Genetic analysis of the chromosome segregation protein Spo0J of Bacillus subtilis: evidence for separate domains involved in DNA binding and interactions with Soj protein

Abstract

SummarySpo0J and Soj belong to the ParB/ParA family of proteins involved in chromosome and plasmid segregation in bacteria. In Bacillus subtilis, Spo0J protein binds to several specific sites, parS, located on both sides of the origin of DNA replication, oriC, and apparently self-associates to form large discrete foci visible by fluorescence microscopy. Soj protein forms large 'patches' probably associated with the nucleoid, which can undergo dynamic, co-operative jumping from nucleoid to nucleoid in the presence of Spo0J. Patches of Soj protein somehow help to bring about the condensation of Spo0J foci. Soj is also a negative regulator of transcription. In the absence of Spo0J, Soj is statically distributed on each of the nucleoids in the cell and blocks the transcription of several sporulation genes. To analyse the functional interaction between Spo0J and Soj further, we have constructed and studied a collection of spo0J mutants. Most of the mutants completely prevent Spo0J from interacting with DNA. One mutation impairs the formation of compact Spo0J foci and simultaneously results in loss of Soj movement. We also isolated one spo0J mutant, in which the frequency of Soj internucleoid oscillation is highly increased. Both mutations affecting the interaction with Soj lie in the N-terminal coding part of spo0J, whereas the substitutions affecting DNA binding lie in the mid-to C-terminal coding region.

An expanded view of bacterial DNA replication

“…To visualize the location of the replication origin, the oriC-labeling system based on the lacO array (18) or on the Spo0J-green fluorescent protein (GFP) fusion (19) were introduced into the wild-type B. subtilis strain (168) and the yabA null mutant (168⌬yabA::phleo). The lacO array was amplified and the strains were grown for microscopy essentially as described in ref.…”

Evolution of Eukaryotic Cysteine Sulfinic Acid Reductase, Sulfiredoxin (Srx), from Bacterial Chromosome Partitioning Protein ParB

“…In accord with the observed evolutionary conservation, mutation of R80 of B. subtilis Spo0J has been shown to impair chromosome segregation. 34 Phylogenetic analysis and evolutionary implications. To gain insight into the evolution of the Srx-ParB superfamily, we constructed a phylogenetic tree from the multiple alignment shown in Figure 1.…”

Transcription Regulation ofezrAand Its Effect on Cell Division ofBacillus subtilis

“…In the initiation of cell division, the tubulin-like cell division protein FtsZ polymerizes at the midcell into a ring structure that is required for subsequent recruitment of other cell division proteins and assembly of the cell division machinery (7,10,29,36,40). Temporally, the division process is tightly coupled to chromosome replication, chromosome segregation, and cell growth to ensure that both daughter cells inherit complete genomes and are of appropriate size and shape (12,16,17,35).Two mechanisms, nucleoid occlusion and the Min system, are involved in selection of the correct mid-cell site for cell division (1,12,17,30). Nucleoid occlusion, although poorly defined, was revealed by the observation that cell division is largely inhibited in the vicinity of the nucleoid of cells in which DNA replication and/or segregation is perturbed (33).…”

“…Two mechanisms, nucleoid occlusion and the Min system, are involved in selection of the correct mid-cell site for cell division (1,12,17,30). Nucleoid occlusion, although poorly defined, was revealed by the observation that cell division is largely inhibited in the vicinity of the nucleoid of cells in which DNA replication and/or segregation is perturbed (33).…”