S-adenosylmethionine decarboxylase-dependent gene expression in planktonic Bacillus subtilis NCIB3610

The polyamine spermidine is not required for normal planktonic growth of Bacillus subtilis but is essential for robust biofilm formation. In a spermidine-deficient mutant of B. subtilis, the structural analogue norspermidine but not homospermidine restored biofilm formation. Intracellular biosynthesis of another spermidine analogue aminopropylcadaverine from exogenously supplied homoagmatine also restored biofilm formation. The differential ability of C-methylated spermidine analogues to replace the function of spermidine in biofilm formation indicated that the aminopropyl side of spermidine is more sensitive to C-methylation. Together, these data indicate that the aminopropyl side of spermidine is essential for its function in biofilm formation, and that the length and symmetry of the molecule is not critical. Transcriptomic analysis of a spermidine-depleted speD mutant of B. subtilis uncovered a nitrogen, methionine and S-adenosylmethionine sufficiency response, resulting in repression of gene expression related to purine catabolism, methionine and S-adenosylmethionine biosynthesis, methionine salvage, and there were indications that membrane status was altered. Consistent with the requirement for spermidine in biofilm formation, expression of the operons for production of the exopolysaccharide and TasA protein components of the biofilm matrix was reduced, as was expression of regulator SinR antagonist slrR. Single-cell analysis indicated that the effect of spermidine depletion was to decrease the number of cells expressing the biofilm matrix operons. Deletion of sinR or ectopic expression of slrR in the spermidine-deficient ΔspeD background restored biofilm formation, indicating that spermidine is required to promote expression of the biofilm regulator slrR. The study consists of two identical experiments separated in time by one week. The first experiment consists of a comparison of gene expression of the parental Bacillus subtilis NCIB3610 strain with a speD mutant (S-adenosylmethionine decarboxylase) that produces a spermidine auxotrophic strain. Both strains were grown in MSgg polyamine-free minimal medium at 37C to an OD of 0.5. For each strain, three independent biological replicates were grown (3610-1, 3610-2, 3610-3 and speD-1,speD-2, speD-3) and analyzed separately. The second experiment was identical and performed one week later with samples 3610-4, 3610-5, 3610-6, speD-4, speD-5 and speD-6.