Roles of the second messenger cyclic di-GMP in environmental adaptation of Sinorhizobium meliloti

Bacteria have evolved various systems for the integration of environmental signals to rapidly coordinate cellular pathways and adapt to changes in their environment. In the quickly advancing field of nucleotide-based second messengers, cyclic dimeric guanosine monophosphate (c-di-GMP) has emerged as...

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Bibliographic Details
Main Author: Schäper, Simon
Contributors: Becker, Anke (Prof. Dr.) (Thesis advisor)
Format: Dissertation
Language:English
Published: Philipps-Universität Marburg 2017
Biologie
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Summary:Bacteria have evolved various systems for the integration of environmental signals to rapidly coordinate cellular pathways and adapt to changes in their environment. In the quickly advancing field of nucleotide-based second messengers, cyclic dimeric guanosine monophosphate (c-di-GMP) has emerged as a key regulatory player whose underlying signaling networks control major adaptational and lifestyle changes. Enzymes that catalyze synthesis and degradation of c-di-GMP, named diguanylate cylases (DGCs) and phosphodiesterases (PDEs), respectively, are near-ubiquitous in the bacterial kingdom. Despite the numerous studies aiming to better understand the role of c-di-GMP in bacteria, knowledge on integration of c-di-GMP networks into other regulatory networks, the molecular inventory of c-di-GMP receptors and molecular mechanisms underlying c-di-GMP-dependent regulation is limited. This study investigated roles of c-di-GMP in environmental adaptation of soil-dwelling Sinorhizobium meliloti, a rod-shaped alphaproteobacterium from the order Rhizobiales that exists either in free-living states or in symbiosis with leguminous plant hosts. The S. meliloti genome encodes 22 proteins putatively involved in synthesis, degradation and binding of c-di-GMP. Single mutations in 21 of these genes did not cause evident changes in surface attachment, swimming motility or exopolysaccharide (EPS) production. Moreover, screening the different phenotypes of S. meliloti c-di-GMP0 mutants revealed no defects in cell viability and symbiotic potency. In contrast, artificially increasing c-di-GMP levels by overproduction of several DGCs promoted production of extracellular matrix components and surface attachment, whereas swimming motility and extracellular accumulation of N-Acyl-homoserine lactones (AHLs) was reduced. The identification of genetic determinants responsible for observed phenotypic changes at elevated c-di-GMP levels proved c-di-GMP-dependent regulation at both transcriptional and post-translational levels. The SMc01790-SMc01796 locus, homologous to the Agrobacterium tumefaciens uppABCDEF cluster governing biosynthesis of a unipolar polysaccharide (UPP), was required for c-di-GMP-stimulated surface attachment, while the stand-alone PilZ domain protein SMc00507 (renamed McrA) acted as c-di-GMP receptor protein involved in regulation of swimming motility. Transcriptome profiling of S. meliloti at elevated c-di-GMP levels revealed upregulation of the uxs1-SMb20463 gene cluster governing biosynthesis of an extracellular polysaccharide (referred to as CUP). Resulting from this finding, AraC-like transcriptional activator SMb20457 (renamed CuxR) was shown to bind c-di-GMP by a mechanism similar to that of PilZ domains, which provided an example of convergent evolution in two distinct protein families. This study demonstrates that the c-di-GMP network in S. meliloti is integrated into other cellular systems, particularly the well-characterized regulatory network for opposing control of EPS biosynthesis and motility. For instance, CuxR-mediated activation of CUP production was counteracted by the global repressor MucR, while both MucR and the AHL-sensitive master regulator ExpR reduced UPP-mediated surface attachment at elevated c-di-GMP levels. Moreover, a new cellular function was assigned to the essential PDE SMc00074 (renamed GdcP), which is linked to cell envelope biogenesis in alpha-rhizobial species. Overall, c-di-GMP-dependent regulation of multiple cellular functions indicated that high c-di-GMP levels favor a sedentary lifestyle of free-living S. meliloti. The switch of single motile bacteria from a planktonic state to a structured community of cells might contribute to environmental adaptation and long-term survival of S. meliloti in its natural soil habitat.
Physical Description:196 Pages
DOI:https://doi.org/10.17192/z2017.0677