spo0J is required for normal chromosome segregation as well as the initiation of sporulation in Bacillus subtilis

Abstract

The spoOJ gene of BaciUlus subtilis is required for the initiation of sporulation. We show that the sporulation defect caused by null mutations in spoOJ is suppressed by a null mutation in the gene located directly upstream from spoOJ, soj (suppressor of spoOJ). These results indicate that Soj inhibits the initiation of sporulation and that SpoOJ antagonizes that inhibition. Further genetic experiments indicated that Soj ultimately affects sporulation by inhibiting the activation (phosphorylation) of the developmental transcription factor encoded by spoOA. In addition, the temperature-sensitive sporulation phenotpe caused by the fts4279 (spoIIN279) mutation was partly suppressed by the soj null mutation, indicating that FtsA might also affect the activity of Soj. Soj and SpoOJ are known to be similar in sequence to a family of proteins involved in plasmid partitioning, including ParA and ParB of prophage P1, SopA and SopB of F, and IncC and KorB of RK2. spoOJ was found to be required for normal chromosome partitioning as well as for sporulation. spoOJ null mutants produced a significant proportion of anucleate cells during vegetative growth. The dual functions of SpoOJ could provide a mechanism for regulating the initiation of sporulation in response to activity of the chromosome partition machinery.The generation of new cells during growth or development requires proper replication, repair, and segregation of chromosomes. Conditions that perturb chromosome replication or damage DNA often inhibit cell cycle progression or development through checkpoint mechanisms. Such regulatory mechanisms are present in organisms as diverse as bacteria, yeasts, and mammals and act to prevent the unproductive formation of cells lacking intact chromosomes.Spore formation by the gram-positive bacterium Bacillus subtilis is a developmental process requiring two different cell types. Each cell type has an intact chromosome and a characteristic and distinct pattern of gene expression. The two cell types are created by formation of an asymmetric division septum early during development. The smaller cell, known as the forespore, develops into the mature spore while enclosed in the larger mother cell (30).One of the most important early events necessary for the initiation of sporulation is the activation of the developmental transcription factor encoded by spoOA. SpoOA is activated by phosphorylation, and SpoOA-P induces expression of several genes, including spoIL4, spoIIE, and spoIIG (4,6,46,47,55,58) that are essential for sporulation and the establishment of cell type-specific gene expression (30). Phosphorylation of SpoOA requires histidine protein kinases, i.e., KinA, -B, and -C (2, 27, 38, 56), and two phospho-transfer proteins, SpoOF and SpoOB (7). The histidine protein kinases autophosphorylate on a histidine residue, and phosphate is transferred to SpoOF, and then from

DNA polymerase template switching at specific sites on the φ29 genome causes the in vivo accumulation of subgenomic φ29 DNA molecules

“…Recently, important progress has been made in the understanding of the active chromosomal segregation process in vegetative and sporulating B. subtilis cells (Ireton et al, 1994;Sharpe and Errington, 1996;Glaser et al, 1997;Lin et al, 1997;Webb et al, 1997). It has been shown that 30% of the chromosome containing oriC has a centromere-like function that, during binary fission and sporulation, migrates actively towards the cell poles (Sharpe and Errington, 1996;Wu and Errington, 1994;Glaser et al, 1997;Webb et al, 1997).…”

“…It has been shown that 30% of the chromosome containing oriC has a centromere-like function that, during binary fission and sporulation, migrates actively towards the cell poles (Sharpe and Errington, 1996;Wu and Errington, 1994;Glaser et al, 1997;Webb et al, 1997). Because accurate segregation is affected in spoOJ mutant strains (Ireton et al, 1994;Sharpe and Errington, 1996), the SpoOJ protein was a prime candidate for being involved in the chromosomal segregation process. Using in vivo cross-linking techniques, the SpoOJ protein was recently shown to be associated with short DNA sequences containing the same DSE (Lin and Grossman, 1998) that is repeatedly present in the 29 genome.…”

“…In particular, after a few rounds of phage DNA replication the large amounts of DSEs available on the phage DNA molecules may efficiently titrate out SpoOJ and possibly other host-encoded proteins required for efficient chromosome segregation. In this respect it is interesting to note that SpoOJ is also involved in regulation of sporulation (Ireton et al, 1994). An alternative effect of the additive partitioning sites on the 29 genome therefore could be that, because of sequestering of the SpoOJ protein, development of the host cell to differentiate into sporulation is prevented, thereby keeping the cell in a state optimal for the infectious cycle of the phage.…”

“…The following B. subtilis strains were used: 110NA (trpC2, spoOA3, su ¹ ) (Moreno et al, 1974); JH642 (trpC2, pheA1 ) (Perego et al, 1988); AG1468 (JH642, ⌬spoOJ ::spc) (Ireton et al, 1994); AG1505 (JH642, ⌬soj-spoOJ ::spc) (Ireton et al, 1994). The last three strains were kindly provided by A. D. Grossman.…”

“…Fifth, sequences nearly identical to the mentioned DSE (maximum of two mismatches) are not only found several times on the chromosome of B. subtilis (Table 2) Subgenomic molecules also accumulate in spoOJ deletion strains of B. subtilis Surprisingly, as mentioned above, the DSE was found to be present in the spoOJ gene of the B. subtilis host chromosome. Besides an essential role in the development of sporulation, the SpoOJ protein is also involved in chromosome segregation (Ireton et al, 1994). By in vivo crosslinking experiments the SpoOJ protein was shown to be associated with this DSE (Lin and Grossman, 1998).…”