Fine-tuning of chemotactic response in E. coli determined by high-throughput capillary assay

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Wiki Entry by Daniel Rubin, AP225, 9/24/2012

General Information

Authors: Heungwon Park, Calin C. Gouet, Thierry Emonet, Phillipe Cluzel

Publication: Park, et al. Fine-Tuning of Chemotactic Response in E. coli Determined by High-Throughput Capillary Assay, (2011) Curr. Microbiol. 62:764-769

Key Words: Random walk, Chemotaxis, E. coli, High-throughput

Summary

Chemotaxis is the primary means by which bacteria know to move toward nutrients and away from harm. The process is typically thought of as a biased random walk composed of random tumbling steps followed by extended periods of movement in a single direction. While the tumbling remains random, regardless of the chemotactic gradient, the periods of straight movement are extended when moving towards an attractant, or away from a repellant. With these simple rules, E. coli are able to navigate a wide variety of conditions.

At a molecular level, this process is a well-choreographed assemblage of many different proteins that interact and dissociate from one-another in different ways. Two of the major proteins responsible for E. coli chemotactic behavior are CheR and CheB. These proteins work together to methylate/demethylate the receptor kinases that are activated by the chemotactic factor outside of the cellular membrane. While the process of chemotaxis has been studied intensively, it is almost universally studied in a bacterial 'swarm' environment. While this has provided insight over the years, it is not a perfect system- other factors are at play. Within a swarm, population growth, and depletion of nutrients can affect the chemotactic behavior of the E. coli, convoluting a more precise understanding of the many proteins necessary for chemotaxis to occur properly.

Park et al. have devised a high-throuput capillary system that allows their research team to deconvolute the swarming behavior of E. coli when following a chemoattractant. Specifically, they use this system to study the robustness of the chemotactic response with respect to the intercellular concentrations of CheR and CheB. Recombinant CheR and CheB (fluorescent fusions) were added by plasmid to the E. coli under inducible promoters, allowing researchers to assay the attraction to L-aspartate with varied concentrations of CheR and CheB present within the cell. To get a sense for the apparatus that was designed, see figure 1:

Figure 1:The device shown was used as a means to test large numbers of E. coli, independently, while ignoring swarming affects. In the top 24 wells, L-aspartate was added within the capillary. In the lower 24 wells, only mobility media was present.

As one might expect, they conclude that while chemotactic behavior is quite robust to altered concentrations of CheR and CheB when bacteria are swarming, it is much more sensitive in a non-swarming scenario. In fact, the chemotactic response is clearly maximized in the case where the ratio of CheR to CheB is equal to that of the wild-type E. coli (Go Evolution!). As figure 2: shows, one can graphically depict this behavior by plotting chemotactic response and efficiency versus the ratio of CheR and CheB to wild type amounts. When each is correlated 1-to-1 with it's wild-type analog, the response is largest.

Figure 2: As the relative concentration of CheR and CheB are adjusted, the chemotactic response and efficiency are altered.

Discussion

While this experiment is quite simple (in theory), it illustrates a key point for those of use studying biology- Simplify your system and utilize reasonable assumptions. Oftentimes, the complexity of biological systems can be overwhelming, so researchers often overlook confounding factors. In this case, it was not only clear to the authors that swarming disrupted the expected data from chemotactic assays but that there was a relatively simple way to engineer around this problem.

Reference

Park, et al. Fine-Tuning of Chemotactic Response in E. coli Determined by High-Throughput Capillary Assay, (2011) Curr. Microbiol. 62:764-769