Victor Gaba

Higher plants and UV-B radiation: balancing damage, repair and acclimation

Abstract Although UV-B is a minor component of sunlight, it has a disproportionately damaging effect on higher plants. Ultraviolet-sensitive targets include DNA, proteins and membranes, and these must be protected for normal growth and development. DNA repair and secondary metabolite accumulation during exposure to UV-B have been characterized in considerable detail, but little is known about the recovery of photosynthesis, induction of free-radical scavenging and morphogenic changes. A future challenge is to elucidate how UV-B-exposed plants balance damage, repair, acclimation and adaptation responses in a photobiologically dynamic environment.

Identification of a primary in vivo degradation product of the rapidly-turning-over 32 kd protein of photosystem II

The 32 kd photosystem II protein of plant chloroplasts is rapidly turned over in the light. The initial events in the degradation of the 32 kd protein were studied. A 23.5 kd breakdown product was identified in Spirodela oligorrhiza membranes using immunological analysis. The 23.5 kd polypeptide was shown to be derived from the amino‐terminal portion of the 32 kd protein using partial proteolytic fingerprinting. An in vivo precursor–product relationship between the 32 kd protein and the 23.5 kd polypeptide was kinetically demonstrated by radiolabeling and pulse‐chase experiments. The cleavage site yielding the 23.5 kd polypeptide was localized to a functionally active region (between helices IV and V) of the 32 kd protein. We propose that an alpha‐helix‐destabilizing ‘degradation’ sequence, bordered by arginine residues 225 and 238, is involved in the formation of the 23.5 kd polypeptide.

Accelerated Degradation of the D2 Protein of Photosystem II Under Ultraviolet Radiation

The D2 protein of photosystem II is relatively stable in vivo under photosynthetic active radiation, but its degradation accelerates under UVB radiation. Little is known about accelerated D2 protein degradation. We characterized wavelength dependence and sensitivity toward photosystem II inhibitors. The in vivo D2 degradation spectrum resembles the pattern for the rapidly turning over D1 protein of photosystem II, with rates being maximal in the UVB region. We propose that D2 degradation, like D1 degradation, is activated by distinct photosensitizers in the UVB and visible regions of the spectrum. In both wavelength regions, photosystem II inhibitors that are known to be targeted to the D1 protein affect D2 degradation. This suggests that degradation of the two proteins is coupled, D2 degradation being influenced by events occurring at the QB niche on the D1 protein.

PHOTORECEPTOR INTERACTION IN PLANT PHOTOMORPHOGENESIS: THE LIMITS OF EXPERIMENTAL TECHNIQUES AND THEIR INTERPRETATIONS

Abstract— Problems concerning the interpretation of interactions of higher plant photomorphogenetic receptors are discussed. The theory that action of a blue light photoreceptor serves only to maintain responsiveness to phytochrome (Responsiveness Theory) is demonstrated to be unable to be properly tested with present techniques. This theory is also unable to explain experimental results any better than an alternative theory that a blue light photoreceptor may require the presence of the active form of phytochrome to express its activity (Presence Theory). This tatter theory is also incapable of being fully tested. There does not appear to be an adequate current theory to explain photoreceptor interactions. Other issues discussed include the use of displacement transducers in growth studies, the induction of phytochrome‐type responses by blue light, and the relative importance of the photoreceptors. New data are introduced on the effect of blue light in the end‐of‐day growth response to phytochrome of the light‐grown Cucumis sativus L. hypocotyl, and on the light equivalence principle in the same species.

Amplified Degradation of Photosystem II D1 and D2 Proteins under a Mixture of Photosynthetically Active Radiation and UVB Radiation: Dependence on Redox Status of Photosystem II

Abstract— Plants exposed to a mixture of photosynthetically active radiation (PAR) and UVB radiation exhibit a marked boost in degradation of the D1 and D2 photosysteni II (PS II) reaction center proteins beyond that predicted by the sum of rates in PAR and UVB alone (amplified degradation). Becausee degradation driven by visible or UVB radiation alone is uncoupled from PS II redox status, it was therefore assumed that the mixed‐light‐amplified component of degradation would behave similarly. Surprisingly, amplified degradation proved to be coupled tightly to the redox status of PS II. We show that inactivation of the PS II water oxidation by heat shock or oxidation of the plastosemiquinone (QA‐) by silicomolybdate nullifies only the amplified component of degradation but not the basic rates of degradation under PAR or UVB alone. The data are interpreted to indicate that formation of plastosemiquinone or an active water‐oxidizing Mn4 cluster, is the UVB chromophore involved in amplified degradation of the D1 and D2 proteins. Furthermore, accumulation of QA‐by 3‐(3,4‐dichlorophenyl)‐1,1‐dimethylurea or 2‐bromo‐3‐methyl‐6‐isopropyl‐4‐nitrophenol stimulated the mixed‐light‐amplified degradation component. Thus, amplified degradation of the D1 and D2 proteins in mixed radiance of PAR plus UVB (which simulates naturally occurring radiance) proceeds by a mechanism clearly distinct from that involved in degradation under PAR or UVB alone.

Ultraviolet-B effects on Spirodela oligorrhiza: induction of different protection mechanisms

Abstract Ultraviolet-B (UV-B) tolerance in plants has mostly been correlated with the presence of screening pigments (e.g. flavonoids) or other reductions in leaf transmittance. We have exploited the rapid turnover of the Photosystem II reaction center protein D1 as a sensitive in vivo probe for UV-B damage. We found that the aquatic monocot, Spirodela oligorrhiza, protects itself from UV-B irradiance using at least three different mechanisms. In one case, protection is correlated to the presence of UV-B screening pigments; in the second, an elevated oxygen-radical detoxifying system parallels UV-B tolerance; while in a third, UV-B tolerance is related to a mechanism involving neither screening pigments nor increased radical scavenging capacity. This demonstrates that, in vivo, a plant can complement its UV-screening and attenuation strategies by other tactics as well.

Photocontrol of hypocotyl elongation in light-grown Cucumis sativus L. : A synergism between the blue-light photoreceptor and phytochrome.

An interaction is demonstrated between the effects of phytochrome and cryptochrome (the specific blue-light photoreceptor) in the inhibition of hypocotyl elongation of light-grown cucumber (Cucumis sativus L.) cv. Ridge Greenline seedlings. At certain fluence rates of blue light the total inhibition response is greater than the sum of the separate responses to each photoreceptor. The threshold for response to blue light is reduced at least 30-fold by additional red-light irradiation. The synergistic effect is demonstrated for two different fluence rates of red light. Synergism is mediated by phytochrome in both the cotyledons and the hypocotyl.

Degradation of the 32 kDa Photosystem II Reaction Center Protein in UV, Visible and Far Red Light Occurs Through a Common 23.5 kDa Intermediate

Abstract A characteristic 23.5 kD a degradation intermediate of the 32 kDa photosystem II reaction center protein is produced upon illumination in UV , visible and far red light. We suggest a similar degradation pathway is employed in these three spectral regions, even though the light can enter the system through different photo receptors.

Photoregulation of Protein Turnover in the PSII Reaction Center

The reaction center of photosystem II (PSII) consists of three proteins: The 32-kDa protein (32K, also referred to as D1), D2, and cytochrome b559 (1,2). It has several features in common with the reaction center from purple bacteria (3,4), including amino acid sequence homology in functional regions (3,5), arrangement of the transmembrane helices (6,7,8), and conservation of the binding sites for chlorophylls, pheophytins, quinones and a non-heme iron (3,4,6,7,9). Furthermore, 32K and the L-subunit of the bacterial reaction center are the site of triazine herbicide action (10–12), and point mutations at conserved residues in these proteins can confer herbicide resistance (3,13–15).

Light-requiring acifluorfen action in the absence of bulk photosynthetic pigments

Abstract The nitrodiphenyl ether herbicide acifluorfen requires light for phytotoxicity even though it alone cannot absorb light. All possible major pigment systems have been implicated as the photoreceptor. We tested whether carotenoids and chlorophyll are essential for phytotoxicity. We used green-photosynthetic (mixotrophic) tomato cell cultures, etiolated cells of the same line (containing carotenoids but no chlorophyll), and carotenoid-free white cells (by continuously culturing etiolated cells on norflurazon). All three cell culture types parallel plants insofar as the first measureable effect is membrane lipoxidation. Acifluorfen at 1 μ M had little effect in darkness, but strongly inhibited growth of all cultures in 40 μmol m −2 sec −1 white light. Acifluorfen at 0.1 μ M did not affect green cells in light, but inhibited the growth of white and etiolated cells. Action spectroscopy showed that 350-nm light was the most effective wavelength inhibiting the growth of white cells with 1 μ M acifluorfen, followed by 550-nm, 450-nm, cool-white fluorescent, and 630-nm light, with only a threefold difference between 350-nm and red light. Far-red light was ineffective. These data demonstrate that in this system, chlorophyll, carotenes, cryptochrome, flavins, and phytochrome cannot be the sole photoreceptor for acifluorfen action. Our data, along with all other published findings are consistent with two hypotheses: (a) that acifluorfen interacts with other moieties to produce broad-spectrum chromophore(s) that react(s) with oxygen, forming active-oxygen species in the light; (b) that acifluorfen stimulates the accumulation of chromophoric photodynamic molecules.