Functional Characterization of c-di-GMP Signaling-Related Genes in the Probiotic Lactobacillus acidophilus.

Jiahui He,Wenjuan Yan,Fang Wang,Wenhao Ruan,Jieli Sun

doi:10.3389/fmicb.2018.01935

Abstract

The bacterial second messenger cyclic diguanylate monophosphate (c-di-GMP) regulates a series of cellular functions, including biofilm formation, motility, virulence, and other processes. In this study, we confirmed the presence of several c-di-GMP related genes and evaluated their activities and functions in Lactobacillus species. Bioinformatic and biochemical analyses revealed that Lactobacillus acidophilus La-14 have an active c-di-GMP phosphodiesterase (PdeA) that may act in the metabolic cycle of c-di-GMP. A GGDEF protein (DgcA) induced two c-di-GMP-dependent phenotypes (low motility and high production of curli fimbriae) in Escherichia coli by heterologously expressed in vivo but showed no diguanylate cyclases activity in vitro while in the expression without the N-terminal transmembrane domain. The degenerated EAL-domain protein (PdeB), encoded by the last gene in the gts operon, serve as a c-di-GMP receptor which may be associated with exopolysaccharide (EPS) synthesis in L. acidophilus. Heterologously expressed GtsA and GtsB, encoded by the gts operon, stimulated EPS and biofilm formation in E. coli BL21. Constitutive expression in L. acidophilus revealed that a high concentration of intracellular DgcA levels increased EPS production in L. acidophilus and enhanced the co-aggregation ability with E. coli MG1655, which may be beneficial to the probiotic properties of Lactobacillus species. Our study imply that the c-di-GMP metabolism-related genes, in L. acidophilus, work jointly to regulate its functions in EPS formation and co-aggregation.

Highlights

Cyclic diguanylate monophosphate (c-di-GMP), formed by the condensation of two GTP molecules, is a secondary messenger that is widely distributed in bacteria and is involved in the regulation of multiple bacterial physiological functions (Hengge, 2009)
The L. acidophilus La-14 genome (NCBI reference sequence: NC_021181.2) contains a gene (LA14_RS07000, dgcA) encoding the GGDEF domain and two genes (LA14_RS07005, pdeA; LA14_RS07010, pdeB) encoding the EAL domain (Figure 1A); these genes may be involved in the metabolic cycle of c-di-GMP
Such adaptations are regulated in various bacteria by proteins with GGDEF and EAL domains, which involve the second messenger c-di-GMP

Summary

Introduction

Cyclic diguanylate monophosphate (c-di-GMP), formed by the condensation of two GTP molecules, is a secondary messenger that is widely distributed in bacteria and is involved in the regulation of multiple bacterial physiological functions (Hengge, 2009). The existence of a c-di-GMP signaling pathway has been confirmed in many Grampositive bacteria, such as Streptomyces coelicolor (den Hengst et al, 2010), Clostridium difficile (Purcell et al, 2012), Bacillus subtilis (Gao et al, 2013), and Listeria monocytogenes (Chen et al, 2014). In these species, c-di-GMP signaling primarily regulates flagellum synthesis, production of adhesion factor in response to surface contact, and production of extracellular polymeric substances (Purcell and Tamayo, 2016; Bedrunka and Graumann, 2017a)

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