Hierarchical mutational events compensate for glutamate auxotrophy of a B acillus subtilis gltC mutant

• Published in: Environmental microbiology reports Vol. 9; no. 3; pp. 279 - 289
• Main Authors: Dormeyer, Miriam, Lübke, Anastasia L., Müller, Peter, Lentes, Sabine, Reuß, Daniel R., Thürmer, Andrea, Stülke, Jörg, Daniel, Rolf, Brantl, Sabine, Commichau, Fabian M.
• Format: Journal Article
• Language: English
• Published: 01-06-2017
• ISSN: 1758-2229; 1758-2229
• DOI: 10.1111/1758-2229.12531

Summary
Glutamate is the major donor of nitrogen for anabolic reactions. The Gram‐positive soil bacterium
Bacillus subtilis
either utilizes exogenously provided glutamate or synthesizes it using the
gltAB
‐encoded glutamate synthase (GOGAT). In the absence of glutamate, the transcription factor GltC activates expression of the GOGAT genes for glutamate production. Consequently, a
gltC
mutant strain is auxotrophic for glutamate. Using a genetic selection and screening system, we could isolate and differentiate between
gltC
suppressor mutants in one step. All mutants had acquired the ability to synthesize glutamate, independent of GltC. We identified (i) gain‐of‐function mutations in the
gltR
gene, encoding the transcription factor GltR, (ii) mutations in the promoter of the
gltAB
operon and (iii) massive amplification of the genomic locus containing the
gltAB
operon. The mutants belonging to the first two classes constitutively expressed the
gltAB
genes and produced sufficient glutamate for growth. By contrast, mutants that belong to the third class appeared most frequently and solved glutamate limitation by increasing the copy number of the poorly expressed
gltAB
genes. Thus, glutamate auxotrophy of a
B. subtilis gltC
mutant can be relieved in multiple ways. Moreover, recombination‐dependent amplification of the
gltAB
genes is the predominant mutational event indicating a hierarchy of mutations.