Discovery of new mechanisms to control metabolism and homeostasis in Bacillus subtilis

Biosynthetic needs of a cell and the generation of precursors and energy in catabolic reactions must be faithfully balanced. Even in model organisms such as Bacillus subtilis, there are still important gaps in our knowledge. The key bottleneck in research is the lack of novel research hypotheses. New concepts and methodologies can help to develop such hypotheses. Here, we discuss how the introduction of proteome-wide protein-protein interaction mapping by in vivo cross-linking, the AI-mediated prediction of structure models for each protein, and the possibility to compare those models highly efficiently aid the development of novel hypotheses. Moreover, the focused use of suppressor screens can help to get new unbiased insights. We demonstrate how these approaches are applied to B. subtilis. Global cross-linking combined with the power of AI provided a testable hypothesis to unravel the long-standing open question of how iron is sensed in B. subtilis and related bacteria. This is of particular importance as iron is the growth-limiting factor for most bacterial pathogens. The isolation of suppressor mutants that are resistant to growth-inhibiting amino acids has identified novel amino acid exporters. Importantly, the corresponding genes belong to the most poorly expressed genes in B. subtilis, and they are only activated under selective pressure by mutations that affect corresponding transcription factors or the promoter regions of the exporter genes. As the approaches discussed here have only recently been brought to wide application, we can expect that they will be very fruitful in gaining a better understanding of metabolism and metabolic homeostasis.