Sodium Lactate Negatively Regulates Shewanella putrefaciens CN32 Biofilm Formation via a Three-Component Regulatory System (LrbS-LrbA-LrbR)

Abstract

The capability of biofilm formation has a major impact on the industrial and biotechnological applications of Shewanella putrefaciens CN32. However, the detailed regulatory mechanisms underlying biofilm formation in this strain remain largely unknown. In the present report, we describe a three-component regulatory system which negatively regulates the biofilm formation of S. putrefaciens CN32. This system consists of a histidine kinase LrbS (Sputcn32_0303) and two cognate response regulators, including a transcription factor, LrbA (Sputcn32_0304), and a phosphodiesterase, LrbR (Sputcn32_0305). LrbS responds to the signal of the carbon source sodium lactate and subsequently activates LrbA. The activated LrbA then promotes the expression of lrbR, the gene for the other response regulator. The bis-(3=-5=)-cyclic dimeric GMP (c-di-GMP) phosphodiesterase LrbR, containing an EAL domain, decreases the concentration of intracellular c-di-GMP, thereby negatively regulating biofilm formation. In summary, the carbon source sodium lactate acts as a signal molecule that regulates biofilm formation via a three-component regulatory system (LrbS-LrbA-LrbR) in S. putrefaciens CN32.IMPORTANCE Biofilm formation is a significant capability used by some bacteria to survive in adverse environments. Numerous environmental factors can affect biofilm formation through different signal transduction pathways. Carbon sources are critical nutrients for bacterial growth, and their concentrations and types significantly influence the biomass and structure of biofilms. However, knowledge about the underlying mechanism of biofilm formation regulation by carbon source is still limited. This work elucidates a modulation pattern of biofilm formation negatively regulated by sodium lactate as a carbon source via a three-component regulatory system in S. putrefaciens CN32, which may serve as a good example for studying how the carbon sources impact biofilm development in other bacteria.KEYWORDS Shewanella putrefaciens CN32, biofilm, c-di-GMP, sodium lactate, threecomponent regulatory system B iofilms are described as the aggregates of microorganisms in which cells are embedded in a self-produced matrix that are adherent to each other or a surface (1, 2). The ability to form biofilms is a significant factor for the survival of bacteria in stressful environments. Biofilms are produced by bacteria to escape antibiotics, oxidative agents, metals, and nutrient limitation conditions (2-5). Numerous environmental factors can affect biofilm formation through different signal transduction pathways. For example, aminoglycoside antibiotics inhibit the inner membrane phosphodiesterase Arr and increase intracellular bis-(3=-5=)-cyclic dimeric GMP (c-di-GMP), which induce biofilm formation in Pseudomonas aeruginosa (6). ␤-Lactam antibiotics promote biofilm

The Regulation of Bacterial Biofilm Formation by cAMP-CRP: A Mini-Review

“…Biofilms have emergent properties that protect their inhabitants' survival in hostile environments, such as those in which the available nutrients have changed (Myszka and Czaczyk, 2009;Zhang et al, 2014;Liu et al, 2017Liu et al, , 2019Wang et al, 2017;Flemming and Wuertz, 2019). Several studies have shown that multiple bacteria form biofilms in response to changes in the types or concentrations of the available carbon sources (Myszka and Czaczyk, 2009;Zhang et al, 2014;Liu et al, 2017). The relationship between nutrient metabolism and biofilm formation is largely unknown.…”

“…Second messengers play an important role in various physiological processes in bacteria. The involvement of the second messenger c-di-GMP in biofilm formation is well established in multiple bacteria (Matsuyama et al, 2016;Liu et al, 2017), and regulatory effects of c-di-GMP on biofilm formation have been carefully reviewed (Christen et al, 2010;Dahlstrom and O'Toole, 2017;Krasteva and Sondermann, 2017). Compared to c-di-GMP, which participates in the whole biofilm cycle (from initial attachment to dispersion), the regulatory effects of cAMP-CRP on biofilm formation seem mostly ancillary.…”

“…The second messenger c-di-GMP is universally recognized as a "switch molecule" that controls the bacterial transition between a planktonic lifestyle and biofilm formation (Dahlstrom and O'Toole, 2017). High c-di-GMP levels induce biofilm formation, whereas low c-di-GMP levels induce planktonic growth (Valentini and Filloux, 2016;Liu et al, 2017). Another well-characterized second messenger cAMP, the cofactor of CRP, is primarily involved in carbon catabolite repression in bacteria.…”

Two-Component Signal Transduction Systems: A Major Strategy for Connecting Input Stimuli to Biofilm Formation

“…HK LrbS responds to the carbon source sodium lactate and triggers the transcriptional regulatory activity of LrbA by phosphorylation, which subsequently upregulates lrbR . Meanwhile, LrbS activates the phosphodiesterase of LrbR, which decreases intracellular c-di-GMP levels, thereby inhibiting biofilm formation (Figure 1C; Liu et al., 2017). The Lrb-three-component and Hno-multi-component signal transduction systems have much more complex regulatory modes in mediating biofilm formation.…”

The Cyclic AMP Receptor Protein, Crp, Is Required for the Decolorization of Acid Yellow 36 in Shewanella putrefaciens CN32

“…Then, the constructed plasmid was transformed into E. coli DH5α and introduced from E. coli DH5α into S. putrefaciens CN32 wild type (WT) using a helper plasmid, pRK2013, by triparental conjugation ( Figurski and Helinski, 1979 ). The conjugation experiment was performed according to a previous study ( Liu et al, 2017a ). Briefly, the donor and recipient strains were conjugated in a 1:1 ratio and spotted on an LB agar plate.…”