Function of and signaling by blue light photoreceptors in Rhodobacter sphaeroides
Zusammenfassung der Projektergebnisse
While a previous project had investigated the role of the AppA photoreceptor on photosynthesis gene regulation, project Kl563/22 was designed to unravel the function of the LOV and CryB photoreceptors in Rhodobacter sphaeroides. Since only one graduate student could work on the project only a small part of the project focused on the LOV protein. Our hypothesis that LOV may control cell attachment by acting on capsule or biofilm formation was not confirmed. A transcriptome study revealed that LOV affects many genes (under different conditions) with a role in amino acid and carbon metabolisms and transport and mainly acts as a repressor. Mechanisms of regulation by LOV could not be investigated further. The main part of our work focused on the CryB protein with a known role in regulation of photosynthesis genes and in in vivo photoreactivation. Transcriptome analyses revealed an effect of CryB (under different growth conditions) on several genes and an important role in oxidative stress defense. Regarding the underlying mechanisms we focused on regulation of photosynthesis genes and showed that a light- and redox-dependent interaction of CryB to AppA affects binding of AppA to the transcriptional repressor PpsR and consequently expression of photosynthesis genes. Thus, two photoreceptors interact to balance the regulation of photosynthesis genes in R. sphaeroides. A collaboration with Lars-Oliver Essen allowed to solve the crystal structure and to identify two additional cofactors in addition to the already identified FAD in the active site. A new type of antenna cofactor (lumazine) and an Fe-S cluster are present in CryB from R. sphaeroides and predicted for proteins of the cryptochrome/photolyase family from other alphaproteo- and cyanobacteria. In addition the amino acids W338-W386-Y387 were shown to function as electron chain to FAD. Based on the structure site-directed amino acid exchanges in CryB were performed to address the function of the cofactors and of the electron transport chain. In cooperation with the Essen group and R. Pokorny (both University of Marburg) we could prove a weak (6-4) photolyase activity for CryB. The Fe-S cluster proved essential for the integrity of the protein. The FAD cofactor in the active site was essential for in vitro repair, it was not essential for in vivo photoreactivation as long as the lumazine cofactor was present. Since the construction of many mutants and the analyses of photolyase function took a lot of time and efforts, the different CryB variants could not yet be tested for their function in gene regulation.
Projektbezogene Publikationen (Auswahl)
- (2012) CryB from Rhodobacter sphaeroides: A unique class of cryptochromes with novel cofactors. EMBO rep. 13: 223-229
Geisselbrecht, Y., Frühwirth, S., Schröder, C., Pierik, A.J., Klug, G., Essen, L.-O.
(Siehe online unter https://doi.org/10.1038/embor.2012.2) - (2012) Effects of the cryptochrome CryB from Rhodobacter sphaeroides on global gene expression the dark or blue light or in presence of singlet oxygen. PLoS ONE 7: e33791
Frühwirth, S., Teich, Klug, G.
(Siehe online unter https://doi.org/10.1371/journal.pone.0033791) - (2012) Interaction of two photoreceptors, AppA and CryB, in the regulation of photosynthesis genes in Rhodobacter sphaeroides. Nucleic Acids Res. 40: 5901-5909
Metz, S., Haberzettl, K., Fruehwirth, S.W., Teich, K., Hasewinkel, C., Klug, G.
- (2012) Role of a short LOV protein in blue light- and singlet oxygendependent gene regulation in Rhodobacter sphaeroides. Microbiology 158: 353-352
Metz, S., Jäger, A., Klug, G.
- (2014) Beyond catalysis: Vitamine B12 as a cofactor in gene regulation. Mol. Microbiol. 91: 635-640
Klug, G.
(Siehe online unter https://doi.org/10.1111/mmi.12490) - (2016) New insights into the bifunctional CryB of Rhodobacter sphaeroides, a bacterial cryptochrome with (6-4) photolyase activity. FEBS J. 283: 4291-4309
von Zadow, A., Ignatz, E., Pokorny, R., Essen, L.-O., Klug G.
(Siehe online unter https://doi.org/10.1111/febs.13924)
