kinA mRNA is missing a stop codon in the undomesticated Bacillus subtilis strain ATCC 6051

Abstract

Several features distinguish laboratory and undomesticated strains of Bacillus subtilis. For example, unlike the laboratory strain 168, the undomesticated strain ATCC 6051 is deficient in sporulation in a rich sporulation medium, 2¾ SG. ATCC 6051 cannot induce transcription of the spoIIG operon, suggesting that this strain has a defect in initiation of sporulation. To determine the genetic difference between 168 and ATCC 6051, the DNA region responsible for the Spo " phenotype was transferred to strain 168. Genetic mapping and DNA sequencing analysis revealed that a stop codon (TAA) for kinA in 168 is replaced with Lys (TAT) in ATCC 6051, making the kinA open reading frame 201 bp longer in the undomesticated strain ATCC 6051. Introduction of kinA from strain 168 restored sporulation in ATCC 6051, indicating that the difference in kinA is responsible for the Spo " phenotype of ATCC 6051. A potential rindependent terminator is located upstream of a stop codon for the extended kinA open reading frame in ATCC 6051. Northern blot analysis showed that transcription of kinA terminated at this terminator, and kinA mRNA is missing a stop codon in ATCC 6051. Moreover, deletion of tmRNA suppresses the sporulation defect in ATCC 6051. These observations indicate that in ATCC 6051 the absence of a stop codon in kinA mRNA affects sporulation, probably by leading to rapid degradation of KinA via the trans-translation process. In ATCC 6051, the kinA mutation affects sporulation but not other Spo0A-dependent phenomena such as biofilm formation, which can be activated by a low level of Spo0A~P. This is due to the fact that KinA activity is kept low during the exponential phase via transcriptional and post-translational regulation. Thus, the stop-codon-less kinA probably affects only sporulation. DNA sequencing of 30 B. subtilis strains revealed that another strain also produces stop-codon-less kinA mRNA. This observation suggests that the lack of a stop codon for kinA mRNA may give rise to a selective advantage under certain conditions. INTRODUCTIONThe Gram-positive soil bacterium Bacillus subtilis makes spores in response to nutrient depletion. Initiation of sporulation is controlled by the multi-component phosphotransfer system, the phosphorelay (Hoch, 1991(Hoch, , 1993. In response to nutrient depletion, a histidine kinase phosphorylates itself, and then this phosphate residue is sequentially transferred to Spo0F, Spo0B and finally Spo0A. Spo0A is a member of a response-regulator family of transcriptional regulators, and phosphorylated Spo0A (Spo0A~P) activates transcription of early sporulation genes.Spo0A~P directly regulates about 120 genes required for various stationary-phase phenomena including sporulation, competence development, production of degradative enzymes, cannibalism and biofilm formation (Molle et al., 2003). Spo0A-regulated genes can be classified into two groups based on Spo0A dose dependency: low-threshold genes require a low level of Spo0A~P for activation or repression of transcription, whereas high-thres...

DegU and Spo0A Jointly Control Transcription of Two Loci Required for Complex Colony Development byBacillus subtilis

“…However, deletion of degS from ATCC 6051 and NCIB3610 results in different swarming motility phenotypes, indicating that there are differences in the DegU regulons of these two strains (23,42). Additionally, ATCC 6051 has recently been found to differ from strain NCIB3610 (used in this study) at at least two locations in its genome, at least one of which influences biofilm formation (24,25). Therefore, it was necessary to establish whether YuaB activates complex colony development by strain NCIB3610 and to assess if, like yvcA transcription, yuaB transcription was activated by DegUϳP, as DegU is known to have regulatory activity in both its nonphosphorylated and phosphorylated states (10).…”

trans-translation-mediated tight regulation of the expression of the alternative ribosome-rescue factor ArfA in Escherichia coli

“…There is a clue to this question. Kobayashi et al (2008) reported that the KinA protein was degraded in an SsrA-dependent manner in Bacillus subtilis when a base-substitution mutation in kinA ORF eliminated the stop codon. Although degradation of the KinA protein is just a result of mutation, it is intriguing to speculate that regulatory system for ArfA expression has its origin in such a mutation-promoted trans-translation.…”

Sporulation during Growth in a Gut Isolate of Bacillus subtilis

“…However, it seems most likely that the efficiency and kinetics of sporulation is not always increased but rather adjusted to specific niches. One wild isolate of B. subtilis, strain NCIB3610, bears a stop codon within the kinA gene, and is conditional for sporulation (73). In MSgg medium, NCIB3610 forms aerial structures, called fruiting bodies, that are preferential sites of spore formation (74), but it sporulates inefficiently in DSM (73).…”

“…One wild isolate of B. subtilis, strain NCIB3610, bears a stop codon within the kinA gene, and is conditional for sporulation (73). In MSgg medium, NCIB3610 forms aerial structures, called fruiting bodies, that are preferential sites of spore formation (74), but it sporulates inefficiently in DSM (73). Spore differentiation is ecologically beneficial, as spores will remain viable under harsh environmental conditions, but it is a time-consuming and energetically costly process, requiring the differential expression of over 10% of the genome over a period of 7 to 8 h (75,76).…”