A review on the ambit and prospects of C3 and C4 plants in Nigeria

Abdulwakeel Ayokun-nun Ajao , Oludare Oladipo Agboola , Sefiu Adekilekun Saheed

1. Department of Botany, Obafemi Awolowo University, Ile-Ife, Nigeria 220005

2. Department of Botany and Plant Biotechnology, University of Johannesburg, P. O. Box 524, Auckland Park, South Africa 2006

3. Department of Botany, University of Lagos, Akoka, Nigeria 101017

1 Introduction

1.1 Photosynthesis in plants

Photosynthesis evolved early in the history of life(Blackenship, 2010). It is the process by which the chloroplast thylakoids of the leaf and other photosynthetic structures harvest light. The resultant chemical energy adenosine triphosphate and nicotinamide adenine dinucleotide phosphate (ATP and NADPH) are used to fix atmospheric carbon dioxide CO2. In C3photosynthesis, CO2is fixed directly via ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco). In C4photosynthesis, CO2is fixed indirectly after primary fixation by phosphoenol pyruvate (PEP)carboxylase and subsequent re-release in adjacent cells not in direct communication with the atmosphere, where CO2is concentrated (Furbank, 2009).For the energy of sunlight to be stored, it must be absorbed by the pigments of the organism. Several types of pigments—such as chlorophylls, carotenoids, and phycobilins-serve this function in various photosynthetic organisms. A portion of the light energy absorbed by the pigments is eventually stored in chemical bonds. This energy conversion is a complex process involving interactions between several pigment molecules and electron-transport proteins. Collectively, these components are called a photosynthetic unit. Serving as antennae, most of the pigments collect light and channel the energy to the reaction cen-ter, where the chemical reactions leading to long-term energy storage take place (Blankenship, 1992).

Over time, photosynthetic research has enhanced our understanding of ecological phenomena and the global environment (Monsi and Saeki, 2005). Indeed,photosynthesis is now an integral component of simulation models that are used to predict the future of our planet. For a long time, improving the efficiency of photosynthesis by artificial modification of photosynthetic proteins and pathways has been considered impossible or unrealistic. Over evolutionary time, photosynthesis has become complex and tightly regulated.However, recent advances have made it possible to manipulate the process using molecular genetic engineering (Andrews and Whiney, 2003; Raines, 2006).The advancement in molecular engineering can undoubtedly have positive influences on crop productivity and yield (Parryet al., 2007) because photosynthetic rate depends on biomass accretion (Kruger and Volin, 2006).

To reawaken the interest of researchers in this line of study, we conducted a review of photosynthesis research in Nigeria, with a view for helping to utilize past work and to project future work for tackling new challenges and the modern application of photosynthesis in the areas of food productivity and climate change. This review is also intended to serve as a guide for possible research on C3and C4plants in Nigeria. Information used in this review was sourced from published research articles and books retrieved from scientific data published in reputable journals.

1.2 C3 photosynthesis

The C3photosynthetic carbon-reduction cycle was first elucidated by Calvin, Bassham, and Benson in the 1950s in a series of experiments using the green alga Chlorella and radio-labelled carbon. This cycle,frequently referred to as the Calvin cycle, uses the products of the light reactions of photosynthesis, ATP and NADPH, to fix atmospheric CO2into carbon skeletons that are used directly for starch and sucrose biosynthesis (Figure 1).

The C3photosynthetic pathway is found in plants under moist or moderate environmental conditions(Winteret al., 1976). In the pathway, CO2is fixed by Rubisco and is synthesized into carbohydrates. This pathway operates only in the mesophyll cells (Boom,2004), and the majority of plants use this photosynthetic pathway. Most photosynthetic organisms utilize this method of carbon fixation. Except for a few taxa, all trees and shrubs and most herbs are C3plants(Boom, 2004). These plants fix CO2using the enzyme Rubisco and convert it in the form of a threecarbon-atom molecule storage product, 3-phosphoglyceric acid (PGA), hence the name C3photosynthesis(Boom, 2004). In the long course of earth's history,there have been periods when conditions for C3plants were less favourable (Berner and Canfield, 1989). Although it is generally assumed that C3is the oldest photosynthetic pathway among higher plants, it is still surprising that far back in the evolutionary history of these plants there had been conditions that are considered unfavorable for the plants (Boom, 2004). This circumstances is because high O2concentrations cause photorespiration in this group of plants, which causes a considerable loss of energy for the plant(Boom, 2004).

Figure 1 C3 pathway (photosynthetic carbon reduction,or PCR, cycle). Carbon initially fixed by RUBISCO is phosphorylated and reduced by the products of the light reactions (ATP and NADPH). The reduced three-carbon sugar-phosphate (triose phosphate,TP) can be either exported from the chloroplast for sucrose synthesis via the chloroplast envelope Pi transporter (Pi TRANS) or retained for starch synthesis or recycling to ribulose bisphosphate,the CO2 acceptor for the Rubisco enzyme.(Source: Furbank and Taylor, 1995)

Molecularly, the C3expression of photosynthetic carbon-reduction-cycle genes in mature leaves is sensitive to environmental and metabolic signals,providing long-term mechanisms for the plant to regulate primary carbon fixation. High levels of glucose and sucrose have been shown to be associated with reduced levels of a number of Calvin cycle mRNAs, including those encoding the small subunit of Rubisco,sedoheptulose-1,7-bisphosphatase, and fructose-1,and 6-isphosphatase. This feedback mechanism,which again appears to act at the level of transcription, might be important for source–sink regulation in the plant (Krappet al., 1993). The signaling pathway involved in glucose repression of gene expression is not known although it has been suggested that the enzyme hexokinase might be involved. Recently, it has been shown that photosynthetic genes can respond at the transcript level to nutrient status (namely nitrogen and phosphorous levels) and this response is modulated by carbohydrate status. This process suggests that the interaction between carbohydrate status and nutrient status may function as a long-term strategy in the control of primary carbon metabolism.

1.3 C4 photosynthesis

The C4photosynthetic pathway is found in plants growing in warm, sunny, and dry conditions (Winteret al., 1976). The leaves of C4plants display Kranz anatomy, in which vascular bundles are surrounded by an outer layer of mesophyll cells and an inner layer of bundle-sheath cells (Brown, 1999). The C4pathway fixes CO2initially by phosphenol pyruvate carboxylase (PEPC) (Figure 2), which is localized in mesophyll cells forming C4acids (malate and aspartate). The C4plants are among the most productive on the planet, and this trait is associated with high rates of photosynthesis and efficient use of water and nitrogen (Brown, 1999). In a typical C4plant, the initial fixation of HCO3-by PEPC and the re-fixation of CO2by Rubisco are compartmented into different cell types although segregation can occur within individual cells (Voznesenskayaet al., 2002).

Figure 2 C4 pathway, simplified to describe only the NADP-malic enzyme type, which transports malate from the mesophyll to the bundle-sheath chloroplasts. CO2 is fixed by the enzyme PEPC to form the C4 acid oxaloacetate(OAA), which is reduced by NADPH from the light reactions to form malate (MAL), the C4, acid that is transported to the bundle-sheath cells. Malate is decarboxylated in the bundle-sheath cells, where the CO2 released is fixed by the PCR cycle in much the same way as in C3 plants.The three-carbon compound pyruvate (PYR) diffuses back to the mesophyll, where it is phosphorylated by ATP to regenerate the carbon acceptor.phosphoenolpyruvate (PEP) (Source:Furbank and Taylor, 1995)

C4photosynthesis is found in many plant species:monocots includingZea,CyperusandSacharrum;dicots such asAmaranth,Flaveria,Euphorbia,Cyperus,Cleome, andBoerhavia(J?rgensen and Ulloa,1994; Ehleringeret al., 1997; Muhaidatet al., 2007).Table 1 presents some of the Nigerian plants with C3and C4features. Such a wide and diverse distribution of the occurrence of the C4pathway indicates that it has evolved independently at many different times(Langdale and Nelson, 1992). Because all C4plants share a number of characteristics in terms of leaf anatomy, physiology, and biochemistry, they stand as excellent examples of convergent evolution. The C4pathway provides a means for maintaining photosynthetic efficiency under conditions of high temperature or water limitation, by greatly reducing or eliminating photorespiration (Ehleringeret al., 1997).Photorespiration is problematic in these situations because increasing temperatures enhance the oxygenase activity relative to the carboxylation activity of the Rubisco enzyme (Ehleringeret al., 1997). In addition,C4plants have a selective advantage under arid conditions. To minimize water loss through transpiration,plants must reduce the opening of their stomata,which leads to reduced carbon CO2uptake and reduced release of oxygen. Because PEPC can fix carbon dioxide from relatively low intracellular concentrations, C4plants show higher rates of photosynthesis than C3plants under conditions that promote high transpiration rates (Sage, 2004). C4plants also show higher photosynthetic efficiencies under conditions of light saturation, such as occurs in open plains or Savannahs (Lattanzi, 2010). It is likely that the ancestors of most contemporary C4and C3–C4intermediate plants evolved between 30 and 50 million years ago,in response to reduced carbon dioxide in the atmosphere, possibly in combination with elevated temperatures or water deficit (Ehleringeret al., 1997).

The occurrence of C3–C4indicates that multiple independent steps are required for evolution of full C4capability. In most C4plants, efficient transfer of metabolites between mesophyll and bundle-sheath cells is as a result of higher plasmodesmatal linkage between these two cells, as compared to C3species(Botha, 1992). In the mesophyll cells of C4plants,chloroplast development is reduced; while in bundle sheath cells, chloroplasts are larger and undergo additional rounds of division (Hatch, 1987). According to Kanai and Edwards (1999), the operation of the C4cycle results in the increased concentration of CO2at the active site of Rubisco in the bundle-sheath cells by suppression of the oxygenase reaction of Rubisco. C4plants have evolved independently from C3plant in different taxonomic groups (Bj?rkman, 1976), and the photosynthesis is more efficient than C3photosynthesis under some environmental conditions. C4plants fix atmospheric CO2through Phosphoenolpyruvate Carboxylase (PEPC) in the mesophyll cells to form oxaloacetate, which is converted to four-carbon decarboxylic acids (malate and aspartate) (Larcher,1995). The four-carbon acids are then translocated to bundle-sheath cells and decarboxylated at the C4position with the release of CO2and C3compound(Larcher, 1995). C4plants have been reported to show higher export rates of photosynthate than C3plants(Leonardos and Grodzinski, 2000). To explain the higher rate of translocation, some researchers have pointed to the presence of a denser vascular system in C4leaves (Crookston and Moss, 1974) and a larger cross-sectional area of phloem (Gallaheret al., 1975)to be responsible for it. The vascular bundle is composed of two kinds of conducting tissues: xylem and phloem. Thus, it appears that C4leaves have a denser hydraulic network than C3leaves. The CO2-concentrating mechanism of the C4pathway gives C4plants an efficient photosynthetic mechanism under low stomatal conductance and thus a higher water-use efficiency and photosynthetic ability under environments of low water availability than is the case for C3plants(Sage, 2004). Nevertheless, it is a generally accepted fact that C4plants evolved from C3plants; and the evolution was accompanied by modifications to anatomical and biochemical features of leaves. A change in the vein density of leaves undoubtedly occurred during the evolution from C3to C4plants (Sage, 2004;Ueno and Sentoku, 2006).

Table 1 List of plants with C3 and C4 photosynthesis in Nigeria

According to Hatch (1987), C4plants are divided into three subtypes, differing in the process of decarboxylation of C4acids; nicotinamide adenine dinucleotide phosphate-malic enzyme (NADP-ME),nicotinamide adenine dinucleotide malic enzyme(NAD-ME), and phosphoenolpyruvate carboxykinase(PCK). The difference in their biochemical function is associated with the structural features of the leaves.While the NADP-ME grasses have a bundle shealth that originated from the mestome sheath, both the NAD-ME and PCK grasses have one that originated from the parenchyma sheath (Dengler and Nelson,1999). The bundle-sheath cells of the C4subtypes also differ in the structure, intracellular position, and quantity of chloroplasts and mitochondria (Yoshimuraet al., 2004). The quantitative balance of photosynthetic tissues (Dengleret al., 1994) and organelles(Yoshimuraet al., 2004) between the mesophyll and bundle-shealth cells reflects the difference in biochemical function of the photosynthetic subtypes.

2 Methods used in determining photosynthetic pathways in plants

2.1 Photosynthetic products

Initial product labeling with14C is the only direct method for photosynthetic pathway determination.Hatchet al. (1967) reported that, in C4plants, as much as 93% of fixed radioactivity appeared in oxaloacetic, malic, and aspartic acids following exposure to14CO2for approximately one second. In contrast,early products of the C3process were 3-PGA and hexose phosphates.

2.2 Carbon-dioxide compensation and photorespiration

The carbon-dioxide compensation point (the point at which photosynthetic CO2uptake equals respiratory CO2evolution when measured in a closed chamber) is an easily quantifiable characteristic. During photosynthesis, a light-induced release of CO2can occur and is referred to as photorespiration, as contrasted with CO2released by mitochondria or dark respiration. Plants with the C4pathway have a photosynthetic CO2compensation in the range of 0～10 ppm,indicating a lack of significant net photorespiration(Downton and Tregunna, 1968). Photorespiration occurs as a normal product of the Calvin cycle within the bundle-sheath cells of C4plants. However, because the mesophyll layer surrounds the bundle sheath, the C4pathway would rapidly refix any photorespiratory CO2and prevent leakage to the atmosphere (Bowes and Ogren, 1972). A much higher CO2compensation point (37～70 ppm) is characteristic of C3plants (Black, 1971). Carbon-dioxide compensation points provide a convenient means of identifying the type of photosynthetic pathway. The low CO2compensation point of C4plants indicates an ability to utilize more external CO2, as compared to use by C3plants.

2.3 Oxygen suppression

Oxygen differentially affects CO2exchange in C3and C4plant species, primarily because of differences in photorespiration. In soybean (Glycine max), and probably other C3species as well, the total O2inhibition consists of two discernible effects (Waller and Lewis, 1979). Oxygen substitutes for CO2in the carboxylase reaction to yield P-glycolate, a C3photorespiratory intermediate. As a result of this substitution, O2competitively inhibits the carboxylase with respect to CO2(Ogren, 1976). During photorespiration, glycolate is oxidized, releasing CO2. Consequently, oxygen depletion reduces glycolate oxidation, thereby increasing photosynthetic CO2assimilation by 40% ～ 50% in species possessing the C3pathway, while having no effect on C4plants (Downes and Hesketh, 1968). The re-fixation of photorespiratory CO2allows C4plants to utilize all of the fixed CO2, thus increasing photosynthetic efficiency. Chollet (1976) postulated that the enzyme complement of the C4pathway increased CO2concentration at the site of the C3carboxylase, reducing the competitive inhibition of O2and minimizing photorespiration. The CO2concentration at the site of the C3carboxylase,coupled with a specialized leaf anatomy allowing recapture of photorespiratory CO2, was apparently responsible for the lack of photorespiration and the absence of an inhibitory effect of 21% O2on net photosynthesis in C4plants (Waller and Lewis, 1979).

2.4 Carbon-isotope discrimination

Carbon-isotope ratio (13C/12C) in plant tissue is characteristically is known to be less than that of atmospheric CO2, indicating that plants preferentially assimilate the lighter of the two isotopes (Troughtonet al., 1974). Carbon-isotope values are defined as the difference in per 13 mil (1 mile = 0.0254 mm) of the13C/12C ratio of the sample relative to a standard and reported as13C‰ (Smith and Epstein, 1971). Details of the procedure are described elsewhere (Park and Epstein, 1960). Higher plants are placed into two categories, those with low13C‰ values (-24‰ to-34‰) and those with high values (-6‰ to -19‰)(Smith and Epstein, 1971). The distinctive difference apparently results from differences in affinity of the enzyme systems of the two pathways for the two isotopes of carbon (Whelanet al., 1973). Thus, the carbon-isotope technique was cited as a reliable method of distinguishing between C3and C4plants (Bender,1971). This method was successfully used by Cornet and Bonhomme (2007) to characterize 23 genotype ofDioscorea; they indicated that the δ13C values for all yam samples studied ranged from -25.39‰ to-30.07‰, which indicated all species had a C3photosynthetic type.

2.5 Leaf anato my

Leaf anatomy provides easily distinguished differences between C3and C4plants. C4photosynthesis is characterized with leaf property called "Kranz" anatomy (after Haberlandt's description in German of a wreathlike arrangement of cells). Kranz anatomy can be described as two distinct concentric layers of chlorenchyma cells formed by a bundle sheath containing most of the chloroplasts, surrounded by an outer layer consisting of a small number of mesophyll cells (Carolinet al., 1973). The visual identific-ation of such arrangements in transverse section has been used in numerous anatomical surveys of leaves to identify the photosynthetic pathway for hundreds of species (Renvoize, 1987; Faniyanet al., 2013; Ayeniet al., 2015a; Ajaoet al., 2017). Plants with the C4photosynthetic pathway generally have well-developed parenchymatic bundle sheaths containing high concentrations of chloroplasts and starch.Bundle-sheath cells utilize the C3photosynthetic process; however, they are surrounded by mesophyll cells containing chloroplasts utilizing the C4photosynthetic process, which fix and then supply CO2for the C3pathway.

Anatomical characteristics that have been used to distinguish plant species with both C3and C4photosynthetic pathways include stomatal density (Oguroet al., 1985; Kim, 2012; Ajaoet al., 2017); interstomata distance (Oguroet al., 1985); stomatal index and stomatal size (Tayloret al., 2010; Ajaoet al., 2017);Kranz tissue (Bruhl and Wilson, 2007; Martins and Alves, 2009); interveinal distance (Nelsonet al.,2005; Soros and Dengler, 1998; Ajaoet al., 2017);leaf thickness (Nelsonet al., 2005; Marshallet al.,2007; Ajaoet al., 2017); mesophyll thickness (Nelsonet al., 2005); intercellular air spaces (Marshallet al., 2007; Ajaoet al., 2017); one-cell criterion (Soros and Dengler, 1998; Ajaoet al., 2017); and maximum lateral cell count (Soros and Dengler, 1998; Bruhl and Wilson, 2007; Ajaoet al., 2017). Leaf vascular traits such as leaf-vein density and vascular spacing are also known to be diverse across C3and C4flowering plant species (Nelson and Dengler, 1997; Ajaoet al., 2017)because of their importance in leaf physiological functions such as photosynthesis and water-use efficiency (Mckownet al., 2010). The most important among these anatomical features is the occurrence of Kranz anatomy.

A number of researchers have used anatomical characteristics to classify plants along C3and C4photosynthetic pathways. For example, Welkie and Caldwell (1970) worked on the leaf anatomy of species in some dicotyledon families, as related to the C3and C4pathways of carbon fixation. Also, Hattersleyet al.(1982) reported on leaf anatomical variations inNeurachneand its relatives, in relation to C3and C4photosynthesis. Oguroet al. (1985) compared leaves of C3and C4species ofPanicum(Poaceae), using anatomical and morphological characteristics. In addition,Dengleret al. (1994) studied the quantitative leaf anatomy of C3and C4species of Poaceae, considering the bundle sheath and mesophyll surface-area relationship. Another interesting report is that of Uenoet al.(2006), which used the criteria of differentiation of some amphibious species ofEleocharisin Cyperacea.The paper reports thatEleocharis viviparahas a unique nature that expresses C4characteristic under terrestrial conditions and C3characteristics under submerged conditions. In a similar way, Fisheret al.(1997) subjected all North American species of the halophytic genusSuaeda(Chenopodiaceae) to C3and C4photosynthetic pathways classification using their leaf anatomy, which agrees with already established classification based on morphological characters. In their report, they showed that C3species belong to the Chenopodina group, while C4species are in the Linbogermen group. Nelsonet al. (2005) studied functional leaf anatomy of some dicots in Canada, using characteristics such as leaf thickness, mesophyll thickness, and intercellular air space to compare crassulacean acid metabolism (CAM) plants with C3and C4plants. Soros and Dengler (1998) studied quantitative leaf anatomy of C3and C4Cyperaceae and comparisons with Poaceae. Huxman and Monson(2003) worked on stomatal responses of C3, C3–C4intermediates, and C4Flaveriaspecies to light and intercellular CO2concentration.

2.6 Leaf and inflorescence morphology

Studies on morphological characteristics (vegetative or floral) that could possibly be used as indicators in the classification of plant species into C3and C4photosynthetic groups are rare (Ayeniet al., 2015b).Interestingly, previous reports have suggested that some morphological characteristics are related and can be linked with the two identified photosynthetic pathways in theCyperusgenus. Muasyaet al. (2009)reported that in this genus, those belonging to the C3photosynthetic pathways tend to possess spikelets arranged in digitate clusters, while the C4counterpart spikelets are usually spicately arranged. This finding is also in line with the report of Larridonet al. (2011),that genusCyperusis most commonly divided into two main infrageneric units, determined by the state of a set of anatomical and inflorescence characteristics. Ayeniet al. (2015b) reported the first known work on the use of morphological characteristics to delimit Nigeria species ofCyperusinto C3and C4pathways.

3 An overview of C3 and C4 plants research in Nigeria

The literature survey conducted on previous work such as Gentry (1993), Jorgensen and Ulloa (1994)and Pinto-Escobar and Mora-Osejo (1966) revealed that C3and C4plants are distributed among 21 genera and 11 families in Nigeria (Table 1).

Out of the 21 genera, 3 general includingEuphorbia,CyperusandBoerhaviahave been classified into C3and C4by Faniyanet al., 2013; Ayeniet al.,2015a, b; Ajaoet al., 2016, 2017. This thus infers that research into this line of thought is still not optimally explored, despite the fact that Nigeria is endowed with vast array of C3and C4plants. Although many of the species listed in the table have been classified based on stable anatomical characters such as stomata type, trichomes, and arrangement of the cell without putting into consideration the photosynthetic pathways exhibited by the plants. All the previously reported work on C3and C4plant in the country employed the use of anatomical character to delimit the species into their respective pathways, with pioneering one being that of Faniyanet al. (2013), where they used the occurrence of Kranz structure, one cell distant count and interveinal distance to classify four species ofEuphorbia. Their study revealed thatE.hirtaandE. hyssopifoliaare C4whileE. heterophylaandE. gramineaare C3. They also found out that characters such as stomatal density, cell sizes, veinstomatal distance, inter-stomatal distance, leaf thickness, mesophyll thickness, and intercellular air spaces were not found to be useful in their classification.

Two years later, Ayeniet al. (2015a) employed leaf anatomical characters to delimit twelve species ofCyperusinto C3and C4. The study revealed the usefulness of a combination of anatomical characters,rather than isolated ones, in the grouping of plant species according to their photosynthetic pathway. The combination of characters that was reported to be useful in their study are Kranz tissue, maximum cell distant count, maximum lateral cell count, interveinal distance, and to some extent, leaf and mesophyll thickness, were also found to be useful to classifyCyperusspecies into their respective photosynthetic pathways. This study confirmed the photosynthetic pathway utilized byC. difformisandC. haspanto be C3, while that ofC. articulatus,C. compressus,C.distans,C. esculentus,C. imbricatus,C. iria,C. rotundus,C. sphacelatusandC. tenuiculmisis C4.

Interestingly, Ayeniet al. (2015b) also reported that floral morphological characteristics have a potential to be useful in the grouping ofCyperusspecies as either C3or C4species. They found that compound umbellate inflorescence, digitate spikelets, and spikelet length not over 1 cm are may be peculiar to C3species, while C4species have simple umbellate inflorescence, spicate spikelets, and spikelet length over 1 cm. The two papers demonstrated a reliable basis for assessing photosynthetic pathways of the investigatedCyperusspecies in Nigeria, using inflorescence morphology and anatomical characters.

Lastly, in continuation of our effort to classically delimit our native species along the photosynthetic lines, Ajaoet al. (2016, 2017), reported the first known photosynthetic delimitation of genusBoerhaviainto C3and C4pathways. In the study, characters such as stomata index, stomata size, inter-stomatal distance, stomatal density, interveinal distance,intercellular air spaces, leaf thickness, mesophyll thickness, Kranz tissue, one cell distant count criterion, maximum lateral cell count criterion, vein density and vein distance was useful in the grouping three speciesB. erecta,B. coccineaandB. repensinto C4plants andB. diffusainto C3plant. Also in the study, the characters including interveinal distance less than 166 μm and maximum lateral count ranging 2～6 were touted to be diagnostic for C4dicotyledons species. The submission is due to the fact that higher interveinal and maximum lateral counts has been reported for monocotyledons. To the best of our knowledge, there is a dearth of information to date on the physiological and biochemical characterization of C3and C4plants in Nigeria. Research into this direction is imperative to complement the previous studies.

In other African countries such as Ghana and South Africa, research into C3and C4plant has reached considerable level. In Ghana, awareness has been created on the need to cultivated C4plants in other to meet the demand for fossil fuel in the next few decades, as it is touted that there will be rise in fossil fuel demand and prices (Blacket al., 2012). In South Africa, Stocket al. (2004) used δ13C values of herbarium specimens of 68 southern African species from 22 genera and eight tribes to delimit the species to either the C3or C4photosynthetic pathway. Out of the 68 species studied, 28 exhibited C4photosynthesis distributed among nine genera of four tribes (Cypereae, Scirpeae, Abildgaardieae and Rhyncosporeae). Their study also revealed the absence of strong relationships in the abundance of C4plant and climatic factors such as altitude and rainfall in the country.Vogelet al. (1978) investigated the distribution of C3and C4grasses in South Africa. They found out that C3and C4grasses co-occupied areas that have minimum of 100 mm of annual rainfall and maximum of 1000 mm. Furthermore, low temperatures below 25 also favors C3grasses when compared to C4counterparts. And, studies focusing on energy dissipation of C4plants under water stress have been reported (Lal and Edwards 1996; Ripleyet al., 2007). The phylogenetic issue onAlloteropsis semialata, a South African grass with C3and C4photosynthesis has also been resolved (Ripleyet al., 2007).

In West Africa at large, the influence of C3and C4vegetation on soil organic matter dynamic in seminatural tropical ecosystems including Sahel savanna,Sudan savanna, savanna woodland and the dry forest has been investigated (Saizet al., 2015). They found out that current environmental conditions encourage the distribution of C3plants over their C4counterparts in more mesic savanna ecosystems of West Africa. In addition, δ13C was also found to be varied with soil across the ecosystem. In addition, the inter-dependence between biotic and abiotic factors undoubtedly has influence on whether soil organic matter dynamics of C3and C4derived vegetation will vary in ecosystems where both vegetation types coexist. In Southern Africa, nitrogen isotopic (δ15N)abundances on soils and C3and C4plants along land use gradients of varying aridity has been investigated(Aranibaret al., 2008). The δ15N values of soils and plants were found to be higher in the areas with significant land use intensity. However, their study also revealed C3plants significantly have higher δ15N than C4counterparts from the same area. In another study conducted by Swapet al. (2004), the mean annual precipitation and the δ15N of the C3vegetation of southern African were found to be inversely proportional whereas, on the contrary, no relationship was found between mean annual precipitation and the δ13C and δ15N signatures of C4vegetation.

4 Future research and its application in food security, global climate change, and biofuel in Nigeria

Photosynthesis is one of the main research objects in many studies on the impact of water deficit on plants (Ghannoum, 2009). Water deficit is one of the worst disasters that affect health and activities in the world. The changes in precipitation patterns and the expansion of waterlogging or of drought-affected areas are expected in the future due to global climate change (Xoconstle-Cazáreset al., 2010). The response of C3photosynthesis to water stress has been well studied (Flexaset al., 2004). In general, the literature points to the fact that C3photosynthesis is negatively affected by water stress, measured as changes in leaf-water potential or relative water content. In contrast, the response of C4photosynthesis to water stress has been less studied. In spite the fact that C4plants make a significant contribution to the global carbon budget, and C4crops, such as maize and sorghum, are pivotal to current and future global food security(Lloyd and Farquhar, 1994; Brown, 1999; Pingali,2001). The CO2-concentrating mechanism, compared to the C3photosynthetic pathway, ensures a higher carbon-assimilation rate and dry-matter production under drought conditions, when stomata closure reduces CO2supply (Larcher, 1995). Consequently,higher stomatal resistance allows a low transpiration rate and high water-use efficiency in C4plants (Zhang and Kirkham, 1995).

Breeding of drought-resistant plants can be an advantageous strategy for avoiding drought-induced damage to agriculture. A suitable alternative to ordinary crops is C4plants, such as maize, sorghum, millet,and amaranth. These plants are characterized by high productivity (Svirskis, 2009), nutritive value, and resistance to water deficit (Liu and Stutzel, 2004; Osborne and Freckleton, 2009). They dominate in arid and hot regions; approximately half of the world's grasses and only from 4%～10% of all plant species use the C4photosynthetic pathway. Nevertheless, high photosynthetic capacity and productivity of these plants determine their essential contribution to global primary production (Osborne and Freckleton, 2009).

As the world population races toward 10 billion,agricultural scientists are realizing that another "green revolution" is needed for crop yields to meet demands for food. In rice, yield potential is limited by the photosynthetic capacity of leaves that, as carbohydrate factories, are unable to fill the larger number of florets of modern rice plants. One potential solution is to introduce a higher-capacity photosynthetic mechanism like the C4pathway into rice. This is the goal of researchers in the international C4. Rice Consortium: to identify and engineer the genes necessary to install C4photosynthesis in rice (Hibberdet al.,2008). Rubisco, the primary CO2-fixing enzyme in rice, is a poor catalyst of CO2under present-day atmospheric conditions. It has a tendency of confusing its substrate CO2with the more abundant O2, as well as being a very slow catalyst of CO2, turning over only once or twice per second. Rubisco's oxygenase activity requires the re-cycling of phosphoglycolate in the photorespiratory pathway, resulting in an energy cost and loss of previously fixed CO2. Many plants have developed active CO2-concentrating mechanisms to overcome Rubisco's inefficiencies among land plants; and this has led to the development of C4photosynthesis, a biochemical CO2-concentrating mechanism. C4photosynthesis arose multiple times in the past 60 million years in warm, semi-arid regions, with early occurrences coinciding with low atmospheric CO2in the late Oligocene (Sageet al., 2011).

A C4rice is achievable with a subset of C4genes,but it will require substantial fine-tuning of biochemistry and anatomy. Particularly intriguing is the need for additional metabolite transport across membranes of organelles in C4photosynthesis (Langdale, 2011).A functional C4-concentrating mechanism in rice would allow for an approximately two-thirds reduction in Rubisco levels, relative to wild-type rice; but Rubisco would be sequestered in bundle-sheath cells and ideally have a greater catalytic turnover rate(Badgeret al., 1998). However, it is expected that another few years of research is required for optimization of the phenotype and field-testing for C4rice to become ready for cultivation in farmers' fields.

Another driver of the current C4research agenda is the global focus on biofuels. Two of the current major biofuel crops, sugarcane and maize, are both C4species. Whereas the future of sugar cane as a fuel crop is almost certain, the use of maize can be defen-ded only in a future in which lignocellulosic fermentation means that grain is not used to produce ethanol.However, another C4species may hold the key to biofuel demands, for example, in the United States of America. The perennial grassMiscanthus giganteusis capable of producing a higher biomass than maize,primarily because it can photosynthesize efficiently for a longer period during the growing season. This increased efficiency is achieved in two ways. First,Miscanthuscan photosynthesize at cooler temperatures than maize, as a consequence of cold-tolerant pyruvate phosphate dikinase (PPdK) activity (Wanget al., 2008). Second, its perennial habit makes it possible for the plant to capture more light early in the season because at that time the canopy is bigger than that of annual crops such as maize (Dohleman and Long, 2009). Current estimates suggest that 9.7 million hectares (1 hectare = 10,000 m2) ofMiscanthuswould provide enough biomass to meet the annual U.S. energy mandate (Somervilleet al.,2010). Given that long-term field trials have shown thatMiscanthusproduces high yields even on poor soils and that 14 million hectares (1 hectare =10,000 m2) of land dropped out of agricultural use in the U.S. between 1997 and 2007(http://www.ers.usda.gov/statefacts/us.htm), this C4perennial could resolve the food versus fuel dilemma in the U.S. for the foreseeable future.

This trending C4research has resulted in one of the largest consortia of plant biologists pursuing a common goal. Nigeria can also optimistically take on this challenge, anticipating that advances in our understanding of C3and C4photosynthesis will better serve humanity in years to come by rejuvenating our economy in the areas of food production and green energy.

The authors thank Dr. Sabiu Saheed for encouraging us to write this review; also, the support of Mr. Alayande Kazeem and Dr. Balogun Fatai Oladunni is greatly appreciated.

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