Two Native Pools of Phytochrome A in Monocots: Evidence from Fluorescence Investigations of Phytochrome Mutants of Rice

Abstract

Fluorescence investigations of phytochrome (phy) in rice (Oryza sativa L. cv. Nipponbare) mutants deficient in phyA, phyB and phyA plus phyB were performed. Total content of the pigment (P(tot)) and its spectroscopic and photochemical characteristics were determined in different parts of the dark-grown and far-red light (FR)-grown coleoptiles. Spectroscopically, phyA in the phyB mutant was identical to phyA in the wild-type (WT) and the extent of the conversion from Pr to lumi-R at 85 K was the same for phyA in both lines and varied similarly, depending on the part of the coleoptile used. The latter finding proved that phyA in rice is heterogeneous and comprises two phyA populations, phyA' and phyA". Functional properties of phyA were also determined. In the dark the phyB mutant had a higher content of phyA, inactive protochlorophyllide (Pchlide633) and active protochlorophyllide (Pchlide655) than WT and its coleoptile was longer, indicating that phyB may affect the development of WT seedlings in the dark. Constant FR drastically reduced the content of phyA, Pchlide633 and Pchlide655 and brought about coleoptile shortening and appearance of the first leaf, whereas pulsed FR of equal fluence was less effective. This suggested that the reactions were primarily of the high irradiance responses type, which are likely to be mediated by phyA'. The effects on protochlorophyllide biosynthesis and growth responses type were more pronounced in the phyB mutant than in the WT seedlings, which can be connected with the higher phyA' content in the phyB mutant and/or phyB interference with its action in WT seedlings. In the phyA mutant induction of Pchlide633 and Pchlide655 biosynthesis was observed under constant FR, indicating that phyC may be responsible for this effect.

Solid‐State NMR Spectroscopy to Probe Photoactivation in Canonical Phytochromes

“…the C3 1 atom consists of a mixture of ( R )‐ and ( S )‐diastereomers as well as the alternative side‐chain conformations adopted by the two methionines, M174 and M269. The possibility that two isoforms exist is consistent with numerous spectroscopic studies over the past 15 years for both plant and Cph1 phytochromes and also noted in the more primitive bacteriophytochromes . There is no evidence that these two isoforms result from preparation artifacts , but even if it were to be the case, still the fact would remain that one state, Pr, tends to exist in two isoforms, whereas the other, Pfr, is structurally homogenous.…”

Electronic transitions and heterogeneity of the bacteriophytochrome Pr absorption band: An angle balanced polarization resolved femtosecond VIS pump–IR probe study

“…Schmidt et al showed the existence of an isosbestic point between the main absorption peak and the shoulder upon temperature change in oat phytochrome (33). These findings were in contrast to a single electronic transition and were interpreted as heterogeneity of the Pr form as reported by Sineshchekov et al (14). Single molecule fluorescence studies on native Agp1 bacteriophytochrome showed clear evidence for heterogeneity at very low temperatures (13).…”

“…Heterogeneity of the chromophore ground state geometry can induce a broadening of absorption lines in the visible absorption spectrum and new, to our knowledge, spectral features in the vibrational pattern due to diverse interactions between the chromophore and the protein binding pocket, e.g., hydrogen bonds (11). For the Pr form, it is still under debate as of this writing whether the shoulder in the absorption spectrum around 620 nm and the biexponential decay with time constants of~3 ps and 30 ps of the isomerization dynamics originate from heterogeneity of the chromophore (12)(13)(14). With our method we are able to identify chromophore heterogeneity and separate contributions from different electronic transitions.…”

Fluorescence and Photochemical Investigations of Phytochrome in Higher Plants

“…The major ones are light-labile phytochrome A (phyA) and light-stable phytochrome B (phyB). phyA mediates all the types of the photoresponses-VLFR, HIR, and also LFR [17][18][19][20][21][22][23][24]. phyB is capable of mediating only the inductive LFR type.…”