ACTA BIOLOGICA SLOVENICA LJUBLJANA 2003 Vol. 46, Št. 2: 49 - 54 Sprejeto (accepted): 2003-10-30 Light-modulation of bark photosynthesis in birch (Betula pendula Roth.) Christiane WITIMANN and Hardy PFANZ * Institute of Applied Botany, University of Duisburg-Essen, FRG *Corresponding address: Univ.-Prof. Dr. H. Pfanz, Institut fiir Angewandte Botanik, Universitat Essen, Universitatsstr. 5 D-45117 Essen; Tel.: 0201-183-2153; Fax: 0201-183-4219; e-mail: hardy.pfanz@uni-essen.de Introduction Abstract. Bark photosynthesis has been shown to be an effective mechanism for stem-intemal refixation of respiratory CO 2 • In young birch trees (Betula pendula Roth.) this function is clearly modulated by the prevailing light intensity regime. Although positive net photosynthesis was not found in intact birch twigs, apparent twig respiration was reduced upon illumination by 65 % in high-light grown birches and even more in shade grown trees (81 %). Compared ona unit area basis the bark chlorenchyma contained up to 55% of the chlorophyll of the concomitant !eaves when grown under 100 % sunlight and even 66 % when trees were grown under low-light (20 % of full sunlight). Light penetration through the periderm of birch twigs and branches is age-dependent and ranges in control trees from roughly 24 % of the incident sunlight in recent-year twigs to 1-3 % in 5-year-old main stems. Peridermal light transmittance was also changed by the light intensity regime. An additional light-reducing peridermal layer present in control trees was not found in shade-grown birches. It was shown that CO 2 -refixation is not limited to the light- exposed outer parts of tree crowns. Our results show that also inner branches of trees are well adapted to function as a rather efficient system to prevent respiratory carbon loss. Key words: Betula, bark photosynthesis, light intensity, carbon fluxes, CO 2 - refixation In nearly all trees, woody shrnbs and bushes chlorophyll-containing tissues can be found in the inner bark !ayers. These chlorenchymes are able to photosynthetically reduce the flux of respiratory CO 2 to the atmosphere and in parallel to evolve thylakoid-bome 0 2 (PFANZ et al. 2002, PFANZ & AscHAN 2000), a process that bas been termed „CO 2 -refixation" or alternatively „corticular photosynthesis" (SPRUGEL & BENECKE 1991, NILSEN 1995, PrLARSKI 1995). Plant productivity depends on the balance between photosynthetic carbon assimilation and the expenditure of fixed carbon by respiration (EDWARDS ET al. 1981 , W ARING & RuNNING 1998). Equi valent gains in whole plant carbon assimilation can be made by increasing the rate of carbon uptake from 50 Acta Biologica Slovenica, 46 (2), 2003 the atmosphere (i.e by the stimulation of leaf photosynthesis) or by decreasing respiratory carbon losses (i.e through bark photosynthesis; CERNUSAK & MARSHALL 2000). In different deciduous trees (e.g. Fagus sylvatica, Populus tremula) stem-intemal refixation of CO2 in young twigs and branches may compensate for 60-90 % of the potential respiratory carbon loss (Prr.ARsKI 1995, WITTMANN Er AL. 2001, AscHAN Er AL. 2001). The light-driven stem-intemal refixation of carbon dioxide may therefore be an important measure to optirnize carbon fluxes at the whole-tree (or even whole-canopy) leve!. Light is one of the major environmental factors regulating plant productivity. There is plenty of knowledge on the effects of different light on the carbon gain of Ieaves, but information on the light- dependency of bark photosynthesis or on the carbon budgets of twigs and branches is still scarce. To illuminate the influence of the light environment on the photosynthetic activity of the inner bark cells we grew young birch trees (6-year-old) in distinct light environments (20 % and 100 % of full sunlight) and studied the effectiveness and the probable adaptive light dependency of stem- intemal carbon refixation as well as related morphological and cyto-chemical parameters. Materials and Methods Plant material Six-years-old birch trees (Betula pendula L.) were grown outside in 20 l plastic containers under sufficient nutrition (Einheitserde Typ T, Balster, Germany) and water supply, realised by periodic fertilisation with Osmocote (Bayer, Germany) and daily irrigation. In early spring 1999, 20 trees were shaded with a tentlike construction, covered with a light-reducing net tissue (light permeability about 20 % of incident sunlight); the other cluster (control, 100 %) was exposed to natura! sunlight conditions. The net structure of the shading tissue as well as the open front and back of the tent enabled permanent air circulation, avoicling overheating during hot summer days. PFD transmittance The quantity and quality of light (photon flux density ; PFD) transmitted through the peridermal layers of birch twigs were studied in 2001. According to PFANZ (1999) tissues were removed with a cork borer and placed for 5 min in isotonic solution. The wet bark samples were sandwiched between two microscopy slides and tightly adjusted over an opening in a black plastic plate towards a quantum sensor (model Ll-190SA, Li-Cor) connected with a quantum radiometer (Li-250, Li-Cor, Lincoln, Neb., USA). PFD transrnission was measured by illurninating it with a 100 W quartz halogen lamp (Xenophot HLX 64625, Osram, Germany) equipped with an infrared filter (NIR filter ST 931619/ KB, Balzers, Lichtenstein) to avoid warrning the samples and producing approximately 1000 µmol m·2 s·1 (see Pfanz 1999). Relative PFD transmission was calculated as a percentage of the incident PFD directly above the sample surface. Photosynthetic pigment determination Disks from leaves or from bark were removed by a calibrated cork borer (5 mm in diameter) and placed in 80 % (v/v) dimethyl sulfoxide (DMSO). Pigment extraction required approximately 2 h at 65°C in the dark. To avoid acidification and a concomitant phaeophytinisation of the chlorophylls, 20 mg Mgz(OH)2CO3 was added . Finally, extract absorbances were measured with a spectrophotometer (UV 160, Shimadzu, Japan) and pigment contents calculated according to standard equations (Wellburn 1994). C. Wittmann and H. Pfanz: Light-modulation of bark photosynthesis in birch (Betula pendula Roth.) 51 Gas-exchange measurement Photosynthetic performance of intact twigs and leaves was studied with a portable gas-exchange system (model LI-6400, Li-Cor Inc., Lincoln, USA). To avoid possible wound respiration only intact twig internodes or single mature leaves stili attached to the twigs were selected for the gas- exchange measurements. Dark respiration was measured after a 30-min shading period of the respective plant parts within the cuvette. Subsequent light response curves were conducted under constant climatic conditions (20°C, 50-55% relative humidity) and a controHed CO 2 supply (350 ppm). High-irradiance CO2-exchange rates were first measured after a 30-rnin period oflight exposure to 2000 µmol photons m-2 s·1• At least ten independent light response curves were assessed for each type of leaf or twig intemode. Curve fittings and calculations of all parameters were done using Sigma Plot 5.0 (SPSS Science Software). Light saturation of CO2-assirnilation was defined at 90% of maximum photosynthetic rate (e.g. von WILLERT ET AL. 1995). Stem internat CO2-re-fixation rates were calculated as the difference between high-irradiance maximum CO 2 -exchange rate and dark respiration. Results and Discussion PFD transmittance Transrnittance of light (qualitatively and quantitatively) through the periderm clearly depends on thickness, cellular structure and optical properties of the outer bark layers (VoGELMANN 1993, PFANZ & AscHAN 2000). With advancing age of the twigs the thickness of the peridermal layers increases; as a result peridermal PFD transmission is reduced. Light penetration through the periderm ofbirch twigs ranged from ca. 24 % in recent-year twigs to 1-3 % in 5-year-old main stems. Growth under limiting light conditions greatly influenced peridermal PFD transmittance. A normally formed additional peridermal layer was not found under low light growth conditions. Thus, the peridermal layers of birch twigs permit a relatively higher light flux under shaded conditions. Pigment content Compared on a unit area basis the bark chlorenchyma contained up to 55 % of the chlorophyll content of concomitant leaves in high-light grown trees and even 66 % in trees grown under low- light conditions. It is well known that shade-leaves optimise the effectiveness of light absorption by an increased pigment density per unit leaf area and this strategy holds also for twigs. The chlorophyll alb ratio in birch chlorenchyma ranged between 2.1 and 2.7. These results are in good agreement with those described for other deciduous species, such as Fagus sylvatica (1.8, LARCHER ET AL. 1988), Populus tremuloides (2.7, KHAROUK ET AL. 1955), Syringa vulgaris (PILARSKI 1999), and Betula pendula (KAuPPI 1991). In general, shade adapted cells within a leafhave a lower chlorophyll alb ratio than cells adapted to high light ( e.g. LICHTENTHALER ET AL. 1981, ANDERSON & 52 Acta Biologica Slovenica, 46 (2), 2003 OsMOND 1987; THERASHIMA & HIKoSAKA 1995). The lower chlorophyll alb ratio of the bark chlorenchyma (as compared to the leaves) is easily explained by the hidden location behind the cork layers. Table 1: Cardinal points of the light response curves of Betula pendula twigs cultivated under different growth light regimes (n = 10; +/- SD) relative light during cultivation [% sunlight] 20% 100 % Age of the twig organ current-year current-year max. net photosynthesis rate [µmol C0 2 m·2 s·'] -0.27 ± 0.05 -0.98 ± 0.12 stem-internal photosynthesis 1 [µmol C0 2 m·2 s·'] 1.07 ± 0.05 1.79±0.12 saturated PFD 2 [µmol photons m·2 s·1] 250 339 dark respiration rate (µmol C0 2 m·2 s·1] -1.34 ± 0.04 -2.77 ± 0.10 Age of the twig organ 1-year-old 1-year-old max. net photosynthesis rate [µmol C0 2 m·2 s·1] -0.16 ± 0.04 -0.44 ± 0.10 stem-internal photosynthesis 1 [µmol C0 2 m·2 s·'] 0.69 ± 0.04 0.98 ± 0.10 saturated PFD 2 [µmol photons m·2 s·'] 250 334 dark respiration rate [µmol C0 2 m·2 s· 1] -0.85 ± 0.04 -1.42 ± 0.09 1 stem-internal or "hidden" photosynthesis (Wittmann et al. 2001) was calculated as the difference between high-irradiance maximum C0 2 -exchange rate and dark respiration. 2 saturated PFD [µmol photons m·2 s·1] was defined at 90 % of maximum photosynthetic rate (e.g. von Willert et al. 1995); calculations were done using Sigma Plot 5.0 (SPSS Science Software). CO2-exchange of twigs and leaves Birches revealed typical hyperbolic light response curves for both, mature leaves and intact twig segments. The measured cardinal points of the light response curves of leaf and twig photosynthesis are given in Tab. l. Besides changes in net photosynthetic rates and apparent dark respiration, distinct changes in stem-intemal photosynthesis were observed (cf. WrTIMANN ET AL. 2001). Stem-intemal CO 2 -refixation was calculated as the difference between the maximum CO 2 -fixation rate and dark respiration. Furthermore, the light saturation of twig photosynthesis clearly differed, being around 340 µmol photons m·2 s·1 in high-light-grown twigs and 250 µmol photons m·2 s·1 in the shade-grown variant (Tab.l). According to LARCHER ET AL. (1994), light saturation of deciduous shade leaves is around 200-500 µE m-2 s·1; corticular photosynthesis is thus performed by extremly shade adapted chloroplasts (as indicated above by the low chl alb ratios). This fact is even more corroborated when photosynthesis of chloroplasts of the tree's pith or wood is studied (PFANZ ET AL. 2002). C. Wittmann and H. Pfanz: Lig:ht-modulation of bark photosynthesis in birch (Betula pendula Roth.) 53 90 - 80 = ~ o 70 ...... .... ~ ...... - ~ i. 60 = .... o C. .... r,.i 50 ...... ~ ~ i. ~ ~ 40 = i. ~ ~ i. "C 30 1 M ~ o o u ~ 20 = ....... 10 o 20% sunlight 100% sunlight Preliminary calculations revealed that in birch grown under full sunlight around 65 % of the respired CO 2 can be refixed within the twigs. With 81 % corticular photosynthesis is even more pronounced under shaded conditions (Fig.!). Figure 1: CO 2 -refixation rates of Betula pendula twigs (current- and 1-year-old) cultivated under different light intensity regimes (means with SE, n = 10). Conclusions Although twig and branch photosynthesis rarely result in positive net CO 2 -fixation, quite a high portion of the respired mitochondrial carbon dioxide is immediately re-used within the tree skeleton and thus contributes substantially to the overall carbon balance and productivity of trees. In young twigs of tree crowns the respiratory CO 2 losses are more efficiently reduced than in older branch parts. Yet, CO 2 -refixation is not restricted to the light-exposed outer parts of the canopy. Our results show that the light use efficiency of inner branches is relatively higher. More morphometrical data are needed to elucidate the quantitative effect of bark photosynthesis on the C-budget of trees. Acknowledgements We would like to thank Gudrun Friesewinkel, Sabine Kuhr and Christa Kosch for technical assistance. Warm thanks also to Dipl. Umweltwiss. 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