Institut für Biochemie, Genetik und Mikrobiologie, Universität Regensburg, D-93040 Regensburg, Germany1
Tel: +49 941 9433162. Fax: +49 941 9433163. e-mail: rudy.schmitt{at}biologie.uni-regensburg.de
Keywords: -proteobacteria, sensory signal transduction, multiple chemotaxis genes, retro-phosphorelay, chemokinesis
a To Professor Wolfram Heumann on his 87th birthday.
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Overview |
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The focus of this article will be on features that distinguish the chemosensory systems of E. coli and Sinorhizobium meliloti. The soil bacterium S. meliloti is a member of the -subclass of Proteobacteria (Olsen et al., 1994
), which contains such diverse species as Acetobacter, Agrobacterium, Caulobacter, Methylomonas and Rhodobacter. These differ from enterobacteria (
-subclass) by their wide range of natural habitats and by their great metabolic flexibility, reflecting a separate evolution of more than 500 million years (Ochman & Wilson, 1987
). By colonizing the lower animal gut, E. coli has adopted a specialized lifestyle, whereas the nitrogen-fixing S. meliloti leads a versatile life, either freely in the soil, or in the rhizosphere of host plants, or ultimately in root nodules, to convert atmospheric nitrogen to biologically usable ammonium. Differences between the specialized enterobacterial and the versatile rhizobial lifestyles are reflected in their recently sequenced genomes: E. coli K-12 contains a single circular chromosome with just 4300 protein-coding genes (Blattner et al., 1997
), while S. meliloti features three circular replicons the actual chromosome and two megaplasmids, pSymA and pSymB with over 6200 protein-coding genes (Galibert et al., 2001
). A large and variable genome is considered essential for growth and survival under the complex environmental conditions prevailing in the soil, the rhizosphere and in plant roots. As for many years the coli-centric view has considered the presence of a single circular chromosome as universally valid, studies of S. meliloti and other
-proteobacteria have revealed an abundance of circular and even linear chromosomes that was previously unseen (Jumas-Bilak et al., 1998
). Similarly, analyses of the chemosensory system of S. meliloti and its relatives (Armitage & Schmitt, 1997
) have revealed departures from the enterobacterial paradigm that expand and newly define previous concepts. This will be illustrated in this review by two novel features that distinguish the S. meliloti chemosensory system from the established enterobacterial scheme.
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Signalling pathways |
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Retro-phosphorylation in the sensory transduction chain as a mechanism for efficient dephosphorylation of the response regulator CheY2-P, and hence for rapid adaptation to new stimuli, was initially considered a peculiarity of S. meliloti (Sourjik & Schmitt, 1998 ). However, an inspection of well-analysed
-proteobacteria and of some recently published genome sequences of other non-enterics revealed the presence of CheY1 and CheY2 paralogues concurrent with the absence of CheZ in many of these bacterial species. We take this as strong suggestive evidence for functional retro-phosphorylation pathways operating in
-proteobacteria, such as Agrobacterium, Caulobacter, Rhodobacter and the rhizobia (Armitage & Schmitt, 1997
), as well as in Bacillus subtilis (Rosario et al., 1994
), Borrelia burgdorferi (Fraser et al., 1997
), Campylobacter jejuni (Parkhill et al., 2000
), Helicobacter pylori (Tomb et al., 1997
), in the cyanobacterium Synechocystis (Kaneko et al., 1996
) and even in the hyperthermophilic Thermotoga maritima (Nelson et al., 1999
) representing the deepest known branch of the bacterial phylogenetic tree (Stetter, 1995
). This suggests that the retro-phosphorelay may have evolved as an original mechanism for the deactivation of sensory response regulators and that the enterobacterial CheYCheZ pathway may have been developed more recently as an adaptation to modern intestinal environments.
It should be noted that whole-genome sequences of various -proteobacteria and of other non-enterics revealed an additional complexity in containing two (or more) operons with multiple copies of the chemosensory (che) genes. Is there a biological concept behind these duplicate pathways? In the photosynthetic Rhodobacter sphaeroides, one of two che operons is dominant directing all known photo- and chemotactic responses, whereas mutants in the second che operon have virtually no effect on bacterial behaviour under laboratory conditions (Hamblin et al., 1997
). Similarly, in S. meliloti, the chromosomal che operon dominantly controls the response to all tested attractants (Greck et al., 1995
; Sourjik & Schmitt, 1996
). A second set of che genes revealed by the complete S. meliloti genome sequence (Galibert et al., 2001
) is located on the pSymA replicon and linked to genes that control type IV pilus synthesis. Together, the taxis- and pilus-coding genes may have a role in gliding (or twitching) motility (Kearns et al., 2001
) appropriate for colonizing the host plant. While it is conceivable that multiple che operons in one cell may interact in a synergistic fashion, the two reported cases rather suggest that these operons specify distinctive sensory pathways operating under diverse and rather specific environmental conditions. It should also be mentioned that multiple CheY paralogues observed in various non-enterics do not preclude, but on the contrary favour, the existence of a functional retro-phosphorelay pathway, as long as CheZ phosphatase is lacking.
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Swimming patterns and flagellar rotary control |
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The flagellar filaments and their mode of rotation differ greatly between enteric bacteria and S. meliloti and its relatives. In E. coli, the six to eight flexible left-handed helical flagella mostly rotate counterclockwise (ccw) to form a bundle that propels the cell forward. A new direction is taken up when some flagella switch to clockwise (cw) rotation, forcing the handedness and helical amplitude to change, which drives the bundle apart and causes the cell to tumble. Unlike E. coli, the soil bacterium S. meliloti is driven by rather rigid, right-handed helical filaments, which facilitate efficient swimming in a viscous environment (Götz et al., 1982 ). Whereas the plain enterobacterial flagellar filaments are assembled from monomeric flagellin subunits, the complex rhizobial filaments consist of four related flagellin subunits that are assembled as functional heterodimers (Scharf et al., 2001
). The complex flagellar fine structure is dominated by three helical ribbons that lock the filament in right-handedness, thus blocking a switch of hand (Cohen-Krausz & Trachtenberg, 1998
). Also, the motor of S. meliloti exhibits unidirectional cw rotation and never switches to ccw rotation. Hence, a swimming S. meliloti cell is driven by the unidirectional cw rotation of its right-handed helical filaments. Unlike in enterobacteria, but similar to R. sphaeroides (Packer et al., 1997
), the flagellar motors can increase and decrease rotary speed (Götz & Schmitt, 1987
; Platzer et al., 1997
). Therefore, these bacteria respond to the addition of chemoattractants by a sustained increase in free-swimming speed, a phenomenon called chemokinesis (Sourjik & Schmitt, 1996
). Full-speed cw rotation causes the four to six right-handed filaments of S. meliloti to form a bundle that pushes the cell forward. A new direction is assumed when the rotary speeds of individual flagella decline at different rates, forcing their helical amplitudes to change asynchronously, which drives the bundle apart and causes the cell to turn or tumble. Hence, to change the direction of a swimming cell, E. coli flagellar rotors switch into reverse gear, whereas S. meliloti by independently slowing individual propellers follows the principle of a caterpillar drive.
In S. meliloti and the related Rhizobium lupini H13-3 (Scharf et al., 2001 ), rotary speed variation of the flagellar motor has a molecular corollary in two new motility proteins, MotC and MotD (Platzer et al., 1997
), present in addition to the ubiquitous MotA/MotB transmembrane proteins that form the energizing proton channels (Fig. 1
). MotC is a periplasmic protein interacting with and presumably stabilizing the periplasmic domain of MotB. The cytoplasmic MotD is essential for motility and, when overexpressed, enhances swimming and rotary speed (Platzer et al., 1997
). In vitro, MotD binds to FliM, the very component of the cytoplasmic portion of the rotor (C-ring) to which CheY2-P also binds to effect a slow-down of rotary speed (E. Eggenhofer, B. Scharf & R. Schmitt, unpublished). MotD and CheY2-P may compete in an antagonistic fashion for binding to the
34 copies of FliM (Thomas et al., 1999
) that form the cytoplasmic surface of the rotor. In such a model, the actual ratio of MotD:CheY2-P bound to the C-ring would thus determine the rotary speed at each moment. Such a mechanism can be tested by monitoring the effects of an inducible resurrection of CheY2-P (or an activated CheY2 derivative) on flagellar rotary speed reduction, as has been described by Scharf et al. (1998)
.
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Conclusions |
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ACKNOWLEDGEMENTS |
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