Credit: Eshel Ben-Jacob Bacterial colonies cultured on agar avoid each other when forced to compete for nutrients, but the mechanism behind the observed growth inhibition has been unclear. Now a new study by collaborating scientists at UC San Diego, University of Texas and Tel Aviv University has explored the behaviour of Paenibacillus dendritiformis cultures, identifying the factors responsible.
The team found that it was not a shortage of food that halted the growth. They found nutrients in the no-man’s land between the colonies, but also a protein that wasn’t present elsewhere on the dish. When a sample of the purified protein was introduced to a fresh dish inoculated with P. dendritiformis, the bacteria formed a lopsided colony that shied away from the spot. In addition, the bacteria at the edge of the colony closest to the suspect protein were dead.
Analysis of the secretions from P. dendritiformis identified the protease, subtilisin, and a 12 kDa protein, termed sibling lethal factor (Slf). Whilst subtilisin promotes growth and expansion of P. dendritiformis colonies, Slf lyses the bacterial cells in culture. Slf is encoded by a gene belonging to a large family of bacterial genes of unknown function, and the gene is predicted to encode a protein of approximately 20 kDa. The team generated recombinant 20 kDa protein, which was found to be inactive. Exposure to subtilisin, however, resulted in cleavage to the active, 12 kDa form. The experimental results, combined with mathematical modelling, show that subtilisin regulates growth of the colony. Below a threshold concentration subtilisin promotes colony growth and expansion, but once it exceeds a threshold, as occurs at the interface between competing colonies, Slf is then secreted into the medium to rapidly reduce cell density by lysis of the bacterial cells. The presence of genes encoding homologs in other bacterial species suggests that this mechanism for self-regulation of colony growth might not be limited to P. dendritiformis.
The study is published in PNAS.