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J S Marshall  J D Stubbs    W C Taylor 《Plant physiology》1996,111(4):1251-1261
To gain an understanding of the molecular events underlying the evolution of C4 photosynthesis, we have undertaken as detailed study of the NADP-malic enzyme gene family in C4 and C3 species of Flaveria. Three genomic clones form the C4 species Flaveria bidentis were characterized and found to encode two highly similar chloroplastic forms of NADP-malic enzyme, termed ME1 and ME2. Genomic southern blotting with gene-specific probes showed that both Me1 and Me2 are found in Flaveria trinervia (C4) and Flaveria pringlei (C3) as well as in F. bidentis. Northern blots demonstrated that Me1 expression in leaves parallels the degree of C4 photosynthesis in seven Flaveria species. Furthermore, whereas Me2 was expressed at a low level in both roots and leaves of F. bidentis, Me1 expression was seen only in leaves and was light-regulated. We discuss these results in the context of the evolution of C4 photosynthesis in Flaveria.  相似文献   

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A cytosolic NADP-malic enzyme (CYTME) has been described previously in several plants, all C3 species. CYTME is distinct from the chloroplastic NADP-malic enzyme (CHLME) that is highly active in C4 species. We show that at least one CytMe gene is present in all Flaveria spp., including C3, C4, and C3-C4 intermediate types. Based on the CytMe expression patterns in Flaveria pringlei (C3) and Flaveria trinervia (C4), we suggest CYTME has several distinct roles, including the supplying of NADPH for cytosolic metabolism, the supporting of wound response or repair, and the balancing of cellular pH in illuminated leaves. These three roles are likely correlated with CytMe mRNAs of apparent sizes 2.0, 2.2, and 2.4 kb, respectively, which differ in the length of the 5' untranslated regions. Various regulatory mechanisms involving RNA processing and translational efficiency are discussed.  相似文献   

5.
C4 photosynthesis is characterized by a division of labour between two different photosynthetic cell types, mesophyll and bundle-sheath cells. Relying on phosphoenolpyruvate carboxylase (PEPC) as the primary carboxylase in the mesophyll cells a CO2 pump is established in C4 plants that concentrates CO2 at the site of ribulose 1,5-bisphosphate carboxylase/oxygenase in the bundle-sheath cells. The C4 photosynthetic pathway evolved polyphyletically implying that the genes encoding the C4 PEPC originated from non-photosynthetic PEPC progenitor genes that were already present in the C3 ancestral species. The dicot genus Flaveria (Asteraceae) is a unique system in which to investigate the molcular changes that had to occur in order to adapt a C3 ancestral PEPC gene to the special conditions of C4 photosynthesis. Flaveria contains not only C3 and C4 species but also a large number of C3-C4 intermediates which vary to the degree in which C4 photosynthetic traits are expressed. The C4 PEPC gene of Flaveria trinervia, which is encoded by the ppcA gene class, is highly expressed but only in mesophyll cells. The encoded PEPC protein possesses the typical kinetic and regulatory features of a C4-type PEPC. The orthologous ppcA gene of the C3 species Flaveria pringlei encodes a typical non-photosynthetic, C3-type PEPC and is weakly expressed with no apparent cell or organ specificity. PEPCs of the ppcA type have been detected also in C3-C4 intermediate Flaveria species. These orthologous PEPCs have been used to determine the molecular basis for C4 enzyme characteristics and to understand their evolution. Comparative and functional analyses of the ppcA promoters from F. trinervia and F. pringlei make it possible to identity the cis-regulatory sequences for mesophyll-specific gene expression and to search for the corresponding trans-regulatory factors.  相似文献   

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The function of the C4 mechanism of photosynthesis depends on the strict compartmentation of the enzymes involved. Here, we investigate the regulatory mechanisms that ensure the mesophyll-specific expression of the C4 isoform of phosphoenolpyruvate carboxylase. We show that 2 kb of the 5[prime] flanking region of the Flaveria trinervia C4 PpcA1 gene is sufficient to direct mesophyll-specific expression of the [beta]-glucuronidase reporter gene in transgenic F. bidentis (C4) plants. In young leaves of seedlings, the activity of this promoter is dependent on the developmental stage of the mesophyll cells. It is induced in a basipetal fashion (leaf tip to base) during leaf development. The promoter region of the orthologous nonphotosynthetic Ppc gene of F. pringlei (C3) induces reporter gene expression mainly in the vascular tissue of leaves and stems as well as in mesophyll cells of transgenic F. bidentis plants. Our experiments demonstrate that during the evolution of the C4 Flaveria species, cis-acting elements of the C4 Ppc gene must have been altered to achieve mesophyll-specific expression.  相似文献   

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To improve the efficiency of CO(2) fixation in C(3) photosynthesis, C(4)-cycle genes were overexpressed in potato and tobacco plants either individually or in combination. Overexpression of the phosphoenolpyruvate carboxylase (PEPC) gene (ppc) from Corynebacterium glutamicum (cppc) or from potato (stppc, deprived of the phosphorylation site) in potato resulted in a 3-6-fold induction of endogenous cytosolic NADP malic enzyme (ME) and an increase in the activities of NAD-ME (3-fold), NADP isocitrate dehydrogenase (ICDH), pyruvate kinase (PK), NADP glycerate-3-P dehydrogenase (NADP-GAPDH), and PEP phosphatase (PEPP). In double transformants overexpressing cppc and chloroplastic NADP-ME from Flaveria pringlei (fpMe1), cytosolic NADP-ME was less induced and pleiotropic effects were diminished. There were no changes in enzyme pattern in single fpMe1 overexpressors. In cppc overexpressors of tobacco, the increase in endogenous cytosolic NADP-ME activity was small and changes in other enzymes were less pronounced. Determinations of the CO(2) compensation point (Gamma*) as well as temperature and oxygen effects on photosynthesis produced variational data suggesting that the desired decline in photorespiration occurred only under certain experimental conditions. Double transformants of potato (cppc/fpMe1) exhibited the most consistent attenuating effect on photorespiration. In contrast, photorespiration in tobacco plants appeared to be diminished most in single cppc overexpressors rather than in double transformants (cppc/fpMe1). In tobacco, introduction of the PEP carboxykinase (PEPCK) gene from the bacterium Sinorhizobium meliloti (pck) had little effect on photosynthetic parameters in single (pck) and double transformants (cppc/pck). In transgenic potato plants, increased PEPC activities resulted in a decline in UV protectants (flavonoids) in single cppc or stppc transformants, but not in double transformants (cppc/fpMe1). PEP provision to the shikimate pathway inside the plastids, from which flavonoids derive, might be restricted only in single PEPC overexpressors.  相似文献   

11.
Stable reciprocal hybrids between Flaveria pringlei (C3) and F. brownii (C4-like) have been produced by standard breeding techniques. There are no differences in the isoelectric focusing patterns of the catalytic subunits of the ribulose-1,5-bisphosphate carboxylase/oxygenase from F. pringlei, F. brownii, or the reciprocal hybrids. The enzyme from both species also contains an identical noncatalytic subunit polypeptide. However, the carboxylase enzyme from F. brownii contains another isomeric form of noncatalytic subunit polypeptide which is resolveable by isoelectric focusing. This isomeric form constitutes about 50% of the total noncatalytic subunits in this species. It comprises only about 10% of the total noncatalytic subunit population in the C3 x C4 plants, but about 42% of the noncatalytic subunits in the reciprocal cross. The concentrations of the holoenzyme in the reciprocal hybrids are comparable to those of the respective maternal parent. We hypothesize that a differential inheritance of parental chloroplasts by the reciprocal hybrids may be associated with this apparent maternal influence on the expression of the noncatalytic polypeptides and the holoenzyme concentration.  相似文献   

12.
Limited information exists regarding molecular events that occurred during the evolution of C(4) plants from their C(3) ancestors. The enzyme β-carbonic anhydrase (CA; EC 4.2.1.1), which catalyses the reversible hydration of CO(2), is present in multiple forms in C(3) and C(4) plants, and has given insights into the molecular evolution of the C(4) pathway in the genus Flaveria. cDNAs encoding three distinct isoforms of β-CA, CA1-CA3, have been isolated and examined from Flaveria C(3) and C(4) congeners. Sequence data, expression analyses of CA orthologues, and chloroplast import assays with radiolabelled CA precursor proteins from the C(3) species F. pringlei Gandoger and the C(4) species F. bidentis (L.) Kuntze have shown that both contain chloroplastic and cytosolic forms of the enzyme, and the potential roles of these isoforms are discussed. The data also identified CA3 as the cytosolic isoform important in C(4) photosynthesis and indicate that the C(4) CA3 gene evolved as a result of gene duplication and neofunctionalization, which involved mutations in coding and non-coding regions of the ancestral C(3) CA3 gene. Comparisons of the deduced CA3 amino acid sequences from Flaveria C(3), C(4), and photosynthetic intermediate species showed that all the C(3)-C(4) intermediates investigated and F. brownii, a C(4)-like species, have a C(3)-type CA3, while F. vaginata, another C(4)-like species, contains a C(4)-type CA3. These observations correlate with the photosynthetic physiologies of the intermediates, suggesting that the molecular evolution of C(4) photosynthesis in Flaveria may have resulted from a temporally dependent, stepwise modification of protein-encoding genes and their regulatory elements.  相似文献   

13.
Evolution of C4 phosphoenolpyruvate carboxylase   总被引:8,自引:0,他引:8  
C4 plants are known to be of polyphyletic origin and to have evolved independently several times during the evolution of angiosperms. This implies that the C4 isoform of phosphoenolpyruvate carboxylase (PEPC) originated from a nonphotosynthetic PEPC gene that was already present in the C3 ancestral species. To meet the special requirements of the C4 photosynthetic pathway the expression program of the C4 PEPC gene had to be changed to achieve a strong and selective expression in leaf mesophyll cells. In addition, the altered metabolite concentrations around C4 PEPC in the mesophyll cytoplasm necessitated changes in the enzyme's kinetic and regulatory properties. To obtain insight into the evolutionary steps involved in these altered enzyme characteristics, and even the order of these steps, the dicot genus Flaveria (Asteraceae) appears to be the experimental system of choice. Flaveria contains closely related C3, C3-C4, and C4 species that can be ordered by their gradual increase in C4 photosynthetic traits. The C4 PEPC of F. trinervia, which is encoded by the ppcA gene class, possesses typical kinetic and regulatory features of a C4-type PEPC. Its nearest neighbor is the orthologous ppcA gene of the C3 species F. pringlei. This latter gene encodes a typical nonphotosynthetic C3-type PEPC which is believed to be similar to the C3 ancestral PEPC. This pair of orthologous PEPCs has been used to map C4-specific molecular determinants for the kinetic and regulatory characteristics of C4 PEPCs. The most notable finding from these investigations was the identification of a C4 PEPC invariant site-specific mutation from alanine (C3) to serine (C4) at position 774 that was a necessary and late step in the evolution of C3 to C4 PEPC. The C3-C4 intermediate ppcA PEPCs are used to identify the sequence of events leading from a C3- to a C4-type PEPC.  相似文献   

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D B?rsch  P Westhoff 《FEBS letters》1990,273(1-2):111-115
The primary structure of NADP-dependent malic enzyme (NADP-ME) of the dicotyledonous C4 plant Flaveria trinervia was determined from sequence analysis of a cDNA clone containing the complete coding region. Comparison of the mature F. trinervia NADP-ME with the maize enzyme reveals extensive sequence similarity. In contrast, no significant similarity can be detected between the putative transit peptides of the two enzymes. This suggests that the corresponding parts of the genes arose independently from each other during evolution of mono- and dicotyledonous C4 plants.  相似文献   

16.
C4 phospho enolpyruvate carboxylases (PEPCase; EC 4.1.1.3) have evolved from ancestral non-photosynthetic (C3) isoforms during the evolution of angiosperms and thereby gained distinct kinetic and regulatory properties. In order to obtain insight into this evolutionary process we have studied the C3 isoforms, ppcB and ppcC, of the C4 dicot Flaveria trinervia (Spreng.) C. Mohr and compared them with the C4 enzyme of this species, ppcA, and its orthologue in the C3 species F. pringlei Gandoger. Phylogenetic analyses indicate that the ppcB PEPCase is the closest relative of the ppcA enzyme. In addition, the presence of ppcB also in the closely related C3 species F. pringlei suggests that this gene was present already in the ancestral C3 species and consequently that ppcA has evolved by gene duplication of ppcB. Investigation of the enzymatic properties of the ppcB and ppcC enzymes showed low and similar K(0.5)-PEP values and limited activation by glucose-6-phosphate, typical of non-photosynthetic PEPCases, at pH 8.0. However, at the more physiological pH of 7.6, the ppcC enzyme displayed a substantially higher K(0.5)-PEP than the ppcB counterpart, indicating their involvement in different metabolic pathways. This indication was strengthened by malate inhibition studies in which the ppcC enzyme showed 10 times higher tolerance to the inhibitor. The ppcA enzyme was, however, by far the most tolerant enzyme towards malate. Interestingly, the increased malate tolerance was correlated with a decrease in enzyme efficiency displayed by the turnover constant k(cat). We therefore suggest that the increased malate tolerance, which is imperative for an efficient C4 cycle, is connected with a decreased enzyme efficiency that in turn is compensated by increased enzyme expression.  相似文献   

17.
Pfundel E  Nagel E  Meister A 《Plant physiology》1996,112(3):1055-1070
The chlorophyll fluorescence characteristics of mesophyll and bundle-sheath thylakoids from plant species with the C4 dicarboxylic acid pathway of photosynthesis were investigated using flow cytometry. Ten species with the NADP-malic enzyme (NADP-ME) biochemical type of C4 photosynthesis were tested: Digitaria sanguinalis (L.) Scop., Euphorbia maculata L., Portulaca grandiflora Hooker, Saccharum officinarum L., Setaria viridis (L.) Beauv., Zea mays L., and four species of the genus Flaveria. This study also included three species with NAD-ME biochemistry (Atriplex rosea L., Atriplex spongiosa F. Muell., and Portulaca oleracea L.). Two C4 species of unknown biochemical type were investigated: Cyperus papyrus L. and Atriplex tatarica L. Pure mesophyll and bundle-sheath thylakoids were prepared by flow cytometry and characterized by low-temperature fluorescence spectroscopy. In pure bundle-sheath thylakoids from many species with C4 photosynthesis of the NADP-ME type, significant amounts of photosystem II (PSII) emission can be detected by fluorescence spectroscopy. Simulation of fluorescence excitation spectra of these thylakoids showed that PSII light absorption contributes significantly to the apparent excitation spectrum of photosystem I. Model calculations indicated that the excitation energy of PSII is efficiently transferred to photosystem I in bundle-sheath thylakoids of many NADP-ME species.  相似文献   

18.
Ueno  O 《Journal of experimental botany》1998,49(327):1637-1646
Cellular localization of photosynthetic enzymes was investigated by immunogold electron microscopy for leaves of nine C4 grasses (three NADP-malic enzyme (NADP-ME)subtype species, three NAD-malic enzyme (NAD-ME) subtype species, and three phosphoenolpyruvate carboxykinase (PCK) subtype species), two C4 sedges (NADP-ME subtype species) and two C4 dicots (an NADP-ME and an NADP/NAD-ME subtype species). In leaves of all species, immunogold labelling was present for phosphoenolpyruvate carboxylase in the cytosol of the mesophyll cells (MC) and for ribulose-1,5-bisphosphate carboxylase/oxygenase in the chloroplasts of the bundle sheath cells (BSC). However, considerable specific variation was found in the intercellular patterns of labelling for pyruvate orthophosphate dikinase (PPDK). In the NADP-ME grasses, two NAD-ME grasses, and the dicots, significant labelling for PPDK was present in the both the BSC and the MC chloroplasts. In the other NAD-ME grass, the PCK grasses, and the sedges, labelling for PPDK was present almost exclusively in the chloroplasts of the MC. These patterns were observed in the leaves of both young seedlings and mature plants. These results indicate that the accumulation of PPDK in leaves of C4 plants is not necessarily restricted to the MC, although the chloroplasts of the MC accumulate more than those of the BSC.Key words: C4 plants, immunolocalization, phosphoenolpyruvate carboxylase, pyruvate orthophosphate dikinase, ribulose-1,5-bisphosphate carboxylase/oxygenase.   相似文献   

19.
B McGonigle  T Nelson 《Plant physiology》1995,108(3):1119-1126
In C4 plants of the NADP-malic enzyme type, an abundant, mesophyll cell-localized NADP-malate dehydrogenase (MDH) acts to convert oxaloacetate, the initial product of carbon fixation, to malate before it is shuttled to the bundle sheath. Since NADP-MDH has different but important roles in leaves of C3 and C4 plants, we have cloned and characterized a nearly full-length cDNA encoding NADP-MDH from Flaveria trinervia (C4) to permit comparative structure/expression studies within the genus flaveria. The dicot genus Flaveria includes C3-C4 intermediate species, as well as C3 and C4 species. We show that the previously noted differences in NADP-MDH activity levels among C3, C4, and C3-C4 Flaveria species are in part due to interspecific differences in mRNA accumulation. We also show that the NADP-MDH gene appears to be present as a single copy among different Flaveria species, suggesting that a pre-existing gene has been reregulated during the evolution from C3 to C4 plants to accommodate the abundance and localization requirements of the C4 cycle.  相似文献   

20.
In the C(4) pathway of photosynthesis two types of malate decarboxylases release CO(2) in bundle sheath cells, NADP- and NAD-dependent malic enzyme (NADP-ME and NAD-ME), located in the chloroplasts and the mitochondria of these cells, respectively. The C(4) decarboxylases involved in C(4) photosynthesis did not evolve de novo; they were recruited from existing housekeeping isoforms. NADP-ME housekeeping isoforms would function in the control of malate levels during hypoxia, pathogen defence responses, and microspore separation, while NAD-ME participates in the respiration of malate in the tricarboxylic acid cycle. Recently, the existence of three enzymatic NAD-ME entities in Arabidopsis, occurring by alternative association of two subunits, was described as a novel mechanism to regulate NAD-ME activity under changing metabolic environments. The C(4) NADP-ME is thought to have evolved from a C(3) chloroplastic ancestor, which in turn would have evolved from an ancient cytosolic enzyme. In this way, the C(4) NADP-ME would have emerged through gene duplication, acquisition of a new promoter, and neo-functionalization. In contrast, there would exist a unique NAD-ME in C(4) plants, which would have been adapted to perform a dual function through changes in the kinetic and regulatory properties of the C(3) ancestors. In addition to this, for the evolution of C(4) NAD-ME, insertion of promoters or enhancers into the single-copy genes of the C(3) ancestors would have changed the expression without gene duplication.  相似文献   

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