HBsu Is Required for the Initiation of DNA Replication in Bacillus subtilis

Nucleoid-associated proteins (NAPs) help structure bacterial genomes and function in an array of DNA transactions, including transcription, recombination, and repair. In most bacteria, NAPs are nonessential in part due to functional redundancy. In contrast, in Bacillus subtilis the HU homolog HBsu is essential for cell viability. HBsu helps compact the B. subtilis chromosome and participates in homologous recombination and DNA repair. However, none of these activities explain HBsu's essentiality. Here, using two complementary conditional HBsu alleles, we investigated the terminal phenotype of the mutants. Our analysis revealed that cells without functional HBsu fail to initiate DNA replication. Importantly, when the chromosomal replication origin (oriC) was replaced with a plasmid origin (oriN) whose replication does not require the initiator DnaA, cells without HBsu initiated DNA replication normally. However, HBsu was still essential in this oriN-containing strain. We conclude that HBsu plays an essential role in the initiation of DNA replication, likely acting to promote origin melting by DnaA, but also has a second essential function that remains to be discovered. IMPORTANCE While it is common for a bacterial species to express multiple nucleoid-associated proteins (NAPs), NAPs are seldomly essential for cell survival. In B. subtilis, HBsu is a NAP essential for cell viability. Here, using conditional alleles to rapidly remove or inactivate HBsu, we show that the absence of HBsu abolishes the initiation of DNA replication in vivo. Understanding HBsu's function can provide new insights into the regulation of DNA replication initiation in bacteria.