Phytochrome-Interacting Proteins.

Phytochromes are photoreceptors of plants, fungi, slime molds bacteria and heterokonts. These biliproteins sense red and far-red light and undergo light-induced changes between the two spectral forms, Pr and Pfr. Photoconversion triggered by light induces conformational changes in the bilin chromophore around the ring C-D-connecting methine bridge and is followed by conformational changes in the protein. For plant phytochromes, multiple phytochrome interacting proteins that mediate signal transduction, nuclear translocation or protein degradation have been identified. Few interacting proteins are known as bacterial or fungal phytochromes. Here, we describe how the interacting partners were identified, what is known about the different interactions and in which context of signal transduction these interactions are to be seen. The three-dimensional arrangement of these interacting partners is not known. Using an artificial intelligence system-based modeling software, a few predicted and modulated examples of interactions of bacterial phytochromes with their interaction partners are interpreted.

Dynamic interplays between three redox cofactors in a DNA photolyase revealed by spectral decomposition.

DNA repair catalyzed by photolyases is accomplished by a light-dependent electron transfer event from a fully reduced flavin adenine dinucleotide to a DNA lesion site. Prokaryotic DNA photolyase, PhrB, possesses a ribolumazine cofactor and a four-iron-four-sulfur cluster in addition to the catalytic flavin, but their functional roles are poorly understood. Here, we employ time-resolved absorption spectroscopy to probe light-induced responses in both solution and single crystals of PhrB. We jointly analyze a large collection of light-induced difference spectra from the wild-type and mutant PhrB obtained under different light and redox conditions. By applying singular value decomposition to 159 time series, we dissect light-induced spectral changes and examine the dynamic interplay between three cofactors. Our findings suggest that these cofactors form an interdependent redox network to coordinate light-induced redox responses. We propose that the ribolumazine cofactor serves as a photoprotective pigment under intense light or prolonged illumination, while the iron-sulfur cluster acts as a transient electron cache to maintain balance between two otherwise independent photoreactions of the flavin and ribolumazine.

Phytochromes in Agrobacterium fabrum.

The focus of this review is on the phytochromes Agp1 and Agp2 of Agrobacterium fabrum. These are involved in regulation of conjugation, gene transfer into plants, and other effects. Since crystal structures of both phytochromes are known, the phytochrome system of A. fabrum provides a tool for following the entire signal transduction cascade starting from light induced conformational changes to protein interaction and the triggering of DNA transfer processes.

Two aspartate residues close to the lesion binding site of Agrobacterium (6-4) photolyase are required for Mg2+ stimulation of DNA repair.

Prokaryotic (6-4) photolyases branch at the base of the evolution of cryptochromes and photolyases. Prototypical members contain an iron-sulphur cluster which was lost in the evolution of the other groups. In the Agrobacterium (6-4) photolyase PhrB, the repair of DNA lesions containing UV-induced (6-4) pyrimidine dimers is stimulated by Mg2+ . We propose that Mg2+ is required for efficient lesion binding and for charge stabilization after electron transfer from the FADH- chromophore to the DNA lesion. Furthermore, two highly conserved Asp residues close to the DNA-binding site are essential for the effect of Mg2+ . Simulations show that two Mg2+ bind to the region around these residues. On the other hand, DNA repair by eukaryotic (6-4) photolyases is not increased by Mg2+ . In these photolyases, structurally overlapping regions contain no Asp but positively charged Lys or Arg. During the evolution of photolyases, the role of Mg2+ in charge stabilization and enhancement of DNA binding was therefore taken over by a postiviely charged amino acid. Besides PhrB, another prokaryotic (6-4) photolyase from the marine cyanobacterium Prochlorococcus marinus, PromaPL, which contains no iron-sulphur cluster, was also investigated. This photolyase is stimulated by Mg2+ as well. The evolutionary loss of the iron-sulphur cluster due to limiting iron concentrations can occur in a marine environment as a result of iron deprivation. However, the evolutionary replacement of Mg2+ by a positively charged amino acid is unlikely to occur in a marine environment because the concentration of divalent cations in seawater is always sufficient. We therefore assume that this transition could have occurred in a freshwater environment.

Evidence for weak interaction between phytochromes Agp1 and Agp2 from Agrobacterium fabrum.

During bacterial conjugation, plasmid DNA is transferred from cell to cell. In Agrobacterium fabrum, conjugation is regulated by the phytochrome photoreceptors Agp1 and Agp2. Both contribute equally to this regulation. Agp1 and Agp2 are histidine kinases, but, for Agp2, we found no autophosphorylation activity. A clear autophosphorylation signal, however, was obtained with mutants in which the phosphoaccepting Asp of the C-terminal response regulator domain is replaced. Thus, the Agp2 histidine kinase differs from the classical transphosphorylation pattern. We performed size exclusion, photoconversion, dark reversion, autophosphorylation, chromophore assembly kinetics and fluorescence resonance energy transfer measurements on mixed Agp1/Agp2 samples. These assays pointed to an interaction between both proteins. This could partially explain the coaction of both phytochromes in the cell.