1 Introduction
Bacillus species like B. subtilis , B.
amyloliquefaciens , B. licheniformis , are Gram-positive,
spore-forming bacteria widely distributed in soil, water, air, etc.
During times of environmental insult such as poor nutrient, the bacteria
undergo developmental changes leading to quorum sensing responses, by
which the cells form multicellular communities known as biofilm
[1,2]. Now it is known that upon harsh environment, the cells
produce a secondary metabolite called surfactin, which is a nonribosomal
lipopeptide biosynthesized by a multienzyme system encoded by the gene
cluster srfA [3-5], as a signal to trigger biofilm formation
[6-8].
Generally, harsh environments lead to a change of cellular redox state,
which is an important parameter to affect surfactin production and
biofilm formation [9]. For example, B. subtilis can switch
from an unicellular state to a multicellular state of biofilm via
responding to a redox switch1. The
NADH/NAD+ ratio, or another indicator of the cellular
redox potential, is sensed by regulatory pathways to control features of
colony wrinkling. Also, colony development can both respond to and
affect redox homeostasis [9]. For instance, isocitrate dehydrogenase
is capable of regulating biofilm formation by modulating intracellular
redox homeostasis [10]. Also, the cells are capable of transferring
glucose to acetoin as a response to enough carbon source and reducing
power, which are essential for robust biofilm growth and conserving
redox balance [11,12].
Despite importance of redox, we know little about how the shifts in
redox state regulates biosynthesis of surfactin for triggering biofilm
formation. Surfactin production can be regulated by the cellular redox
status via some regulatory proteins such as Spx and PerR. Spx interacts
with RNA polymerase to control transcription in response to oxidative
stress [13,14], which plays a key role in maintaining the redox
homeostasis exposed to disulfide stress, ensuring an immediate response
to oxidative stress [15]. The redox state of cytoplasm is the major
effector driving Spx activation, which activity is enhanced by
reversible formation of a disulfide bond and thereby directly modulated
by the intracellular redox status [16]. The transcription ofspx is negatively regulated by the repressor PerR and is
positively regulated by a sigma factor of SigB [15]. SigB is a
general stress response transcription factor [17], and PerR senses
H2O2 to mediate adaptation to peroxide
stress. PerR also represses oxidative stress resistance genes including
the catalase katA , alkylhydroperoxide reductase ahpC , iron
uptake repressor (fur ), and perR itself [18,19]. After
oxidation PerR is inactivated, so the oxidative response genes includingspx are capable of being successfully expressed. Thereby, PerR
plays an auxiliary role in coordinating the disulfide stress responses
[19].
Previously, we reported that mutation of srfA resulted in a very
seriously defective growth and biofilm formation but could be restored
by glucose in B. amyloliquefaciens WH1 [20]. Glucose an its
intermediate metabolites such as NADH are reductive substances, we
hypothesized that these reductants might influence biofilm formation via
some regulators such as Spx and PerR to regulate surfactin production as
a response to the change of ceullular redox homeostasis. In this study,
we characterized the interplay among redox state, surfactin production
and biofilm formation, and found the redox status influenced by oxidants
and reductants could affect biofilm formation via regulation of
surfactin production in B. amyloliquefaciens . Moreover,
some reductants such as glucose could also influence biofilm formation
by a surfactin-independent way in this bacterium.