Survey
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
229 Photosynthesis Research 53: 229–241, 1997. c 1997 Kluwer Academic Publishers. Printed in the Netherlands. Regular paper Chloroplast redox regulation of nuclear gene transcription during photoacclimation? Dion G. Durnford & Paul G. Falkowski Environmental Biophysics and Molecular Biology Program, Department of Applied Science, Building 318, Brookhaven National Laboratory, Upton, NY 11973, USA Received 1 November 1996; accepted in revised form 26 February 1997 Key words: light-harvesting antenna, plastid signal, plastoquinone pool Abstract The role of the redox state of ferredoxin/thioredoxin within the chloroplast is well established for the feedback regulation of enzyme activity in the Calvin cycle. However, evidence has emerged also suggesting that chloroplast electron transport components regulate plastid and nuclear gene expression. Using the unicellular green alga, Dunaliella tertiolecta, as a model organism, we have shown that the cell acclimates to changes in growth irradiance by altering the abundance and activities of photosynthetic components, in particular the light harvesting complexes (LHC). Pharmacological data suggests that light intensity is sensed through the redox status of the plastoquinone pool leading to the regulation of nuclear encoded genes, such as Lhcb. This signal may be transduced through a redox regulated protein kinase that (in)directly interacts with the nuclear transcription apparatus. The redox state of the plastoquinone pool seems to play a pivotal role in sensing cellular energy status and in regulating photosynthetic capacity. Other cellular pathways, including carbon fixation, carbohydrate metabolism and nutrient assimilation have been shown to have feedback influences on photosynthesis, that may be sensed by the redox state of the plastoquinone pool. Introduction All organisms have, to varying degrees, the capacity to adjust metabolic pathways in response to environmental cues. Physiological acclimation requires signal transduction pathways and feedback mechanisms that link gene expression to changes in the environment. One of the most important cues for photosynthetic organisms is light. Phytochrome was the first light responsive element discovered, and the role of this chromophore in regulating both developmental and reproductive responses in higher plants is well described (Smith 1995). There is also compelling evidence for a blue light receptor(s) in cyanobacteria, eucaryotic algae and higher plants, although the molecular identity of this chromophore remains elu? The US Government’s right to retain a non-exclusive, royaltyfree licence in and to any copyright is acknowledged. sive (Short and Briggs 1994). Both phytochrome and the blue-light responsive elements appear to be much more sensitive to spectral quality or photoperiod rather than photon flux densities that are relevant to photosynthetic electron flow. However, some evidence has suggested that photoacclimation responses are transduced via metabolic signals arising from the photosynthetic apparatus. Here, we review the evidence for regulation of nuclear gene expression based on the redox status of specific carriers in the photosynthetic electron transport chain. For additional information and perspective, the reader is referred to other reviews on redox control of gene transcription (Allen 1993; Allen 1995; Levings and Siedow 1995). Yoshihiko Fujita, to whom this issue is dedicated, was amongst the first researchers to ascribe light-induced changes in the photosynthetic electron transport components to electron transport imbalances. Fujita and his collaborators’ pioneering work increased ICPC: PIPS No.: 139262 BIO2KAP pres712b.tex; 4/11/1997; 11:22; v.7; p.1 230 Figure 1. Dunaliella tertiolecta cultures grown under high (left) and low (right) irradiance. Under high light conditions (in excess of 1500 mol quanta m 2 s 1 ) the cell contains lower amounts of chlorophyll, a higher content of carotenoids per cell and a chl a/b ratio of around 8. The deep green culture on the right, containing more chlorophyll per cell, was grown under low light intensity (ca. 70 mol quanta m 2 s 1 ) and has a chl a/b ratio of around 4. Photograph is courtesy of Dr. Assaf Sukenik. our understanding of how a cell perceives changes in its light environment, especially as it relates to the adjustment of photosystem stoichiometry. In this review we consider Fujita’s observations, along with others, in the context of the experimental evidence indicating possible molecular mechanisms for sensing changes in photon flux density (PFD) and the transduction of the signal leading to photoacclimatory changes. Photoacclimation We define photoacclimation as reversible phenotypic changes in activities and/or concentrations of enzymes or other biosynthetic molecules, such as pigment protein complexes and photosynthetic electron transport components, that compensate for changes in spectral irradiance, independent of plant developmental pro- pres712b.tex; 4/11/1997; 11:22; v.7; p.2 231 cesses (Falkowski and LaRoche 1991). Photoacclimation occurs on a variety of time scales (Falkowski et al. 1994; Anderson et al. 1995). On short time scales of minutes to hours, plants undergo changes in response to high frequency (subdaily) variations in light intensity. These responses include the dissipation of excess excitation energy via xanthophyll cycle carotenoids (Demmig-Adams and Adams 1996) and by state transitions where reversible phosphorylation of LHC II leads to complementary alterations in the effective absorption cross section of PS II and PS I (Bonaventura and Myers 1969; Bennett 1983). As we shall describe, these phenomena are part of a nested series of photoacclimatory responses, which, on the longer time scales, can lead to changes in gene expression. The latter requires a signal transduction pathway that couples an irradiance sensor to both nuclear and plastid gene expression. Although many higher plants can photoacclimate (Anderson et al.1988; Chow et al. 1990; Kim et al. 1993; Anderson et al. 1995), their ability to do so is limited as compared to algae (Bennett 1987; Falkowski and LaRoche 1991). In higher plants a light gradient can exist within the leaf, generating degrees of sun and shade adapted chloroplasts within a single plant (Nishio et al. 1994). This gradient reveals physiological plasticity that is coupled to development through differentiation. However, many higher plants have evolutionarily adapted (i.e. are selected) to either a sun or shade environment and display phenotypes characteristic of either one or the other light regime (Boardman 1977). In contrast, algae generally have an extremely high degree of physiological plasticity that permits rapid and reversible adjustments to changes in the average growth irradiance. Green unicellular algae in particular have a remarkable ability to survive, and acclimate to, a wide range of irradiance levels. Over the past several years, we have studied photoacclimation in the halotolerant chlorophyte, Dunaliella tertiolecta. This organism is easy to culture, has a single cupshaped chloroplast, lacks a cell wall, does not undergo sexual recombination (and therefore the study of irradiance responses can be distinguished from developmental processes), and displays changes in photoacclimatory variables such as cellular chlorophyll that are large and easily measured (Figures 1 and 2). Within a few hours of a transition from high light (e.g. 700 mol quanta m 2 s 1 ) to low light (70 mol quanta m 2 s 1 ), D. tertiolecta redirects the end products of carbon fixation (i.e. carbohydrates) towards the production of proteins. One of the first detectable Figure 2. Kinetics of photoacclimation in Dunaliella tertiolecta following a high light (700 mol quanta m 2 s 1 ) to low light (70 mol quanta m 2 s 1 ) transition. (A) Changes in chlorophyll per cell (open triangles) and the Chl a/b ratio (closed circles) over a 5 day period. (B) Kinetics of LHC II (closed circles) and RC II changes (open triangles) during a high light to low light transition. Figure is a modification from Sukenik et al. (1990). proteins to change is LHC II. A 10 fold decrease in irradiance induces a 4 fold increase in Lhcb mRNA within 9 hours, and ultimately leads to a 3-fold increase in LHC II complexes (LaRoche et al. 1991). Nuclear run-on transcription and mRNA stability experiments have established that Lhcb gene expression is regulated at the level of transcription (Escoubas et al. 1995). The changes in LHC II are subsequently followed by a net increase in other components of the photosynthetic electron transport chain, such that the ratios of the components remain unchanged, irrespective of the growth irradiance (Sukenik et al. 1987a, 1990). A new steady state level of all photosynthetic components is reached within ca. 72 h (Figure 2). pres712b.tex; 4/11/1997; 11:22; v.7; p.3 232 Hypotheses to account for the photoacclimation response What is the molecular basis of photoacclimation? At least three hypotheses can be developed to account for the light intensity induced changes in Lhc gene expression. The first postulates a photoreceptor, such as phytochrome, or a blue light receptor(s) that somehow senses variations in photon flux density and causes changes in nuclear gene expression. A second hypothesis invokes a feedback from a light intensity regulation of chlorophyll synthesis (which is confined to the plastid), to either gene expression and/or protein stability. The third hypothesis proposes a feedback to photosynthetic electron transport or some end product(s) of photosynthetic metabolism that influences gene expression. Let us examine the evidence for each hypothesis noting that they are not mutually exclusive. Evidence for a photoreceptor Photoreceptors are characterized by spectrally-induced reversible responses that are not directly coupled to metabolism. The best characterized photoreceptors are the phytochromes, a conserved family of proteins complexed with an open tetrapyrrole chromophore. The chromophore is activated through a red light induced conversion of the Pr to the Pfr form. In higher plants there are two main types of phytochromes (Furuya and Schäfer 1996), and there is evidence for two distinct signal transduction pathways, one involving G protein activation and the other being calcium/calmodulin dependent (Bowler et al. 1994). The evolutionary origin of phytochrome is unclear. The chromophore has structural similarities to phycobilins, including the covalent bonding of the tetrapyrrole to the protein via a sulfide bridge from cysteine. Phytochrome-like gene sequences have been reported in the green alga, Mesotaenium; however, red/far red reversibility has not been demonstrated (Morand et al. 1993). No phytochrome sequences have been detected in a genomic library of Dunaliella tertiolecta (Harry Smith, personal communication), and attempts to isolate the chromophore from green algae have not been successful (HW Siegelman, personal communication). Although there is some evidence for phytochromelike proteins in chlorophyte algae, there is little evidence indicating a role for phytochrome in light mediated responses in unicellular algae. Perhaps this should not be surprising. In aquatic systems, spectral irradi- ance is highly modified by the medium itself. Absorption and scattering by water leads to rapid depletion of red and far-red light with depth, but permits a large fraction of blue and blue/green light to penetrate (Kirk 1994). Hence, chromophores keyed to red/far-red variations would seem to have little value for planktonic cells in aquatic environments. While phytochrome probably does have cyanobacterial origins (Hughes et al. 1997), it would appear that the utilization of this chromophore as a major light responsive element emerged relatively late in the evolution of photosynthetic organisms, and has been utilized in terrestrial plants primarily as a sensor of photoperiod rather than of light intensity (Anderson et al. 1995). Some cyanobacteria respond to changes in the relative level of red and green irradiance (chromatic adaptation) by adjusting phycoerythrin and /or phycocyanin levels within the phycobilisome; this response is thought to be sensed by a photoreceptor (Grossman et al. 1993). A possible candidate for this receptor has been identified that has some sequence similarity to both phytochrome and ethylene receptors (Kehoe and Grossman 1996). Chromatic adaptation is independent of electron transport rates in the filamentous cyanobacterium, Calothrix (Campbell et al. 1993), adding further evidence for a light receptor mediated response. Blue light responses have been more extensively examined in higher plants and are attributed to the control of numerous cellular processes (reviewed by Short and Briggs 1994). In algae, a blue light receptor(s) is implicated in the regulation of LHC expression (Kindle 1987; Humbeck et al. 1988; Hermsmeier et al. 1991; Melis et al. 1996), cell division (Munzner and Voigt 1992) and chlorophyll biosynthesis (Matters and Beale 1995). In Chlamydomonas, the induction of Lhc expression in blue light is DCMU-insensitive, suggesting that the signal is independent of photosynthesis (Kindle 1987). However, other studies using Chlamydomonas and Scenedesmus indicate that light intensity and light quality are both factors influencing photoacclimation (Humbeck et al. 1988; Melis et al. 1996). While reversible spectral responses are often interpreted as evidence for a photoreceptor, in algae it is difficult to distinguish between the effects of light quality on a specific target molecule and indirect effects on photosynthetic electron transport (as will be discussed below). In contrast to higher plant leaves, because individual algal cells are optically thin, their optical absorption cross section are highly spectrally dependent. Hence, exposing cells to blue light and red light pres712b.tex; 4/11/1997; 11:22; v.7; p.4 233 at equal photon fluence rates does not translate to an equal rate of photon absorption. Rather, for most algal systems, the Soret bands of the antenna pigments are more efficient at photon absorption than the Q bands of chlorophylls; there is virtually no absorption by cells in far-red light. A physiologically more meaningful test for spectral responses that are independent of photosynthesis would be to expose cells to different wavelengths of light at fluence rates that elicit identical electron transport rates. To our knowledge, such experiments have not been performed. This issue becomes critical to evaluating potential signal transduction pathways. For example, in cyanobacteria light intensity and light quality can be manipulated to give the same photoacclimation state; however, the responses have been correlated with changes in the redox poise of the plastoquinol pool rather than with a photoreceptor (Fujita et al. 1987; Murakami and Fujita 1991b, 1993). Evidence for feedback from the chlorophyll biosynthetic pathway Regulation of LHC expression during photoacclimation can be hypothesized to occur indirectly through the light controlled biosynthesis of chlorophyll. Two metabolic steps were identified as potentially important areas in the regulation of chlorophyll biosynthesis: the synthesis of -amino levulinic acid (ALA) and the conversion of protochlorophyllide to chlorophyllide (Reinbothe and Reinbothe 1996). The primary substrate for synthesis of the tetrapyrrole precursor, ALA, is glutamic acid and its synthesis occurs in a three step reaction that utilizes ATP, NADPH and a tRNAglu . One of enzymes involved in this pathway, glutamate-1-semialdehyde aminotransferase, is tightly regulated at the transcriptional level indicating a potentially important step in the photo-regulation of chlorophyll biosynthesis (Matters and Beale 1994, 1995). Another potential light responsive point in the chlorophyll biosynthetic pathway is the photocatalyzed reduction of protochlorophyllide to chlorophyllide (Reinbothe et al. 1996). In angiosperms, protochlorophyllide oxidoreductase (POR) is light dependent, thus the completion of the chlorophyll biosynthetic pathway requires light. Many, but not all, algae are able to synthesize chlorophyll in the dark due to the presence of a light-independent POR. Conifers, which can also synthesize chlorophyll in the dark, seem to have both a light-dependent and light-independent form of POR (see Reinbothe and Reinbothe 1996). The abili- ty of some organisms, especially algae, to synthesize chlorophyll in the dark might suggest that low light or dark induced changes in chlorophyll levels are not solely attributed to the regulatory feedback of protochlorophyllide. Light intensity induced chlorophyll accumulation could, in principle, lead to an increased synthesis of reaction centers and antenna complexes that would be capable of increasing the effective absorption cross section of the photosynthetic apparatus (Ley and Mauzerall 1982). However, the hypothesis implies that chlorophyll exerts its control through the posttranslational stabilization of LHC apoproteins and/or through mRNA stabilization or transcriptional regulation, signaled by a chlorophyll biosynthetic intermediate. Post-translational stabilization of LHC II apoproteins by the binding of chlorophyll has been reported for higher plants and green algae (Apel and Kloppstech 1980; Michel et al. 1983). To test if this mechanism could account for the LHC II changes induced by decreasing irradiance levels, Mortain-Bertand et al. (1990) blocked the formation of ALA from glutamate with gabaculine and found that 35 S labeled LHC II apoproteins were stable in the light. However, the cells did not accumulate more LHC II apoproteins upon transfer to low light. These experiments suggest that while chlorophyll synthesis is not required in Dunaliella for accumulation of LHC II apoproteins, the posttranslational stabilization by chlorophylls is required for an increase in total cellular LHC II levels during acclimation to reduced irradiance levels. LHC synthesis may also be controlled at the transcriptional level through feedback regulation by chlorophyll biosynthesis intermediates. In the green algae Chlamydomonas and Dunaliella, the presence of gabaculine or levulinic acid did not affect the normal light induced increase in Lhc mRNA accumulation through dark to light or high-light to lowlight transitions (Johanningmeier and Howell 1984; LaRoche et al. 1991). These latter results suggested that chlorophyll biosynthesis precursors between ALA and chlorophyll are not required for the observed, enhanced rate of Lhc gene expression. However, blocking synthesis at later stages with inhibitors or by mutation caused reductions in the accumulation of Lhc mRNA following dark to light transitions (Johanningmeier and Howell 1984). These changes were later correlated to transcription rates by nuclear run-on assays (Jasper et al. 1991) and were interpreted as evidence that precursors of chlorophyll biosynthesis pres712b.tex; 4/11/1997; 11:22; v.7; p.5 234 feedback negatively to regulate Lhc transcription and hence accumulation. It should be stressed that acclimation to low light is not equivalent to deprivation of light. Cells entrained in a light/dark cycle exhibit a diel cycle in cellular chlorophyll, typically with an increase in chlorophyll immediately prior to the photophase and a decrease prior to the scotophase. This rhythm is opposite that of photoacclimation. Yet cells entrained in a light/dark cycle acclimate to the average photon flux density to which they are exposed within a photoperiod (Post et al. 1984). Thus, while the results of LaRoche et al. (1991) and Johanningmeier and Howell (1984) may appear to conflict, both studies indicate that chlorophyll biosynthesis is requisite for acclimation to low photon fluence rates; however, the signalling for increased Lhcb mRNA transcription does not appear to require a chlorophyll precursor. Evidence for feedback from photosynthetic electron transport The first clue implicating a role of the chloroplast electron transfer components in the control of nuclear gene expression came from the observations of Beale and Appelman (1971) who reported that when Chlorella cultures were grown in the presence of sublethal concentrations of DCMU (0.5-510 6 M) cellular chlorophyll increased. As LHC II, let alone its gene sequence, had not yet been discovered, the interpretation of these results was limited to a vague understanding that photosynthetic metabolism influenced the abundance of pigments within the cell. Subsequent studies confirmed the increase in chlorophyll per cell in other unicellular green algae (Nau and Melis 1992; Escoubas et al. 1995). Koenig (1990) reported that the addition of atrazine to Anacystis cultures, led to an increased synthesis of phycocyanin, a component of the light harvesting phycobilisome complexes, that mimicked a shade acclimated phenotype. Koenig suggested D1 somehow sensed light intensity. Fujita and colleagues explored the interaction between photosynthetic electron transport and the regulation of photosynthesis in cyanobacteria by taking advantage of the strong spectral differences in the optical absorption cross sections of the two photosystems in these organisms. Using different spectral qualities of irradiance, rather than pharmacological inhibitors, they were able to demonstrate alterations in PS I/PS II stoichiometry (Fujita et al. 1987; Murakami and Fujita 1991a, b, 1993). In Synechocystis, light quality and quantity induced similar changes (Murakami and Fujita 1991a). This was hypothesized to occur in order to maintain an optimal quantum efficiency, and was somehow regulated via the redox state of the plastoquinone pool or the Cyt b6 f complex; hence it was central to the photoacclimation process. The effects of DCMU on gene expression were examined by Escoubas et al. (1995), who reported that the addition of the herbicide to high-light grown cultures of D. tertiolecta induced a two fold increase in Lhc mRNA levels. These changes resembled a response to a reduction in the light intensity (LaRoche et al. 1991) correlating DCMU action to the process of photoacclimation as observed with cyanobacteria (Koenig 1990). Moreover, since DCMU specifically acts within the chloroplast by inhibiting the reduction of plastoquinol (Trebst 1980), these data provided a specific link between chloroplast function and the regulation of nuclear gene transcription. When D. tertiolecta cultures were grown in the presence of DBMIB (10 7 M) at low light intensity, photosynthetic electron transport was reduced by half though there was little observable effect on respiration (Escoubas et al. 1995). Application of the inhibitor resulted in a reduction in the amount of chlorophyll per cell and a 75% decrease in the steady state Lhc mRNA levels even though the cultures were maintained under low light conditions. DBMIB prevents the reoxidation of plastoquinol (Trebst 1980) and mimicks an increase in light intensity that is antithetic to the response seen with DCMU treatment under high light. The inverse effects of DCMU and DBMIB strongly suggest that the balance between the reduced and oxidized forms of plastoquinone are involved in the sensing of light intensity. Signal transduction and nuclear-chloroplast interactions In Dunaliella the changes in nuclear encoded Lhc mRNA levels in response to chloroplast specific inhibitors of photosynthetic electron transport demonstrate that a plastid to nucleus signal must exist. Evidence suggesting the existence of such a signal or ‘plastid factor’ originally arose through the examination of photooxidative stress in plant seedlings (Oelmüller 1989; Taylor 1989). Seedlings deficient in carotenoids, either through mutation or norflurazon treatment, were exposed to high light to induce pres712b.tex; 4/11/1997; 11:22; v.7; p.6 235 photooxidation. Under non-photooxidative light conditions (ie. grown under low intensity white light or farred light) the seedlings accumulate both chlorophyll and Lhc mRNA (Oelmüller and Mohr 1986; Burgess and Taylor 1987). However, upon exposure to high intensity light, chlorophyll is destroyed through photooxidation and there is a rapid reduction in the amount of detectable Lhcb, RbcS, and plastocyanin mRNA (Mayfield and Taylor 1984; Oelmüller and Mohr 1986; Burgess and Taylor 1987). This was attributed primarily to transcriptional changes (Burgess and Taylor 1988). These studies addressed chloroplast biogenesis, not photoacclimation, and are complicated by the fact that the seedlings have an independent energy source. Moreover, one of the Arabidopsis GUN mutants (for genomes uncoupled) that expresses Lhcb genes under norflurazon induced photooxidation, has wild type levels of Lhcb expression under a variety of light quality, quantity and exogenously applied sucrose levels in mature plants (Susek et al. 1993). This indicates that there may be a separate signal transduction pathway for chloroplast development and photoacclimation. While the exact nature of this plastid factor(s) is unknown, some of its characteristics can be inferred indirectly using chlorophyll per cell as an indicator for photoacclimation in Dunaliella. In Dunaliella, a high light to low light transition results in a three fold increase in the amount of chlorophyll per cell that correlates to changes in Lhc mRNA abundance (LaRoche et al. 1991) and an increase in transcription (Escoubas et al. 1995). However, the addition of sub-micromolar amounts of the cytosolic phosphatase inhibitors okadaic acid, microcystin-LR or tautomycin prevents the increase in chlorophyll per cell by about 44%, partially inhibiting the normal photoacclimatory response. This suggests that phosphorylation of a protein in the cytosol or nucleus is involved in the light intensity induced signal transduction pathway. It was proposed that in Dunaliella Lhc gene transcription was repressed under high irradiance and activated under low light (LaRoche et al. 1991). Several putative regulatory motifs were detected in a Dunaliella Lhc promoter by homology to those found in higher plants and were hypothesized to be involved in light intensity mediated control (Escoubas et al. 1995; Terzaghi and Cashmore 1995). Utilizing the electrophoretic mobility shift assay, a DNA binding complex, found only in high light cellular extracts, was identified (Escoubas et al. 1995; Durnford and Falkowski, unpublished data). Binding specificity was localized to a 100 bp region at or near a G-box motif (Durnford and Falkowski, unpublished data). This motif has been implicated in light mediated responses for a variety of nuclear encoded genes (Terzaghi and Cashmore 1995). Binding kinetics of this DNA-protein complex during light transition experiments are correlated with Lhc mRNA levels suggesting a regulatory role for the DNA binding protein during photoacclimation (Durnford and Falkowski, unpublished data). Molecular basis of redox regulation: The protein phosphorylation hypothesis A sudden change in light intensity will immediately cause an imbalance in the electron flow between PS II and PS I. Upon a transfer to a higher irradiance (Figure 3B) there is an over-excitation of PS II resulting in a reduction of the plastoquinone pool that exceeds its reoxidation by the cytochrome b6 f complex. This imbalance in the redox state is sensed by a thylakoid bound protein kinase(s) (Bennett 1991; Allen 1992). Once activated, the thylakoid kinase(s), are known to phosphorylate LHC II. This induces a state II condition where a population of phosphorylated LHC II becomes dissociated from the PS II reaction center (Bonaventura and Myers 1969; Bennett 1983). Consequently, the excitation pressure at PS II is reduced, alleviating the imbalance in electron transport sensed through the redox state of the plastoquinone pool. A similar activation pathway may occur if these shortterm adjustments are not sufficient to fully compensate for the imbalance in the redox status. Electron transfer through the plastoquinone pool is the rate limiting step of the light reactions and is an ideal sensor site, as it functions like a capacitor, storing electrons and buffering against minor variations in the redox rates at Qa and the Cyt b6 f complex. Signal transduction could conceivably involve a chloroplast phosphoprotein intermediate (CPP, Figure 3A) that is released from the membrane upon phosphorylation (Bhalla and Bennett 1987). The mode of signal transduction between the chloroplast and nucleus is unknown although the phosphatase inhibitor experiments suggest the involvement of a phosphorylation cascade. Lhc gene repression under high light intensity may be mediated by the specific interaction of cis-acting regulatory proteins binding to the Lhc promoter. There is evidence correlating an increase in light intensity to the rapid appearance of a specific DNA binding protein and a reduction in mRNA levels, suggestive of a repressor (Durnford and Falkows- pres712b.tex; 4/11/1997; 11:22; v.7; p.7 236 Figure 3. A working hypothesis for the molecular basis of photoacclimation in Dunaliella tertiolecta. In the low light acclimated state (A) Dunaliella contains a larger number of reaction centers and increased levels of LHC II. Lhc transcription rates and mRNA abundance are high under such conditions. Upon a transition to a high light environment (B), the plastoquinone pool becomes reduced (PQrd ) due to excess excitation energy. Under such conditions the redox active LHC II kinase(s) (LK) would become activated and induce state transitions via the reversible phosphorylation of LHC II. The activation of the same or a similar redox sensitive kinase may initiate a signal transduction pathway involving a chloroplast phosphoprotein (CPP). A phosphorylation cascade may trigger the repression of Lhc gene transcription, preventing the accumulation of unnecessary transcripts. These processes eventually lead to a new steady state level of components acclimated to the higher irradiance level (C). In a reverse transition, the plastoquinone pool would become predominantly oxidized (PQox ) due to the reduced availability of light. This would cause an inactivation of the kinase and signal the resumption of Lhc transcription through a removal of the DNA binding complex (D). ki, unpublished data). Lowering light intensity and consequently permitting the re-oxidation of the plastoquinone pool would relax the redox signal (Figure 3D). This could lead to the reverse cellular response, alleviating the repression of Lhc gene expression and allowing for the synthesis of additional antennae during photoacclimation. Coordination between electron transport and gene regulation Photosynthetic electron transport is sensitive to many environmental and cellular metabolic cues in addition to light. Amongst these are temperature, nutrient availability, and carbon metabolite levels. All of these cues have observed effects on photosynthetic rates and chloroplast protein levels. Understanding the interactions between these diverse pathways and signals is essential to fully comprehend the regulation of cellular processes. Let us first consider the photosynthesis- pres712b.tex; 4/11/1997; 11:22; v.7; p.8 237 irradiance response and later relate it to the above mentioned environmental and cellular cues. At low PFD, the rate of photon absorption by PS II is a product of the functional absorption cross section, PSII , and spectral irradiance, I. As irradiance increases to a saturated value (Pmax ), the assimilation rate of electrons is determined by the product of the number of photosynthetic units, n, and the maximum turnover rate 1/ . There is a single point on the photosynthesisirradiance (PI) curve where the rate of photon absorption exactly balances that of electron consumption: PSII I = n= (1) The irradiance level at which this balance occurs is the intercept of the initial slope of the PI curve divided by Pmax , and is sometimes designated Ik in the biophysical literature. The rate limiting step in light saturated photosynthesis is carbon fixation, not photosynthetic electron transport (Sukenik et al. 1987b). A transition from a low- to a high-light environment would necessitate a reduction in the number of reaction centres and/or the size of the contributing light-harvesting antennae in order to avoid chronic photoinhibition as a result of feedback from the dark reactions (Smith et al. 1990). It should be noted that Rubisco is not a light-regulated gene in Dunaliella, hence changes in growth irradiance lead to marked changes in the ratio of photosynthetic electron components to Rubisco. The changes in these ratios are linearly correlated with 1/ (Sukenik et al. 1987b, Fisher et al. 1989). Thus, in principle, any process that affects the assimilation of photosynthetically generated electrons in PS II could lead to a feedback to the biosynthetic components of the electron transport chain via the redox poise of the PQ pool. Temperature acclimation studies in algae can also be interpreted with regard to Equation 1. Temperature acclimation responses in different algae are variable; however, they usually involve changes in a number of cellular processes, including alterations in maximum photosynthetic rates, the optimum temperature for photosynthesis, and changes in the activity of Rubisco and other Calvin cycle enzymes (Davison 1991). Generally, the chlorophyll content per cell decreases with reductions in growth temperature (Morris and Glover 1974; Thompson et al. 1992) though little change (Morris and Glover 1974; Levasseur et al. 1990) or even an increase in chlorophyll per cell (Mortain-Bertrand et al. 1988) has been observed in some species. Manipulations of both light intensity and temperature with green algal cultures demonstrate that a high light-acclimated phenotype, defined by a high chlorophyll a/b ratio and low amounts of chlorophyll per cell, can be induced under low light conditions simply by decreasing the growth temperature (Maxwell et al. 1994). This light/temperature regime caused reductions in LHC II protein and steady state mRNA levels that were correlated to the ‘excitation pressure’ at PS II (i.e. the product of PSII and I) and, consequently, the redox state of the plastoquinone pool (Maxwell et al. 1995). Under low temperature conditions, the rate of enzyme mediated reactions is expected to decrease significantly (Davison 1991), causing reductions in the carbon fixation rate and consequently increasing . The cell could acclimate by an increase in the ratio of Calvin cycle enzymes to electron transport components (Mortrain-Bertrand et al. 1988), thus decreasing . Alternatively, the functional absorption cross section (PSII ) could be reduced by limiting the energy transfer efficiency to PS II (Levasseur et al. 1990) or reducing the LHC II antennae size to prevent over excitation of the photosystems (Davison 1991; Maxwell et al. 1995). Both types of responses are predicted through the equation based on the PI curve. In this sense, the response to temperature is similar to the phenotypic changes observed during photoacclimation (Davison 1991) that may be responding to a common cellular signal originating from the plastoquinone pool. However, thermal and photoacclimation are not entirely equivalent responses and regulatory differences are apparent (Machalek et al. 1996). Metabolic coordination Photoacclimation requires the coordination of both nuclear and chloroplast gene expression in order to synthesize both reaction center and antennae complexes. Thioredoxin, which undergoes a reversible, photosynthesis-dependent reduction of a disulphide bond, is important in the regulation of the reductive pentose phosphate cycle, and functions by altering the redox state of several key enzymes (Buchanan 1991). Thioredoxin has also been implicated in the translational regulation of the chloroplast encoded psbA transcript (D1 protein) in the green alga Chlamydomonas (Danon and Mayfield 1994). In this study a purified protein complex could be induced to bind to the 50 untranslated region of the psbA mRNA transcript by reduction of the extract with DTT or with a reduced thioredoxin preparation. Artificially oxidizing the puri- pres712b.tex; 4/11/1997; 11:22; v.7; p.9 238 fied mRNA protein complex (RNP) eliminated binding of the complex in vitro. These results are consistent with environmental factors that could control the translation of the psbA protein via the redox state of thioredoxin. Such factors as light, temperature or CO2 supply could therefore couple the rate of photosynthetic electron transport to plastid gene expression. The processes of photosynthetic electron transport, CO2 fixation and nitrogen metabolism are tightly coupled due to the requirements for ATP, reductant and protein cofactors such as ferredoxin or thioredoxin. The assimilation of nitrate for the biosynthesis of amino acids and other molecules requires photosynthetically reduced ferredoxin. In plants and algae, nitrogen limitation suppresses many cellular functions, including chlorophyll and protein synthesis, and the expression of photosynthesis-related genes (Turpin 1991; Huppe and Turpin 1994). In Chlamydomonas, nitrogen limitation reduces protein and chlorophyll levels, as well as the expression of Lhc and RbcS nuclear encoded transcripts, while increasing the formation of starch (Plumley and Schmidt 1989). These responses resemble acclimation to high light, suggesting common or interacting sensing and signal transduction pathways for the control of nuclear gene expression. In another green alga, Selenastrum, the ATP dependent assimilation of NH4 + induces a reversible state transition, a short-term photoacclimation response (Turpin and Bruce 1990). The decreased requirement for reductant and ferredoxin in this process, as compared to nitrate uptake, is thought to limit the rate of photosynthetic electron transport, causing a plastoquinone redox state-mediated state transition in an attempt to compensate for changes in . In principle, this provides a mechanism linking nitrogen metabolism to photosynthesis through a feedback regulatory network although it does not preclude the existence of other regulatory points. The processes of starch synthesis, typically occurring in the light, and degradation, occurring in the dark, are keyed to the cellular energy status and are tightly regulated to prevent futile cycling within the chloroplast. The addition of an exogenous energy source such as glucose or acetate has an inhibitory effect upon photosynthetic rates, and on the accumulation of chlorophyll and nuclear transcripts in light-grown cultures (reviewed by Sheen 1994; Koch 1996). Both Lhc and RbcS gene expression in algae and plants are negatively affected in the presence of an exogenous carbon source (Kindle 1987; Kilb et al. 1996), and this is likely a universal response to the addition of carbon metabolites (Sheen 1994). Run-on transcription assays (Krapp et al. 1993) and promoter deletion studies (Sheen 1990) utilizing photosynthetic genes demonstrated that these responses are transcriptionally regulated by specific factors. Hexokinase substrates appear to be pivotal in the feedback regulation of sugar metabolism; hexokinase somehow signals changes in cellular energy status, which in turn regulates photosynthetic components (Jang and Sheen 1994). However, it will be difficult to distinguish between regulatory contributions from sugar metabolic enzymes and those signals originating as a result of feedback to photosynthesis. The complex interactions linking cellular metabolism, nutrient assimilation and photosynthesis to regulatory processes in the nucleus are summarized in Figure 4. Research directed towards understanding the regulatory feedback and signal transduction interactions involved in acclimation to light and other environmental cues is in its infancy. Understanding the interactions between these regulatory pathways is essential to elucidating how photosynthetic organisms integrate multiple environmental cues simultaneously in order to coordinate the synthesis and maintenance of the myriad of proteins and secondary gene products that constitute the photosynthetic apparatus. Figure 4. Summary of proposed interactions between cellular metabolism and photosynthesis in the control of nuclear and chloroplast gene expression. The figure suggests a complex web of interaction controlling gene expression through a feedback to photosynthesis as well as direct, independent signals for the sensing of cellular energy status. pres712b.tex; 4/11/1997; 11:22; v.7; p.10 239 Acknowledgments This research was supported by the US Department of Energy under contract No. DEAC02-76CH00016. References Allen JF (1992) Protein phosphorylation in regulation of photosynthesis. Biochim Biophys Acta 1098: 275-335 Allen JF (1993) Redox control of transcription: sensors, reponse regulators, activators and repressors. FEBS Lett 332(3): 203-207. Allen JF (1995) Thylakoid protein phosphorylation, state1-state 2 transitions, and photosystem stoichiometry adjustment: redox control at multiple levels of gene expression. Physiol Plantarum 93: 196-205 Anderson JM, Chow WS and Goodchild DJ (1988). Thylakoid membrane organisation in sun/shade acclimation. Aust J Plant Physiol 15: 11–26. Anderson JM, Chow WS and Park YI (1995) The grand design of photosynthesis: acclimation of the photosynthetic apparatus to environmental cues. Photosynth Res 46: 129–139 Apel K and Kloppstech K (1980) The effect of light on the biosynthesis of the light-harvesting chlorophyll a/b protein. Planta 150: 426–430 Beale SI and Appleman D (1971) Chlorophyll synthesis in Chlorella. Plant Physiol. 47: 230–235 Bennett J (1983) Regulation of photosynthesis by reversible phosphorylation of the light harvesting chlorophyll a/b protein. Biochem J 212: 1–13 Bennett J (1991) Protein phosphorylation in green plant chloroplasts. Annu Rev Plant Physiol Plant Mol Biol 42: 281–311 Bennett J, Schwender JR, Shaw EK, Tempel N, Ledbetter M and Williams RS (1987) Failure of corn leaves to acclimate to low irradiance. Role of protochlorophyllide reductase in regulating levels of five chlorophyll-binding proteins. Biochim Biophys Acta 892: 118–129 Bhalla P and Bennett J (1987) Chloroplast phosphoproteins: Phosphorylation of a 12-kDa stromal protein by the redox-controlled kinase of thylakoid membranes. Arch Biochem Biophys 252(1): 97–104 Boardman NK (1977) Comparative photosynthesis of sun and shade plants. Ann Rev Plant Physiol. 28: 355–377 Bonaventura C and J. Myers (1969) Fluorescence and oxygen evolution from Chlorella pyrenoidosa. Biochim Biophys Acta 189: 366–383 Bowler C, Neuhaus G, Yamagata H and Chua NH (1994) Cyclic GMP and calcium mediate phytochrome phototransduction. Cell 77: 73–81 Buchanan BB (1991) Regulation of CO2 assimilation in oxygenic photosynthesis: The ferredoxin/thioredoxin system. Arch Biochem Biophys 288(1): 1–9 Burgess DG and Taylor WC (1987) Chloroplast photooxidation affects the accumulation of cytosolic mRNAs encoding chloroplast proteins in maize. Planta 170: 520–527 Burgess DG and Taylor WC (1988) The chloroplast affects the transcription of a nuclear gene family. Mol Gen Genet 214: 89–96 Campbell D, Houmard J and Tandeau de Marsac N (1993) Electron transport regulates cellular differentiation in the filamentous cyanobacteria Calothrix. Plant Cell 5: 451–463 Chow WS, Melis A and Anderson JM (1990) Adjustment of photosystem stoichiometry in chloroplasts improve the quantum efficiency of photosynthesis. Proc Natl Acad Sci USA 87: 7502–7506 Danon A and Mayfield SP (1994) Light-regulated translation of chloroplast messenger RNAs through redox potential. Science 266: 1717–1719 Davison IR (1991) Environmental effects on algal photosynthesis: temperature. J Phycol 27: 2–8 Demmig-Adams B and Adams WW (1996) The role of xanthophyll cycle carotenoids in the protection of photosynthesis. Trends Plant Sci 1(1): 21–26 Escoubas JM, Lomas M, LaRoche J and Falkowski PG (1995) Light intensity regulation of cab gene transcription is signaled by the redox state of the plastoquinone pool. Proc Natl Acad Sci USA 92: 10237–10241 Falkowski PG and LaRoche J (1991) Acclimation to spectral irradiance in algae. J Phycol 27: 8–14 Falkowski PG, Greene R and Kolber Z (1994) Light utilization and photoinhibition of photosynthesis in marine phytoplankton. In: Baker NR and Bowyer JR (eds) Photoinhibition of Photosynthesis, pp 407–432. BIOS Scientific Publishers, Oxford Fisher T, Shurtz-Swirski R, Gepstein S and Dubinsky Z (1989) Changes in the levels of Ribulose-1,5-bisphosphate Carboxylase/Oxygenase (Rubisco) in Tetraedron minimum (Chlorophyta) during light and shade adaption. Plant Cell Physiol 30: 221–228 Fujita Y, Murakami A and Ohki K (1987) Regulation of photosystem composition in the cyanobacterial photosynthetic system: The regulation occurs in response to the redox state of the electron pool located between the two photosystems. Plant Cell Physiol 28(2): 283–292 Furuya M and Schäfer E (1996) Photoperception and signalling of induction reactions by different phytochromes. Trends Plant Sci 1: 301–307 Grossman AR, Schaefer MR, Chiang GG and Collier JL (1993) The phycobilisome, a light-harvesting complex responsive to environmental conditions. Microbiol Rev 57(3): 725–749 Hermsmeier D, Mala E, Schulz R, Thielmann J, Galland P and Senger H (1991) Antagonistic blue- and red-light regulation of cab-gene expression durning photosynthetic adaptation in Scenedesmus obliquus. J Photochem Photobiol B: Biol. 11: 189– 202 Hughes J, Lamparter T, Mittmann F, Hartmann E, Gärtner W, Wilde A and Börner (1997) A prokaryotic phytochrome. Science 386: 663 Humbeck K, Hoffmann B and Senger H (1988) Influence of energy flux and quality of light on the molecular organization of the photosynthetic apparatus in Scenedesmus. Planta 173: 205–212 Huppe HC and Turpin DH (1994) Integration of carbon and nitrogen metabolism in plant and algal cells. Annu Rev Plant Physiol Plant Mol Biol 45: 577–607 Jang JC and Sheen J (1994) Sugar sensing in higher plants. Plant Cell 6: 1665–1679 Jasper F, Quednau B, Kortenjann M and Johanningmeier (1991) Control of cab gene expression in synchronized Chlamydomonas reinhardtii cells. J Photochem Photobiol B: Biol 11: 139–150 Johanningmeier U and Howell SH (1984) Regulation of lightharvesting chlorophyll-binding protein mRNA accumulation in Chlamydomonas reinhardtii. J Biol Chem 259: 13541–13549 Kehoe DM and Grossman AR (1996) Similarity of a chromatic adaptation sensor to phytochrome and ethylene receptors. Science 273: 1409–1412 Kilb B, Wietoska H and Godde D (1996) Changes in the expression of photosynthetic genes precede loss of photosynthetic activi- pres712b.tex; 4/11/1997; 11:22; v.7; p.11 240 ties and chlorophyll when glucose is supplied to mature spinach leaves. Plant Sci 115: 225–235 Kim JE, Glick RE and Melis A (1993) Dynamics of photosystem stoichiometry adjustment by light quality in chloroplasts. Plant Physiol 102: 181–190 Kindle KL (1987) Expression of a gene for a light-harvesting chlorophyll a/b-binding protein in Chlamydomonas reinhardtii: Effects of light and acetate. Plant Mol Biol 9: 547–563 Kirk, JTO (1994) Light and Photosynthesis in Aquatic Ecosystems. Cambridge University Press, Cambridge Koenig F (1990) Shade adaptation in cyanobacteria. Photosynth Res 26: 29–37 Koch KE (1996) Carbohydrate-modulated gene expression in plants. Annu Rev Plant Physiol Plant Mol Biol 47: 509–540 Krapp A, Hofmann B, Schaefer C and Stitt M (1993) Regulation of the expression of rbcS and other photosynthetic genes by carbohydrates: a mechanism for the ‘sink regulation’ of photosynthesis? Plant J 3: 817–828 LaRoche J, Mortain-Bertrand A and PG Falkowski PG (1991) Light intensity-induced changes in cab mRNA and light harvesting complex II apoprotein levels in the unicellular chlorophyte Dunaliella tertiolecta. Plant Physiol 97: 147–153 Levasseur ME, Morissette JC, Popovic R and Harrison PJ (1990) Effects of long term exposure to low temperature on the photosynthetic apparatus of Dunaliella tertiolecta (Chlorophyceae). J Phycol 26: 479–484 Levings III CS and Siedow JN (1995) Regulation by redox poise in chloroplasts. Science 268: 695–696 Ley AC and Mauzerall DC (1982) Absolute absorption cross sections for Photosystem II and the minimum quantum requirement for photosynthesis in Chlorella vulgaris. Biochim Biophys Acta 680: 95–106 Machalek KM, Davison IR and Falkowski PG (1996) Thermal acclimation and photoacclimation of photosynthesis in the brown alga Laminaria saccharina. Plant Cell Environ 19: 1005–1016 Matters GL and Beale SI (1994) Structure and light-regulated expression of the gsa gene encoding the chlorophyll biosynthetic enzyme, glutamate 1-semialdehyde aminotransferase, in Chlamydomonas reinhardtii. Plant Mol Biol 24: 617–629 Matters GL and Beale SI (1995) Blue-light-regulated expression of genes for two early steps of chlorophyll biosynthesis in Chlamydomonas reinhardtii. Plant Physiol 109: 471–479 Maxwell DP, Falk S, Trick CG and Huner NPA (1994) Growth at low temperature mimics high-light acclimation in Chlorella vulgaris. Plant Physiol 105: 535–543 Maxwell DP, Laudenbach DE and Huner NPA (1995) Redox regulation of light-harvesting complex II and cab mRNA abundance in Dunaliella salina. Plant Physiol 109: 787–795 Mayfield SP and Taylor WC (1984) Carotenoid-deficient maize seedlings fail to accumulate light-harvesting chlorophyll a/b binding protein (LHCP) mRNA. Eur J Biochem 144: 79–84 Melis A, Murakami A, Nemson JA, Aizawa K, Ohki K and Fujita Y (1996) Chromatic regulation in Chlamydomonas reinhardtii alters photosystem stoichiometry and improves quantum effciency of photosynthesis. Photosynth Res 47: 253–265 Michel H, Tellenbach M and Boschetti A (1983) A chlorophyll b-less mutant of Chlamydomonas reinhardtii lacking in the lightharvesting chlorophyll a/b-protein complex but not in its apoproteins. Biochim Biophys Acta 725: 417–424 Morand LZ, Kidd DG and Lagarias JC (1993) Phytochrome levels in the green alga Mesotaenium caldariorum are light regulated. Plant Physiol 101: 97–103 Morris I and Glover HE (1974) Questions on the mechanism of temperature adaptation in marine phytoplankton. Mar Biol 24: 147–154 Mortain-Bertrand A, Bennett J and Falkowski PG (1990) Photoregulation of the light-harvesting chlorophyll protein complex associated with photosystem II in Dunaliella tertiolecta. Plant Physiol 94: 304–311 Mortain-Bertrand A, Descolas-Gros C and Jupin H (1988) Growth, photosynthesis and carbon metabolism in the temperature marine diatom Skeletonema costatum adapted to low temperature and low photon-flux density. Mar Biol 100: 135–141 Munzner P and Viogt J (1992) Blue light regulation of cell division in Chlamydomonas reinhardtii. Plant Physiol 99: 1370–1375 Murakami A and Fujita Y (1991a) Steady state of photosynthetic electron transport in cells of the cyanophyte Synechocystis PCC 6714 having different stoichiometry between PS I and PS II: Analysis of flash-induced oxidation-reduction of cytochrome f and P700 under steady state of photosynthesis. Plant Cell Physiol 32(2): 213–222 Murakami A and Fujita Y (1991b) Regulation of photosystem stoichiometry in the photosynthetic system of the cyanophyte Synechocystis PCC 6714 in response to light intensity. Plant Cell Physiol 32(2): 223–230 Murakami A and Fujita Y (1993) Regulation of stoichiometry between PSI and PSII in response to light regime for photosynthesis observed with Synechocystis PCC 6741: Relationship between redox state of the Cyt b6 f complex and the regulation of PSI formation. Plant Cell Physiol 34(8): 1175–1180 Nauš J and Melis A (1992) Response of the photosynthetic apparatus in Dunaliella salina to sublethal concentrations of the herbicide 3(30 ,40 -dichlorophenyl)-1,1-dimethyl urea. Photosynthetica 26(1): 67–78 Nisho J, Sun J and Vogelmann TC (1994) Photoinhibition and the light environment within leaves In: Baker NR and Bowyer JR (eds) Photoinhibition of Photosynthesis, pp 407-432. BIOS Scientific Publishers, Oxford Oelmüller R (1989) Photooxidative destruction of chloroplasts and its effect on nuclear gene expression and extraplastidic enzyme levels. Photochem Photobiol 49(2): 229–239 Oelmüller R and Mohr H (1986) Photooxidative destruction of chloroplasts and its consequences for expression of nuclear genes. Planta 167: 106–113 Plumley FG and Schmidt GW (1989) Nitrogen-dependent regulation of photosynthetic gene expression. Proc Natl Acad Sci USA 86: 2678–2682 Post AF, Dubinsky Z, Wyman K and Falkowski PG (1984) Kinetics of light-intensity adaptation in a marine planktonic diatom. Mar Biol 83, 231–238 Reinbothe S and Reinbothe C (1996) The regulation of enzymes involved in chlorophyll biosynthesis. Eur J Biochem 237: 323– 343 Reinbothe S, Reinbothe C, Lebedev N and Apel K (1996) PORA and PORB, two light-dependent protochlorophyllide-reducing enzymes of angiosperm chlorophyll biosynthesis. Plant Cell 8: 763–769 Sheen J (1990) Metabolic repression of transcription in higher plants. Plant Cell 2: 1027–1038 Sheen J (1994) Feedback control of gene expression. Photosynth Res 39: 427–438 Short TW and Briggs WR (1994) The transduction of blue light signals in higher plants. Annu Rev Plant Physiol Plant Mol Biol 45: 143–171 Smith BM, Morrissey PJ, Guenther JE, Nemson JA, Harrison MA, Allen JF and Melis A (1990) Response of the photosynthetic pres712b.tex; 4/11/1997; 11:22; v.7; p.12 241 apparatus in Dunaliella salina (green algae) to irradiance stress. Plant Physiol 93: 1433–1440 Smith H (1995) Physiological and ecological function within the phytochrome family. Annu Rev Plant Physiol Plant Mol Biol 46: 289–315 Sukenik A, Bennett J and Falkowski PG (1987a) Light-saturated photosynthesis limitation by electron transport or carbon fixation? Biochim Biophys Acta 891: 205–215 Sukenik A, Bennett J, Mortain-Bertrand A and Falkowski PG (1990) Adaptation of the photosynthetic apparatus to irradiance in Dunaliella tertiolecta. Plant Physiol 92: 891–898 Sukenik A, Wyman KD, Bennett J and Falkowski PG (1987b) A novel mechanism for regulating the excitation of Photosystem II in a green alga. Nature 327: 704–707 Susek RE, Ausubel FM and Chory J (1993) Signal transduction mutants of Arabidopsis uncouple nuclear Cab and RbcS gene expression from chloroplast development. Cell 74: 787–799 Taylor WC (1989) Regulatory interactions between nuclear and plastid genomes. Ann Rev Plant Physiol Plant Mol Biol 40: 211–233 Terzaghi WB and Cashmore AR (1995) Light-regulated transcription. Annu Rev Plant Physiol Plant Mol Biol 46: 445–474 Thompson PA, Guo MX and Harrison PJ (1992) Effects on variation in temperature on the biochemical composition of eight species of marine phytoplankton. J Phycol 28: 481–488 Trebst A (1980) Inhibitors in electron flow: tools for the functional and structural localization of carriers and energy conservation sites. Meth Enzymol 69: 675-715 Turpin DH (1991) Effects of inorganic N availability on algal photosynthesis and carbon metabolism. J Phycol 27: 14–20 Turpin DH and Bruce D (1990) Regulation of photosynthetic light harvesting by nitrogen assimilation in the green alga Selenastrum minutum. FEBS Lett 263: 99–103 pres712b.tex; 4/11/1997; 11:22; v.7; p.13