The EAL‐like protein STM1697 regulates virulence phenotypes, motility and biofilm formation in Salmonella typhimurium

Abstract

SummaryThe ubiquitous second messenger c-di-GMP regulates the switching of bacterial lifestyles from motility to sessility and acute to chronic virulence to adjust bacterial fitness to altered environmental conditions. Conventionally, EAL proteins being c-di-GMP phosphodiesterases promote motility and acute virulence phenotypes such as invasion into epithelial cells and inhibit biofilm formation. We report here that in contradiction, the EAL-like protein STM1697 of Salmonella typhimurium suppresses motility, invasion into HT-29 epithelial cell line and secretion of the type three secretion system 1 effector protein SipA, whereas it promotes rdar biofilm formation and CsgD expression. STM1697 can, however, functionally replace the EAL-like protein STM1344 and vice versa, whereby both proteins neither degrade nor bind c-di-GMP. Like STM1344, STM1697 suppresses the transcription of class 2 and class 3 flagella regulon genes by binding to FlhD, a component of the master regulator of the flagella regulon FlhD 4C2 and act additively under numerous conditions. Interestingly, the interaction interface of STM1697 with FlhD 2 is distinct from its paralogue STM1344. We predict that the stand alone EAL domain proteins STM1697 and STM1344 belong to a subclass of EAL domain proteins in S. typhimurium, which are all involved in motility, biofilm and virulence regulation through interaction with proteins that regulate flagella function.

Bistable Expression of CsgD in Salmonella enterica Serovar Typhimurium Connects Virulence to Persistence

“…The importance of SPI-1 in this process was confirmed, since planktonic cells and multicellular aggregates from a ⌬SPI-1 mutant strain had nearly equal colonization efficiencies. Furthermore, the reduction in virulence that we observed for the ⌬csgD strain, together with recent connections between c-di-GMP, virulence (28), and motility (52), suggest that the master biofilm regulator, CsgD, can modulate the virulence capacity of S. Typhimurium.…”

“…One possible explanation for this correlation is that the rdar morphotype is an adap-tation necessary for life outside the host. However, there have been recent links between the rdar morphotype, immune stimulation, and invasion (28,29), making it difficult to fully understand this physiology. Although they are informative, a potential problem with strain-to-strain comparisons is that they could miss fluctuations that exist within individual populations of cells.…”

Stand-Alone EAL Domain Proteins Form a Distinct Subclass of EAL Proteins Involved in Regulation of Cell Motility and Biofilm Formation in Enterobacteria

“…Recent studies of the EAL domain sequences and structures provided significant insights into the mechanism of its PDE activity (1,6,7). These studies characterized a number of EAL domains that functioned as c-di-GMP-specific PDEs and also some inactivated domains that were devoid of enzymatic activity (8)(9)(10). Based on the conservation of the active-site motifs and the dimerization loop (11), EAL domains have been categorized into three main classes: (i) bona fide c-di-GMP PDEs, (ii) EAL domains with a degenerate loop 6 that might or might not have PDE activity, and (iii) EAL domains that lack the key catalytic residues and have no enzymatic activity (11,12).…”

“…Based on the conservation of the active-site motifs and the dimerization loop (11), EAL domains have been categorized into three main classes: (i) bona fide c-di-GMP PDEs, (ii) EAL domains with a degenerate loop 6 that might or might not have PDE activity, and (iii) EAL domains that lack the key catalytic residues and have no enzymatic activity (11,12). The latter class could be further subdivided into those EAL domains that have retained the ability to bind c-di-GMP and serve as c-di-GMP receptors and those domains that do not even bind c-di-GMP (9,10,13,14).…”

“…These proteins display no detectable PDE activity, do not readily form dimers, and do not bind c-di-GMP (9, 10), falling into class 3b of the above-mentioned classification (12). Nevertheless, these proteins have been shown to regulate motility and cell adherence and affect bacterial cytotoxicity towards macrophages (9,10,31,32). This regulation involves a direct interaction with the FlhD component of FlhD 4 C 2 , the master regulator of flagellar gene expression, to prevent the interaction of FlhD 4 C 2 with DNA and to stimulate proteolysis by the ClpXP protease (10,33,34).…”