Dynamics of a microbe's memory revealed by light
Combining optical measurements with theoretical modeling, researchers at AMOLF, IBM and Harvard University have succeeded in characterizing the molecular memory that controls the locomotion of bacteria – the simplest forms of cellular life. The results are published in the prestigious journal Molecular Systems Biology on 22 June.
When searching for better environments, microscopic living things such as bacteria operate very differently from macroscopic organisms such as humans. They explore their local environment by swimming along fairly straight trajectories (called runs) that are occasionally interrupted by brief periods of chaotic reorientation (called tumbles). To achieve movement towards favorable directions through this 'random walk' behavior, they use a sensory network of molecules that constantly monitors the cell’s locale during the movement. It also makes comparisons of the current environment with those experienced in the past. In this way, spatial features of the environment are experienced over time, and the organism must 'remember' conditions that were experienced in the past, and compare them with what they sense in the present. Information about the past is stored in the form of chemical modifications on receptor proteins involved in the sensing. The newly published study – a collaboration between AMOLF group leader Tom Shimizu
, IBM theoretical physicist Yuhai Tu, and Harvard professor Howard C. Berg – has revealed the dynamics of this 'short-term memory' of the bacterium Escherichia coli, by measuring molecular interactions as they happen within live cells.
The experiments exploit a physical process known as Förster resonance energy transfer (FRET), to quantify the strength of interaction between protein molecules in the signaling pathway. The key idea is that when fluorescently labeled molecules come into close physical proximity, they can exchange energy between one another. And this transfer of energy results in measurable changes in the emitted light. By careful consideration of which molecules to label and how, FRET experiments can be designed to study a wide variety of molecular processes, from the distance between two labeled sites on a single molecule, to the number of interacting pairs in a chemically reacting ensemble of molecules. In the present study, the authors measured FRET between a central protein of the network, known as a 'response regulator', and another enzyme that degrades its activity.
The protein molecules are labeled genetically, by fusing the DNA sequence of their genes to those of green fluorescent protein (GFP) – a naturally fluorescent protein isolated from jellyfish. GFP is an ideal label for FRET in live cells, but it can also impair the function of the labeled molecule. Protein fluorophores such as GFP are quite large and bulky, so using them as labels can easily disrupt the properties of the original protein. Thus, obtaining GFP-labeled proteins that retain their original function already requires some degree of luck. But getting a pair suitable for FRET requires further serendipity, as the physical mechanism requires the fused GFP’s to come within a very small (<10nm) distance, known as the Förster radius, of one another.
In the current study, careful analysis of a theoretical model of the system allowed the authors to use a single FRET pair to measure input-output relationships of different functional parts, or 'modules', of the pathway. This was made possible by modulating the concentration of stimulus molecules over time to produce various input waveforms, such as steps, ramps or sinusoids, much like those used by engineers to study electrical circuits.
The results shed light both on the molecular mechanisms at work inside these living cells, as well as the functional design of the sensory network. In particular, the authors characterized the way in which signals are amplified provided by the 'receptor module', as well as the speed of chemical reactions within the 'adaptation module'. Together, these modules determine the bacterium’s sensitivity to chemical gradients it experiences as it wanders about in search for better environments.
While cells – the basic unit of life – are microscopic in dimensions, they demonstrate a myriad of seemingly sentient behavior. The present study demonstrates how experiment and theory can be combined to reveal the physical basis of such behaviors at the molecular level.
Reference
A modular gradient-sensing network for chemotaxis in Escherichia coli revealed by responses to time-varying stimuli, Thomas S. Shimizu, Yuhai Tu, and Howard C. Berg, Molecular Systems Biology 6:382 (2010).
Information
For more information, please contact:
Dr. Tom Shimizu, FOM-instituut AMOLF, telefoon (020) 754 71 00