Both Subunits of the Dimeric Plant Photoreceptor Phytochrome Require Chromophore for Stability of the Far-red Light-absorbing Form

Abstract

The dimeric plant photoreceptor phytochrome is converted from its inactive red light-absorbing form (Pr) into the active far-red light-absorbing form (Pfr) upon light absorption. Dynamics of Pfr generation and of thermal Pfr-to-Pr conversion are of fundamental importance for inducing adequate responses to light signals. Here, we analyzed the role of subunit interactions on spectroscopic properties of dimeric phytochrome A. Using a coexpression system and affinity chromatography, we prepared mixed phytochrome dimers that can incorporate the essential chromophore only in one subunit. We demonstrate that such mixed dimers have unaltered difference spectra. In contrast, dark reversion differed greatly between Pfr-Pfr homodimers and Pfr-Pr heterodimers, the former being about 100-fold more stable. Temperature dependence of reaction rates revealed an additional stabilization of about 4 kcal/mol in homodimers. Consequences of these findings are discussed in relation to the biological function of, and functional diversification between, phytochrome family members.

Light-Independent Phytochrome Signaling Mediated by Dominant GAF Domain Tyrosine Mutants ofArabidopsisPhytochromes in Transgenic Plants

“…In this regard, red light can only produce a maximum of 85% Pfr for phyA owing to the spectral overlap of Pr and Pfr forms (Lagarias et al, 1987). The percentage of photoconversion is even lower for phyB due to dark reversion, especially under lower fluence rates of light (Hennig and Schä fer, 2001). Since the light-independent activation of the phyB Y276H mutant is not constrained by the photoequilibrium limitations of the wild type and its lack of dark reversion would sustain its activation in darkness (or under low fluence rates of light), the hyperactivated phenotype of the PHYB Y276H plants is consistent with a photoinsensitive, nondark-reverting allele of activated phyB.…”

“…Since phyA is an obligate homodimer (Jones and Quail, 1986;Hennig and Schä fer, 2001), heterodimers between phyA Y242H and wild-type phyA subunits could explain the dominant-negative phenotype. Indeed, heterodimerization could also explain the dominant-negative phenotypes observed by expression of variously truncated (and potentially weakly functional alleles) of both PHYA and PHYB in wild-type genetic backgrounds (Boylan et al, 1994;Wagner et al, 1996).…”

Characterization of the requirements for localization of phytochrome B to nuclear bodies

“…k d ϭ ln(2)͞t 1/2 , t 1/2 ϭ 180 s. Therefore, k d ϭ 3.8 ϫ 10 Ϫ3 s Ϫ1 . Because PfrPfr conformer of phyB has a much slower dark-reversion rate than that of PfrPr (24), the Pfr percent is underestimated by this method in high-fluence rates of R.…”

“…The k d of phyB PfrPr to PrPr was calculated from t 1/2 of yeast expressed phyB (23,24). k d ϭ ln(2)͞t 1/2 , t 1/2 ϭ 180 s. Therefore, k d ϭ 3.8 ϫ 10 Ϫ3 s Ϫ1 .…”

Phytochromes function as thermosensors inArabidopsis

“…Phytochromes are interconvertible photoreceptors (14)(15)(16): phyB switches between an inactive Pr state and an active Pfr state upon absorbing red-and far-red light respectively. In a thermal relaxation reaction, Pfr in the dark spontaneously reverts to the Pr state (17)(18)(19). We hypothesized that the reversion of Pfr to the inactive Pr state may contribute to the subsequent increase in expression of the warm temperature transcriptome during the night.…”