Superhochauflösendes Mikroskop STORM/PALM
Final Report Abstract
Bacterial transformation is the import and inheritable integration of external DNA. As such, it is believed to be a major evolutionary force. A key step is the import of DNA through the outer membrane. We have characterized the spatio-temporal dynamics of DNA during import and residence in the periplasm of the Gram-negative pathogen Neisseria gonorrhoeae. We found that the periplasm can serve as a reservoir for imported DNA that can fill within five minutes by importing DNA from the environment. The amount of imported DNA roughly corresponds to the size of a phage genome. The high sensitivity of the STORM microscope was used to show that periplasmic DNA-binding protein ComE is homogeneously distributed in the periplasm in the absence of extracellular DNA. It relocates rapidly to imported DNA when external DNA is added to competent gonococci. As ComE governs the carrying capacity of the periplasm, we propose that it might condense DNA thus linking DNA uptake to its compaction. Our findings strongly support the idea that the periplasm masses DNA independently of transport through the inner membrane. DNA uptake through the outer membrane is driven by a powerful molecular machine. Using the STORM microscope, we showed that the machine transports single- and double stranded DNA at comparable rates, provided that the species-specific DNA uptake sequence is double-stranded. Moreover, we found that single-stranded DNA taken up into the periplasm is rapidly degraded by the thermonuclease Nuc. We conclude that the secondary structure of transforming DNA is important for the recognition of self-DNA but not for the transport process through the outer membrane.
Publications
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(2014) Concerted spatio-temporal dynamics of imported DNA and ComE DNA uptake protein during gonococcal transformation, PLoS Pathog., 10(4), e1004043
Gangel, H., Hepp, C., Müller, S., Oldewurtel, E.R., Aas, F.E., Koomey, M., Maier, B.
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(2016) Single-stranded DNA uptake during gonococcal transformation, J. Bacteriol., 198(18), 2515-23
Hepp, C., Gangel, H., Henseler, K., Günther, N., Maier, B.
