gene deletion, we show that Ori utilization requires Cdc6, and that origin deletion results in increased time in lag phase and a moderate decrease of growth rate in mutants. The number of chromosomes are quite similar between both strains during exponential and early stationary phases but differs after 24h of growth where ΔTbOriC has only 6 chromosomes/cell compared to 10 for the reference strain (WT). Following 1hr of growth in fresh media, ΔTbOriC strains contains 3 chromosome copies/cell, whereas the WT contains only 1. We hypothesize that the T. barophilus might degrade DNA to obtain energy to start replication and cell division, whereas the ΔTbOriC must maintain more chromosomal copies in order to initiate DNA replication in the absence of an origin or replication. Finally, we analyzed the role of Ori at temperatures above or below the optimal temperature, revealing that Ori is important to start growth at those temperatures, suggesting that replication origins may be involved in stress response.