Supplementary Material for: Global Transcriptional Analysis of Bacillus licheniformis Reveals an Overlap between Heat Shock and Iron Limitation Stimulon

In this study, we characterized the heat shock stimulon of the important industrial microorganism Bacillus licheniformis using DNA microarrays. While sharing a high degree of homology with the closely related model organism Bacillus subtilis, the heat shock stimulon of B. licheniformis exhibited several novel and unexpected features. Most notably, heat shock in B. licheniformis resulted in decreased amounts of mRNA from the ytrABCEF operon, encoding a putative acetoin uptake system, and stimulated the transcription of purine biosynthesis and iron uptake genes. Unexpectedly, deletion of the ytrEF genes did not affect acetoin uptake, but increased heat sensitivity. To investigate the connection between heat stress and iron uptake further, we analyzed the iron limitation response of B. licheniformis by DNA microarrays and concluded that the response mostly involves the genes related to iron uptake and metabolism, while the only heat shock gene affected by iron limitation was clpE. We also attempted to delete the fur gene (encoding the ferric uptake repressor), but unexpectedly found it to be essential in B. licheniformis. Using the fluorescent protein-encoding reporter gene under control of the dhb promoter, which responded to both heat shock and iron-starvation, we confirmed the overlap between these responses.