Negative interplay between biofilm formation and competence in the environmental strains ofBacillus subtilis

Abstract

12 13 14 15a. These two authors contributed equally to this work. 16 (She contributed to about 50% of the results in the initial submission and the 17 revision; Hunter initiated the project and contributed to about 30% of the 18 results in the initial submission.) Abstract 31 Environmental strains of the soil bacterium Bacillus subtilis have valuable 32 applications in agriculture, industry, and biotechnology. They are capable of forming 33 robust biofilms and demonstrate excellent biological control activities in plant 34 protection. However, environmental strains are genetically less accessible, a sharp 35 contrast to the laboratory strains well known for their natural competence and a 36 limitation toward their application. In this study, we observed that robust biofilm 37 formation of the environmental strains greatly reduces the rate of competent cells 38 within the biofilm. By using the model strain 3610, we reveal a cross-pathway 39 regulation that allows biofilm matrix producers and competence-developing cells to 40 undergo mutually exclusive cell differentiation. We show that the competence 41 activator ComK represses the key biofilm regulatory gene sinI by directly binding to 42 the sinI promoter, thus blocking competent cells from simultaneously becoming 43 matrix producers. In parallel, the biofilm activator SlrR represses competence 44 through three distinct mechanisms, involving both genetic regulation and cell 45 morphological changes. We discuss potential implications of limiting competence in 46 a bacterial biofilm. 47 48 3 Importance 49The soil bacterium Bacillus subtilis is capable of forming robust biofilms, a 50 multicellular community important for its survival in the environment. B. subtilis also 51 exhibits natural competence, the ability of cells to acquire genetic materials directly 52 from the environment. By investigating competence development in situ during B. 53 subtilis biofilm formation, we reveal that robust biofilm formation, an important 54 feature of the environmental strains of B. subtilis, often greatly reduces the rate of 55 competent cells within the biofilm. We characterize a cross-pathway regulation that 56 allows cells associated with these two developmental events to undergo mutually 57 exclusive cell differentiation during biofilm formation. Finally, we discuss potential 58 biological implications of limiting competence in a bacterial biofilm. 59 60 Bacillus subtilis is a soil-dwelling, spore-forming bacterium widely present in 61 nature, and plays important roles in environment, agriculture, and industry. B. subtilis 62 is also a plant growth-promoting rhizobacterium (PGPR), and a biological control 63 agent that shows various beneficial activities in plant protection (1). Biological control 64 by B. subtilis is attributed to a number of important abilities of the bacterium, 65 including antibiotic production, inhibition of pathogenic fungi and parasites, induction 66 of plant systemic resistance, and formation of plant root-associated biofilms (2-4).

Identification of New Chromosomal Loci Involved in com Genes Expression and Natural Transformation in the Actinobacterial Model Organism Micrococcus luteus

“…A model such as this one was also proposed for the divergently overlapping comEA/EC and comER genes in B. subtilis [ 16 ]. In a more recent work [ 66 ], it has been shown that comER plays an important role in biofilm formation and sporulation, and that competence has a negative correlation with biofilm formation [ 67 ], supporting the stated hypothesis. When analyzing if the gene Mlut_14660 and the operon Mlut_14650/40/30 and their specific genetic context is conserved in other bacteria, we found that they are present and have the same or similar surroundings on the chromosomes of several different Actinobacteria genera (see Figure 5 ; only selection of chromosomal regions of related genera and species with similar gene arrangements depicted).…”

Extensive cellular multitasking within Bacillus subtilis biofilms

“…In combining reporter constructs for the 14 genes of interest, we created 182 strains; this included all 91 possible pairwise combinations of the 14 genes in each of the two color orientations (e.g., both P hag - YPet ∷ amyE , P tapA - mTurq ∷ lacA and P tapA - YPet ∷ amyE , P hag - mTurq ∷ lacA to control for differences in fluorescent protein expression levels). Only six such dual-reporter strain pairs ( hag-tapA ; hag-sspB ; tapA-sspB ; srfAA-tapA ; skfA-tapA ; and tapA-comGA ) have been explicitly examined in the literature previously (23, 29, 30, 32, 33). All strains containing dual reporters grew similarly to wild-type B. subtilis (Figure S3).…”