Variability of leaf photosynthetic characteristics in rice and its relationship with resistance to water stress under different nitrogen nutrition regimes

The negative effects of water stress on rice can be alleviated by NH₄⁺ nutrition. However, the effects of mixed nitrogen (N) nutrition (NO₃^? + NH₄⁺) on resistance to water stress are still not well known. To investigate the response of rice growth to water stress and its relationship with photosynthetic characteristics, a hydroponic experiment supplying different N forms was conducted. Compared with NO₃^? nutrition, mixed‐N and NH₄⁺ nutrition greatly alleviated the reduction of leaf area, chlorophyll content, and photosynthesis under water stress, whilst subsequently maintaining higher biomass. In contrast, water stress inhibited the root‐shoot ratios in NH₄⁺‐ and mixed‐N‐supplied plants, indicating reduced root growth and higher photosynthate availability to shoots. The following key observations were made: (1) a similar stomatal limitation and low proportion of activated Rubisco were observed among the three different N nutrition regimes; (2) increased mesophyll conductance in NH₄⁺‐ and mixed‐N‐supplied plants simultaneously stimulated leaf photosynthesis and improved the water use efficiency and (3), the maximum carboxylation rate and actual photochemical efficiency of photosystem II in NH₄⁺‐ and mixed‐N‐supplied plants were significantly higher than that in NO₃^?‐supplied plants, thus resulting in higher photochemical efficiency under water stress. In conclusion, mixed‐N and NH₄⁺ nutrition may be used to develop strategies for improved water stress resistance and stimulated biomass production under conditions of osmotic stress and possibly drought.     

Photosynthetic limitations in olive cultivars with different sensitivity to salt stress

Olive (Olea europea L) is one of the most valuable and widespread fruit trees in the Mediterranean area. To breed olive for resistance to salinity, an environmental constraint typical of the Mediterranean, is an important goal. The photosynthetic limitations associated with salt stress caused by irrigation with saline (200 mm ) water were assessed with simultaneous gas‐exchange and fluorescence field measurements in six olive cultivars. Cultivars were found to possess inherently different photosynthesis when non‐stressed. When exposed to salt stress, cultivars with inherently high photosynthesis showed the highest photosynthetic reductions. There was no relationship between salt accumulation and photosynthesis reduction in either young or old leaves. Thus photosynthetic sensitivity to salt did not depend on salt exclusion or compartmentalization in the old leaves of the olive cultivars investigated. Salt reduced the photochemical efficiency, but this reduction was also not associated with photosynthesis reduction. Salt caused a reduction of stomatal and mesophyll conductance, especially in cultivars with inherently high photosynthesis. Mesophyll conductance was generally strongly associated with photosynthesis, but not in salt‐stressed leaves with a mesophyll conductance higher than 50 mmol m^?2 s^?1. The combined reduction of stomatal and mesophyll conductances in salt‐stressed leaves increased the CO₂ draw‐down between ambient air and the chloroplasts. The CO₂ draw‐down was strongly associated with photosynthesis reduction of salt‐stressed leaves but also with the variable photosynthesis of controls. The relationship between photosynthesis and CO₂ draw‐down remained unchanged in most of the cultivars, suggesting no or small changes in Rubisco activity of salt‐stressed leaves. The present results indicate that the low chloroplast CO₂ concentration set by both low stomatal and mesophyll conductances were the main limitations of photosynthesis in salt‐stressed olive as well as in cultivars with inherently low photosynthesis. It is consequently suggested that, independently of the apparent sensitivity of photosynthesis to salt, this effect may be relieved if conductances to CO₂ diffusion are restored.     

Leaf chloroplast ultrastructure and photosynthetic properties of a chlorophyll-deficient mutant of rice

Leaf chloroplast ultrastructure and photosynthetic properties of a natural, yellow-green leaf mutant (ygl1) of rice were characterized. Our results showed that chloroplast development was significantly delayed in the mutant leaves compared with the wild-type rice (WT). As leaves matured, more grana stacks formed concurrently with increasing leaf chlorophyll (Chl) content. Except for the lower intercellular CO₂ concentration, the ygl1 plants had a higher leaf net photosynthetic rate, stomatal conductance, and transpiration rate than those of the WT plants. Under equal amounts of Chl, the excitation energy of PSI and PSII was much stronger in the mutant than that in the WT. The ygl1 plants showed higher nonphotochemical quenching and lower photochemical quenching. They also exhibited higher actual photochemical efficiency of PSII with a higher electron transport rate. Under the light of 200 μmol(photon) m^?2 s^?1, the ygl1 mutant showed lesser deepoxidation of violaxanthin in the xanthophyll cycle than WT, but it increased substantially under strong light conditions. In conclusion, the photosynthetic machinery of the ygl1 remained stable during leaf development. The plants were less sensitive to photoinhibition compared with WT due to the active xanthophyll cycle. The ygl1 plants were efficient in both light harvesting and conversion of solar energy.     

Diffusional conductance to CO2 is the key limitation to photosynthesis in salt‐stressed leaves of rice (Oryza sativa)

Salinity significantly limits leaf photosynthesis but the factors causing the limitation in salt‐stressed leaves remain unclear. In the present work, photosynthetic and biochemical traits were investigated in four rice genotypes under two NaCl concentration (0 and 150 mM) to assess the stomatal, mesophyll and biochemical contributions to reduced photosynthetic rate (A) in salt‐stressed leaves. Our results indicated that salinity led to a decrease in A, leaf osmotic potential, electron transport rate and CO₂ concentrations in the chloroplasts (C_c) of rice leaves. Decreased A in salt‐stressed leaves was mainly attributable to low C_c, which was determined by stomatal and mesophyll conductance. The increased stomatal limitation was mainly related to the low leaf osmotic potential caused by soil salinity. However, the increased mesophyll limitation in salt‐stressed leaves was related to both osmotic stress and ion stress. These findings highlight the importance of considering mesophyll conductance when developing salinity‐tolerant rice cultivars.     

Gas exchange responses of Chesapeake Bay tidal marsh species under field and laboratory conditions

Summary Laboratory and field gas exchange measurements were made on C₃ (Scirpus olneyi Gray) and C₄ (Spartina patens (Ait.) Mahl., Distichlis spicata (L.) Green) species from an irregularly flooded tidal marsh on the Chesapeake Bay. Laboratory measurements were made on plants grown from root stocks that were transplanted to a greenhouse and grown under high light and high nutrient conditions. The two C₄ species were similar in their laboratory gas exchange characteristics: both had higher net carbon exchange rates, higher mesophyll conductances, higher photosynthetic temperature optima and lower leaf conductances than the C₃ species. The laboratory photosynthetic water use efficiency of the C₄ species was approximately three times that of the C₃ species.Field gas exchange responses of the above species were measured in situ a Chesapeake Bay tidal marsh. Despite differences in biological potential measured in the laboratory, all three species had similar in situ carbon exchange rates on a leaf area basis. On a dry weight basis, leaves of the two C₄ species had about 1.4 times higher light saturated CO₂ assimilation rates than the C₃ species. Light saturation of CO₂ exchange occurred at photosynthetic photon flux densities of 80 n Einstein cm^-2s^-1, compared with 160 n Einstein cm ^-2s^-1 in the laboratory grown plants. Spartina patens and Scirpus olneyi had similar daily CO₂ assimilation rates, but the daily transpiration rate of the C₃ species was almost twice that of the C₄ species. Spartina patens showed greater seasonal decrease in photosynthesis than Distichlis spicata and Scirpus olneyi. The two C₄ grass species maintained higher mesophyll conductances and photosynthetic water use efficiencies than the C₄ sedge.     

Photosynthetic response of Tempranillo grapevine to climate change scenarios

Photosynthesis in C₃ plants is CO₂ limited and therefore any increase in Rubisco carboxylation substrate may increase net CO₂ fixation, unless plants experience acclimation or other limitations. These aspects are largely unexplored in grapevine. Photosynthesis analysis was used to assess the stomatal, mesophyll, photochemical and biochemical contributions to the decreasing photosynthesis observed in Tempranillo grapevines (Vitis vinifera) from veraison to ripeness, modulated by CO₂, temperature and water availability. Photosynthesis and photosystem II photochemistry decreased from veraison to ripeness. The elevated CO₂ and temperature increased photosynthesis, but transiently, in both well irrigated (WI) and water‐stressed plants. Photosynthetic rates were maxima 1 week after the start of elevated CO₂ and temperature treatments, but differences with treatments of ambient conditions disappeared with time. There were not marked changes in leaf water status, leaf chlorophyll or leaf protein that could limit photosynthesis at ripeness. Leaf total soluble sugars remained at ripeness as high as 2 weeks after the start of treatments. On the other hand, and as expected, CO₂ diffusional limitations impaired photosynthesis in grapevine plants grown under water scarcity, stomatal and mesophyll conductances to CO₂ decreased and in turn low chloroplastic CO₂ concentrations limited photosynthetic CO₂ fixation. In summary, photochemistry and photosynthesis from veraison to ripeness in Tempranillo grapevine were dominated by a developmental‐related decreasing trend that was only transiently influenced by elevated CO₂ concentrations.     

Diffusional limitations explain the lower photosynthetic capacity of ferns as compared with angiosperms in a common garden study

Ferns are thought to have lower photosynthetic rates than angiosperms and they lack fine stomatal regulation. However, no study has directly compared photosynthesis in plants of both groups grown under optimal conditions in a common environment. We present a common garden comparison of seven angiosperms and seven ferns paired by habitat preference, with the aims of (1) confirming that ferns do have lower photosynthesis capacity than angiosperms and quantifying these differences; (2) determining the importance of diffusional versus biochemical limitations; and (3) analysing the potential implication of leaf anatomical traits in setting the photosynthesis capacity in both groups. On average, the photosynthetic rate of ferns was about half that of angiosperms, and they exhibited lower stomatal and mesophyll conductance to CO₂ (g_m), maximum velocity of carboxylation and electron transport rate. A quantitative limitation analysis revealed that stomatal and mesophyll conductances were co‐responsible for the lower photosynthesis of ferns as compared with angiosperms. However, g_m alone was the most constraining factor for photosynthesis in ferns. Consistently, leaf anatomy showed important differences between angiosperms and ferns, especially in cell wall thickness and the surface of chloroplasts exposed to intercellular air spaces.     

An Arabidopsis mutant of inositol pentakisphosphate 2‐kinase AtIPK1 displays reduced arsenate tolerance

Arsenate [As(V)] toxicity is considered to be derived from similarities in the chemical properties of As(V) and phosphate (Pi). An Arabidopsis thaliana mutant of inositol pentakisphosphate 2‐kinase (AtIPK1), atipk1‐1, has previously exhibited lower level of phytate and higher level of Pi, relative to wild‐type (WT). Here, atipk1‐1 displayed hypersensitivity to As(V) stress and less As(V) uptake when compared to WT. Overexpression of AtIPK1 controlled by the CaMV 35S promoter partially rescued the As(V)‐sensitive phenotype of atipk1‐1. When compared to control Pi status, addition of Pi enhanced As(V) tolerance of both WT and atipk1‐1 plants, while the arsenic concentration was less reduced in the latter genotype. Despite the higher Pi level in atipk1‐1 than did WT plants, the mutant suffered more severe Pi starvation under Pi limitation stress, indicating that Pi homeostasis was altered in the mutant. Gene expression analysis of WT and atipk1‐1 plants showed the diverse effect of As(V) stress on Pi starvation‐dependent regulation of Pi‐responsive genes. Our study suggested that a particular mechanism of As(V) toxicity existed in atipk1‐1 mutant, and may offer new insights into the interactions between Pi homeostasis and As(V) detoxification in plants.     

Effects of weather during leaf development on photosynthetic characteristics of soybean leaves

Net photosynthetic rates and mesophyll conductances at 25 °C at light saturation and air levels of carbon dioxide and oxygen were measured on recently fully expanded leaflets of second trifoliolate leaves of soybeans (Glycine max cv. Kent). Plants were grown outdoors in pots at Beltsville, Maryland with 14 planting times from May through August, 1983. Air temperature and humidity, and photosynthetically active radiation (PAR) were measured for the expansion periods of the second trifoliolate leaves. Rates of net photosynthesis ranged from 24 to 33 mol m^–2 s^–1, and mesophyll conductances from 0.24 to 0.35 cm s^–1 for the different planting dates. Mean 24-h air temperatures ranged from 20.6 to 29.0 °C, and mean daily PAR ranged from 29.4 to 58.4 mol m^–2 d^–1 for the leaf expansion periods. There was a positive relationship between photosynthetic characteristics and PAR during leaf expansion, and a negative relationship between photosynthetic characteristics and leaf expansion rates, with 96% of the variation in photosynthetic characteristics accounted for by these two variables. Leaf expansion rates were highly correlated with air temperature.     

Chloride as a macronutrient increases water‐use efficiency by anatomically driven reduced stomatal conductance and increased mesophyll diffusion to CO2

Chloride (Cl^?) has been recently described as a beneficial macronutrient, playing specific roles in promoting plant growth and water‐use efficiency (WUE). However, it is still unclear how Cl^? could be beneficial, especially in comparison with nitrate (NO₃^?), an essential source of nitrogen that shares with Cl^? similar physical and osmotic properties, as well as common transport mechanisms. In tobacco plants, macronutrient levels of Cl^? specifically reduce stomatal conductance (g_s) without a concomitant reduction in the net photosynthesis rate (A_N). As stomata‐mediated water loss through transpiration is inherent in the need of C₃ plants to capture CO₂, simultaneous increase in photosynthesis and WUE is of great relevance to achieve a sustainable increase in C₃ crop productivity. Our results showed that Cl^?‐mediated stimulation of larger leaf cells leads to a reduction in stomatal density, which in turn reduces g_s and water consumption. Conversely, Cl^? improves mesophyll diffusion conductance to CO₂ (g_m) and photosynthetic performance due to a higher surface area of chloroplasts exposed to the intercellular airspace of mesophyll cells, possibly as a consequence of the stimulation of chloroplast biogenesis. A key finding of this study is the simultaneous improvement of A_N and WUE due to macronutrient Cl^? nutrition. This work identifies relevant and specific functions in which Cl^? participates as a beneficial macronutrient for higher plants, uncovering a sustainable approach to improve crop yield.     

Silicon partially preserves the photosynthetic performance of rice plants infected by Monographella albescens

Leaf scald, caused by Monographella albescens, is one of the major diseases in rice worldwide. This study investigated the effect of silicon (Si) on the photosynthetic gas exchange parameters [net CO₂ assimilation rate (A), stomatal conductance to water vapour (g_s), transpiration rate (E)] and internal CO₂ concentration (C_i), chlorophyll (Chl) fluorescence a parameters [minimal fluorescence (F₀), maximum fluorescence (F_m), maximum quantum yield of photosystem II (F_v/F_m)], photochemical quenching coefficient (q_p), effective quantum yield of PSII [Y(II)], quantum yield of regulated energy dissipation [Y(NPQ)] and quantum yield dissipation non‐regulated [Y(NO)] and the concentrations of pigments in rice plants grown in nutrient solutions containing either 0 (?Si) or 2 mM Si (+Si) and non‐inoculated or inoculated with M. albescens. Leaf scald severity decreased with higher foliar Si concentration. For the inoculated +Si plants, A, g_s and E were significantly higher in comparison with the inoculated ?Si plants, in which C_i was significantly increased. Similarly, the concentrations of Chl_a, Chl_b, total Chl_a+b and carotenoids were higher for the +Si plants in comparison with the ?Si plants. Changes in the images of Chl a fluorescence were first observed precisely on the ?Si plants leaves in comparison with the +Si plants. A decrease of q_P and Y(II) in inoculated ?Si plants, in comparison with the inoculated +Si plants, was accompanied by an increase in Y(NPQ) and Y(NO). Notably, the extent of the leaf areas was much more evident for Y(II) and q_P in comparison with F₀, F_m and F_v/F_m, suggesting that Y(II) and q_P were good predictors in detecting the early effects of leaf scald on the leaf photosynthesis. For the +Si non‐inoculated plants, changes in Y(II) were associated with alterations in both Y(NPQ) and Y(NO) compared with non‐inoculated ?Si plants. In conclusion, the photosynthetic performance (as demonstrated by the gas exchange and Chl a fluorescence parameters) and the pigment pools of rice plants infected with M. albescens were preserved by Si supply and, therefore, provided an increase in rice resistance against leaf scald.     

Bioimaging of multiple elements by high‐resolution LA‐ICP‐MS reveals altered distribution of mineral elements in the nodes of rice mutants

Inter‐vascular transfer in rice (Oryza sativa) nodes is required for delivering mineral elements to developing tissues, which is mediated by various transporters in the nodes. However, the effect of these transporters on distribution of mineral elements in the nodes at a cellular level is still unknown. Here, we established a protocol for bioimaging of multiple elements at a cellular level in rice node by laser ablation‐inductively coupled plasma‐mass spectrometry (LA‐ICP‐MS), and compared the mineral distribution profile between wild‐type (WT) rice and mutants. Both relative comparison of mineral distribution normalized by endogenous ¹³C and quantitative analysis using spiked standards combined with soft ablation gave valid results. Overall, macro‐nutrients such as K and Mg were accumulated more in the phloem region, while micro‐nutrients such as Fe and Zn were highly accumulated at the inter‐vascular tissues of the node. In mutants of nodal Zn transporter OsHMA2, Zn localization pattern in the node tissues did not differ from that of WT; however, Zn accumulation in the inter‐vascular tissues was lower in uppermost node I but higher in the third upper node III compared with the WT. In contrast, Si deposition in the mutants of three nodal Si transporters Lsi2, Lsi3 and Lsi6 showed different patterns, which are consistent with the localization of these transporters. This improved LA‐ICP‐MS analysis combined with functional characterization of transporters will provide further insight into mineral element distribution mechanisms in rice and other plant species.     

Depletion of leaf-type ferredoxin-NADP(+) oxidoreductase results in the permanent induction of photoprotective mechanisms in Arabidopsis chloroplasts

Arabidopsis thaliana contains two photosynthetically competent chloroplast‐targeted ferredoxin‐NADP⁺ oxidoreductase (FNR) isoforms that are largely redundant in their function. Nevertheless, the FNR isoforms also display distinct molecular phenotypes, as only the FNR1 is able to directly bind to the thylakoid membrane. We report the consequences of depletion of FNR in the F₁ (fnr1 × fnr2) and F₂ (fnr1 fnr2) generation plants of the fnr1 and fnr2 single mutant crossings. The fnr1 × fnr2 plants, with a decreased total content of FNR, showed a small and pale green phenotype, accompanied with a marked downregulation of photosynthetic pigment‐protein complexes. Specifically, when compared with the wild type (WT), the quantum yield of photosystem II (PSII) electron transport was lower, non‐photochemical quenching (NPQ) was higher and the rate of P700⁺ re‐reduction was faster in the mutant plants. The slight over‐reduction of the plastoquinone pool detected in the mutants resulted in the adjustment of the reactive oxygen species (ROS) scavenging systems, as both the content and de‐epoxidation state of xanthophylls, as well as the content of α‐tocopherol, were higher in the leaves of the mutant plants when compared with the WT. The fnr1 fnr2 double mutant plants, which had no detectable FNR and possessed an extremely downregulated photosynthetic machinery, survived only when grown heterotrophically in the presence of sucrose. Intriguingly, the fnr1 fnr2 plants were still capable of sustaining the biogenesis of a few malformed chloroplasts.     

Photosynthetic characteristics and tolerance to photo-oxidation of transgenic rice expressing C4 photosynthesis enzymes

The photosynthetic characteristics of four transgenic rice lines over-expressing rice NADP-malic enzyme (ME), and maize phosphoenolpyruvate carboxylase (PC), pyruvate,orthophosphate dikinase (PK), and PC+PK (CK) were investigated using outdoor-grown plants. Relative to untransformed wild-type (WT) rice, PC transgenic rice exhibited high PC activity (25-fold increase) and enhanced activity of carbonic anhydrase (more than two-fold increase), while the activity of ribulose-bisphosphate carboxylase/oxygenase (Rubisco) and its kinetic property were not significantly altered. The PC transgenic plants also showed a higher light intensity for saturation of photosynthesis, higher photosynthetic CO₂ uptake rate and carboxylation efficiency, and slightly reduced CO₂ compensation point. In addition, chlorophyll a fluorescence analysis indicates that PC transgenic plants are more tolerant to photo-oxidative stress, due to a higher capacity to quench excess light energy via photochemical and non-photochemical means. Furthermore, PC and CK transgenic rice produced 22–24% more grains than WT plants. Taken together, these results suggest that expression of maize C₄ photosynthesis enzymes in rice, a C₃ plant, can improve its photosynthetic capacity with enhanced tolerance to photo-oxidation. This revised version was published online in June 2006 with corrections to the Cover Date.     

CG hypomethylation leads to complex changes in DNA methylation and transpositional burst of diverse transposable elements in callus cultures of rice

CG methylation (^mCG) is essential for preserving genome stability in mammals, but this link remains obscure in plants. OsMET1‐2, a major rice DNA methyltransferase, plays critical roles in maintaining ^mCG in rice. Null mutation of OsMET1‐2 causes massive CG hypomethylation, rendering the mutant suitable to address the role of ^mCG in maintaining genome integrity in plants. Here, we analyzed ^mCG dynamics and genome stability in tissue cultures of OsMET1‐2 homozygous (?/?) and heterozygous (+/?) mutants, and isogenic wild‐type (WT). We found ^mCG levels in cultures of ?/? were substantially lower than in those of WT and +/?, as expected. Unexpectedly, ^mCG levels in 1‐ and 3‐year cultures of ?/? were 77.6% and 48.7% higher, respectively, than in shoot, from which the cultures were initiated, suggesting substantial regain of ^mCG in ?/? cultures, which contrasts to the general trend of ^mCG loss in all WT plant tissue cultures hitherto studied. Transpositional burst of diverse transposable elements (TEs) occurred only in ?/? cultures, although no elevation of genome‐wide mutation rate in the form of single nucleotide polymorphisms was detected. Altogether, our results establish an essential role of ^mCG in retaining TE immobility and hence genome stability in rice and likely in plants in general.     

Study on salt tolerance with YHem1 transgenic canola (Brassica napus)

5‐Aminolevulinic acid (5‐ALA) has been suggested for improving plant salt tolerance via exogenous application. In this study, we used a transgenic canola (Brassica napus), which contained a constituted gene YHem1 and biosynthesized more 5‐ALA, to study salt stress responses. In a long‐term pot experiment, the transgenic plants produced higher yield under 200 mmol L^?1 NaCl treatment than the wild type (WT). In a short‐term experiment, the YHem1 transformation accelerated endogenous 5‐ALA metabolism, leading to more chlorophyll accumulation, higher diurnal photosynthetic rates and upregulated expression of the gene encoding Rubisco small subunit. Furthermore, the activities of antioxidant enzymes, including superoxide dismutase, guaiacol peroxidase, catalase and ascorbate peroxidase, were significantly higher in the transgenic plants than the WT, while the levels of O₂·^? and malondialdehyde were lower than the latter. Additionally, the Na⁺ content was higher in the transgenic leaves than that in the WT under salinity, but K⁺ and Cl^? were significantly lower. The levels of N, P, Cu, and S in the transgenic plants were also significantly lower than those in the WT, but the Fe content was significantly improved. As the leaf Fe content was decreased by salinity, it was suggested that the stronger salt tolerance of the transgenic plants was related to the higher Fe acquisition. Lastly, YHem1 transformation improved the leaf proline content, but salinity decreased rather than increased it. The content of free amino acids and soluble sugars was similarly decreased as salinity increased, but it was higher in the transgenic plants than that in the WT.     

The promoters of two carboxylases in a C4 plant (maize) direct cell-specific, light-regulated expression in a C3 plant (rice)

C₄ plants have two carboxylases which function in photosynthesis. One, phosphoenolpyruvate carboxylase (PEPC) is localized in mesophyll cells, and the other, ribulose bisphosphate carboxylase (RuBPC) is found in bundle sheath cells. In contrast, C₃ plants have only one photosynthetic carboxylase, RuBPC, which is localized in mesophyll cells. The expression of PEPC in C₃ mesophyll cells is quite low relative to PEPC expression in C₄ mesophyll cells. Two chimeric genes have been constructed consisting of the structural gene encoding β-glucuronidase (GUS) controlled by two promoters from C₄ (maize) photosynthetic genes: (i) the PEPC gene (pepc) and (ii) the small subunit of RuBPC (rbcS). These constructs were introduced into a C₃ cereal, rice. Both chimeric genes were expressed almost exclusively in mesophyll cells in the leaf blades and leaf sheaths at high levels, and no or very little activity was observed in other cells. The expression of both genes was also regulated by light. These observations indicate that the regulation systems which direct cell-specific and light-inducible expression of pepc and rbcS in C₄ plants are also present in C₃ plants. Nevertheless, expression of endogenous pepc in C₃ plants is very low in C₃ mesophyll cells, and the cell specificity of rbcS expression in C₃ plants differs from that in C₄ plants. Rice nuclear extracts were assayed for DNA-binding protein(s) which interact with a cis-regulatory element in the pepc promoter. Gel-retardation assays indicate that a nuclear protein with similar DNA-binding specificity to a maize nuclear protein is present in rice. The possibility that differences in pepc expression in a C₃ plant (rice) and C₄ plant (maize) may be the result of changes in cis-acting elements between pepc in rice and maize is discussed. It also appears that differences in the cellular localization of rbcS expression are probably due to changes in a trans-acting factor(s) required for rbcS expression.