Abstract
The spatial location of proteins in living cells can be critical for their function. For example, the E. coli chemotaxis machinery is localized to the cell poles. Here we describe the polar localization of the serine chemoreceptor Tsr using a strain synthesizing a fluorescent Tsr-Venus fusion at a low level from a single-copy chromosomal construct. Using photobleaching and imaging during recovery by new synthesis, we observed distinct asymmetry between a bright (old) pole and a dim (new) pole. The old pole was shown to be a more stable cluster and to recover after photobleaching faster, which is consistent with the hypothesis that newly synthesized Tsr proteins are inserted directly at or near the old pole. The new pole was shown to be a less stable cluster and to exchange proteins freely with highly mobile Tsr-Venus proteins diffusing in the membrane. We propose that the new pole arises from molecules escaping from the old pole and diffusing to the new pole where a more stable cluster forms over time. Our localization imaging data support a model in which a nascent new pole forms prior to stable cluster formation.
Highlights
About 4% of Escherichia coli membrane proteins are localized at the cell poles, which is critical for their function [1]
We recently found that the majority of newly synthesized Tsr accumulate at the old poles of E. coli and diffuse at different rates at the polar and cylindrical surface regions [22]
We found through direct single molecule counting, a total of 2356 new fluorescent spots among 1956 cells were observed over a 5 minute time scale, with 630 cells displaying more than one event, implying an upper bound for the protein production rate of 0.24 proteins per cell per min
Summary
About 4% of Escherichia coli membrane proteins are localized at the cell poles, which is critical for their function [1]. The polar localization of chemoreceptors, like the serine chemoreceptor Tsr, is required sensing environmental signals [1]. Tsr is a highly abundant transmembrane methyl-accepting chemotaxis protein (MCP) and is stably localized at the poles in variably sized clusters [2], which are necessary for collaborative signalling [3]. Tsr is one of five MCPs that form heterotrimeric membrane complexes at the poles [4]. The polar distribution of Tsr is asymmetric and changes during the cell cycle.
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