Downstream signaling mechanisms of phytochrome dimers
Cell Biology
Final Report Abstract
This project set out to investigate how phytochrome A (phyA) dimers transmit light signals, particularly focusing on the biologically active Pr–Pfr state. While the work was hampered by technical difficulties—such as repeated cloning failures and the lack of some expected molecular interactions— it nonetheless yielded several important findings relevant to plant photoreceptor biology. We optimized the custom multi-color SiMPull (single-molecule pull-down) setup to work with multiple colors. This device enabled us to image phytochrome dimerization and interaction at the singlemolecule level, preoviding information that complements traditional bulk assays. Although we were unable to consistently detect interactions between phyA and its signaling partners FHY1, PIF1, and NOT9B, likely due to weak binding affinities that limit the useability of the method, the data pointed to interesting new possibilities, including cotranslational dimerization and a potential preference for homodimer formation, both of which are entry points for further study. Then we extended our analysis to the type II phytochromes (phyB–E), and photobleaching and colocalization experiments revealed clear evidence of homodimerization for phyB, phyC, and phyE. We also confirmed that phyB and phyC form heterodimers. Unexpectedly, no heterodimers between phyB and phyE were detected in crude plant extracts, contradicting earlier reports and suggesting the need for more detailed follow-up work. In the scope of the PhD thesis but after the funding period of the DFG grant, we also analyzed physiological responses of phytochrome mutants. Measurements of properties like bolting time, leaf number, and hypocotyl length under different light conditions and temperatures uncovered distinct phenotypic differences. So far, no peer-reviewed publication has resulted from the project, but the plant physiologiy experiments will be included into a future manuscript, and the type II phytochrome dimerization experiments as well, once the remaining constructs are finished and measured.
Publications
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Downstream signaling of phytochrome dimers. PhD Thesis, University of Freiburg.
Lesch K. L.
