Journal of Experimental Botany Advance Access published May 26, 2014 Journal of Experimental Botany doi:10.1093/jxb/eru212
Research Paper
Constraints to hydraulic acclimation under reduced light in two contrasting Phaseolus vulgaris cultivars Steven L. Matzner*, David D. Rettedal, Derek A. Harmon and MacKenzie R. Beukelman Department of Biology, Augustana College, Sioux Falls, SD 57197, USA * To whom correspondence should be addressed. E-mail: [email protected]
Abstract Two cultivars of Phaseolus vulgaris L. were grown under three light levels to determine if hydraulic acclimation to light occurs in herbaceous annuals and whether intraspecific trade-offs constrain hydraulic traits. Acclimation occurred in response to reduced light and included decreased stomatal density (SD) and increased specific leaf area (SLA). Reduced light resulted in lower wood density (WD); decreased cavitation resistance, measured as the xylem pressure causing a 50 % reduction in stem conductivity (P50); and increased hydraulic capacity, measured as average leaf mass specific transpiration (E(LM)). Significant or marginally significant trade-offs between P50 and WD, WD and E(LM), and E(LM) and P50 reflected variation due to both genotype and environmental effects. A trade-off between WD and P50 within one cultivar indicated that morphological adjustment was constrained. Coordinated changes in WD, P50, and E(LM) within each cultivar in response to light were consistent with trade-offs constraining plasticity. A wateruse efficiency (WUE, measured as δ13C) versus hydraulic capacity (E(LM)) trade-off was observed within each cultivar, further indicating that hydraulic trade-offs can constrain acclimation. Larger plants had lower hydraulic capacity (E(LM)) but greater cavitation resistance, WD, and WUE. Distinct hydraulic strategies were observed with the cultivar adapted to irrigated conditions having higher stomatal conductance and stem flow rates. The cultivar adapted to rainfed conditions had higher leaf area and greater cavitation resistance. Hydraulic trade-offs were observed within the herbaceous P. vulgaris resulting from both genotype and environmental effects. Trade-offs within a cultivar reflected constraints to hydraulic acclimation in response to changing light. Key words: Cavitation, herbaceous, hydraulic conductivity, light, plasticity, trade-offs.
Introduction As evolutionary selection occurs at the level of the phenotype, both genotypic differences and acclimation in response to environmental conditions are important for an organism’s immediate survival as well as for future fitness. The ability to adjust water transport capacity or drought tolerance is particularly important as water deficit is a primary factor limiting crop yield (Boyer, 1982). With the threat of climate change and rising demands for scarce water resources it is increasingly important to understand both the extent and the limits of hydraulic adjustment within plants.
Comparing different species, xylem structural changes have been shown to vary markedly within woody plants (Pockman and Sperry, 2000; Maherali et al., 2004; Pratt et al., 2007; Ducrey et al., 2008; Markesteijn et al., 2011). These studies focused on the adaptive significance of variation and evolutionary trade-offs between traits related to cavitation resistance or water transport capacity. In contrast to the abundance of studies on hydraulic conductivity and cavitation resistance in woody plants, similar hydraulic studies of herbaceous plants are much less prevalent. Only
Abbreviations: δ13C, carbon isotope ratio; gs, stomatal conductance; kh, hydraulic conductivity; kL, leaf specific conductivity; E(LA), leaf area specific whole plant transpiration; E (LM), leaf mass specific whole plant transpiration; P50, cavitation resistance; SD, stomatal density; SLA, specific leaf area; WD, wood density; WUE, water-use efficiency. © The Author 2014. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: [email protected]
Downloaded from http://jxb.oxfordjournals.org/ at Virginia Commonwealth Universtiy on May 28, 2014
Received 31 January 2014; Revised 11 April 2014; Accepted 22 April 2014
Page 2 of 10 | Matzner et al. herbaceous plants to agriculture, understanding the ability and limits of hydraulic adjustment is critical in attempting to breed for adaptive phenotypic plasticity over a range of conditions (Nicotra and Davidson, 2010). To test hydraulic trade-offs in response to light within an herbaceous annual and the importance of hydraulic acclimation, two cultivars of Phaseolus vulgaris were used that differed in both growth and hydraulic strategies (Mencuccini and Comstock, 1999). Othello is Middle American in origin and selected under irrigated conditions; G4523 is Andean in origin and selected under rain-fed conditions. Othello has smaller leaves and higher rates of transpiration compared with G4523. Specific questions addressed include: (i) are there hydraulic trade-offs within a herbaceous species in response to decreasing light, similar to evolutionary tradeoffs observed within interspecific studies of woody species; (ii) are their coordinated phenotypic shifts (plasticity) in hydraulic traits within P. vulgaris in response to light, and is this plasticity also constrained by trade-offs?
Materials and methods Plant material and experimental design In 2007 and 2008, two Phaseolus vulgaris L. (common bean) cultivars, Othello (Durango race) and G4523 (Nueva Grenada race) (Mencuccini and Comstock, 1999), were grown from seeds under three light levels in a GC-20 Plant Growth Chamber (ENCONAIR Ecological Chambers Inc., Winnipeg, Mb., Canada) at Augustana College, Sioux Falls, South Dakota, USA. The experiment was repeated in 2008 to ensure adequate replication because of space limitations. Replication ranged between 7–12 plants for each cultivar and treatment combination (2–8 in 2007; 4–5 in 2008). Plants were germinated in May or June and grown for 6–9 weeks before harvesting. Plants in 2008 were allowed to grow longer owing to low survival rates for low-light plants in 2007. Growth chamber temperatures were set at 25 °C daytime, and 20 °C at night. Humidity fluctuated with outside conditions, averaging 51.6% daytime, and 55.4% at night. Plants were grown in 4.2 l pots and exposed to low (AV=67.1 μmoles m–2 s–1 SE±7.5), medium (AV=311.9 μmoles m–2 s–1 SE±30.1), and high (AV=604.5 μmoles m–2 s–1 SE±82.8) light levels (top of canopy). Treatments were imposed within one week of germination and continued for the duration of the experiment. Shade cloth was used for low-light treatments, a fibreglass screen was used for medium light, and the high-light treatments were uncovered. Soil media was composed of potting soil, black dirt, peat moss, sand, perlite (1:1:1:0.5:0.5, by volume) with a bulk density of 0.70 kg l–1, and a total porosity of 0.74 (calculated assuming a particle density of 2.65 kg l–1). Treatments were watered at least three times per week with a 1/10 strength nutrient solution (Johnson et al. 1957). Supplemental slow-release fertilizer (approximately 100 g) was provided (14, 14, 14 NPK, Scotts-Sierra, Marysville, Ohio, USA). Biomass, leaf, and root measures Upon harvest, leaf and root area were measured using an Epson V700 photo-scanner and analysed using either WinFolia or WinRhizo software (Regent Instruments Inc.). Plant material was dried for at least 24 h at 70 °C; stem, root, and leaf mass were weighed separately and used to calculate root:shoot ratios. Specific leaf area was calculated from the leaf area divided by biomass. In 2008, stomatal density was measured on the ventral surface of two upper canopy leaves. Leaves were coated with liquid bandage (New-Skin®) which was allowed to dry, peeled off, and the stomatal impressions were counted within a 1 cm2 area.
Downloaded from http://jxb.oxfordjournals.org/ at Virginia Commonwealth Universtiy on May 28, 2014
a few comparisons have been made of cavitation resistance between species of herbaceous plants (Kocacinar and Sage, 2003; Rosenthal et al., 2010). Within-species comparisons of cavitation resistance between different genotypes of corn (Li et al., 2009), sugarcane (Neufeld et al., 1992), and rice (Stiller et al., 2003) have also been made. Plasticity in water-related traits has also been studied in herbaceous plants; however, most studies have focused on changes in biomass allocation or water-use efficiency (WUE; Heschel et al., 2004; Agrawal et al., 2008; Franks, 2011). Within herbaceous plants, studies have looked at embolism and refilling in maize and sunflower (McCully et al., 1998; Stiller and Sperry, 2002). Several studies have focused specifically on long-term structural acclimation in hydraulic traits within herbaceous annual plants and have demonstrated the importance of plasticity in hydraulic traits (Li et al., 2005; Holste et al., 2006; Rosenthal et al., 2010; von Arx et al., 2012). Plasticity in hydraulic traits in response to environmental changes within woody species has also received attention (Barigah et al., 2006; Domec et al., 2009; Schoonmaker et al., 2010; Corcuera et al., 2011; Plavcová and Hacke, 2012). For example, studies of acclimation to increased irradiance within woody species have observed that xylem vessel size or leaf specific whole-plant conductance increased (Lemoine et al. 2002a; Barigah et al., 2006) and cavitation resistance was higher (Cochard et al. 1999; Lemoine et al. 2002a; Barigah et al., 2006; Schoonmaker et al., 2010). Recently, Plavcová and Hacke (2012) demonstrated that constraints to phenotypic plasticity (similar to observed evolutionary trade-offs) can occur within a single hybrid Poplar exposed to various environmental treatments. If trade-offs can constrain plasticity within a woody species, these tradeoffs may also constrain plasticity within herbaceous annuals. Several interspecific hydraulic trade-offs have received attention, but perhaps the best studied is the “safety versus efficiency” trade-off with safety usually measured as the xylem pressure causing a 50% reduction in stem conductivity (P50) and “efficiency” measured as vessel size, specific conductivity, leaf specific conductivity (kL), or some other measure of the capacity to supply water to the canopy (Zimmermann, 1983; Ducrey et al., 2008; Poorter et al., 2010; Markesteijn et al., 2011). In our study, the leaf mass specific transpiration rate (E(LM)) averaged over 3–5 days was used as the capacity to supply water to the canopy. A “safety versus mechanical support” trade-off has also been suggested between greater xylem cavitation resistance and the cost of xylem mechanical reinforcement, often measured as wood density (WD) or conduit wall thickness (Hacke and Sperry, 2001; Pratt et al., 2007; Markesteijn et al., 2011). Trade-offs have been suggested between construction costs such as WD and measures of hydraulic efficiency such as kL (Santiago et al., 2004; McCulloh et al., 2011) or measures of growth (Poorter et al., 2010; Sobrado, 2010) or between hydraulic efficiency and WUE (Santiago et al., 2004; Sobrado, 2010). These evolutionary trade-offs constrain hydraulic traits within woody species in interspecific studies and in comparing populations within a species. Similar trade-offs may also constrain hydraulic traits within herbaceous plants owing to genetic differences or through constraints on plasticity. Given the importance of
Constraints to hydraulic acclimation in beans | Page 3 of 10
Vulnerability curves Xylem cavitation was induced on basal, central axis stems (same stems used to measure kh), using the centrifugal method (Alder et al., 1997). Cavitation was quantified by measuring the decline in flow rates after exposing stems to xylem pressures of –0.1, –0.5, –1.0, –1.5, and –2.0 MPa and expressed as a percent loss of conductivity (PLC) from initial hydraulic conductivity (kh) values. Relationship between xylem pressure and PLC were graphed and fitted with a regression line, and P50 (xylem pressure resulting in 50 % loss in conductivity) was calculated from the equation of the line. Wood density Frozen basal, central axis stem sections (2.5 cm) were longitudinally cut and the pith, phloem, and epidermis removed following Hacke et al. (2000) protocols. Samples were degassed under vacuum for 24 h. Fresh volume was determined by Archimedes’ principle; stems were submerged and displacement weight converted to fresh volume. Samples were dried (70 °C) for 48 h and WD (g cm–3) was calculated as the ratio of dry weight to fresh volume. Leaf carbon isotopes (δ13C) Leaf samples were dried, ground (
Research Paper
Constraints to hydraulic acclimation under reduced light in two contrasting Phaseolus vulgaris cultivars Steven L. Matzner*, David D. Rettedal, Derek A. Harmon and MacKenzie R. Beukelman Department of Biology, Augustana College, Sioux Falls, SD 57197, USA * To whom correspondence should be addressed. E-mail: [email protected]
Abstract Two cultivars of Phaseolus vulgaris L. were grown under three light levels to determine if hydraulic acclimation to light occurs in herbaceous annuals and whether intraspecific trade-offs constrain hydraulic traits. Acclimation occurred in response to reduced light and included decreased stomatal density (SD) and increased specific leaf area (SLA). Reduced light resulted in lower wood density (WD); decreased cavitation resistance, measured as the xylem pressure causing a 50 % reduction in stem conductivity (P50); and increased hydraulic capacity, measured as average leaf mass specific transpiration (E(LM)). Significant or marginally significant trade-offs between P50 and WD, WD and E(LM), and E(LM) and P50 reflected variation due to both genotype and environmental effects. A trade-off between WD and P50 within one cultivar indicated that morphological adjustment was constrained. Coordinated changes in WD, P50, and E(LM) within each cultivar in response to light were consistent with trade-offs constraining plasticity. A wateruse efficiency (WUE, measured as δ13C) versus hydraulic capacity (E(LM)) trade-off was observed within each cultivar, further indicating that hydraulic trade-offs can constrain acclimation. Larger plants had lower hydraulic capacity (E(LM)) but greater cavitation resistance, WD, and WUE. Distinct hydraulic strategies were observed with the cultivar adapted to irrigated conditions having higher stomatal conductance and stem flow rates. The cultivar adapted to rainfed conditions had higher leaf area and greater cavitation resistance. Hydraulic trade-offs were observed within the herbaceous P. vulgaris resulting from both genotype and environmental effects. Trade-offs within a cultivar reflected constraints to hydraulic acclimation in response to changing light. Key words: Cavitation, herbaceous, hydraulic conductivity, light, plasticity, trade-offs.
Introduction As evolutionary selection occurs at the level of the phenotype, both genotypic differences and acclimation in response to environmental conditions are important for an organism’s immediate survival as well as for future fitness. The ability to adjust water transport capacity or drought tolerance is particularly important as water deficit is a primary factor limiting crop yield (Boyer, 1982). With the threat of climate change and rising demands for scarce water resources it is increasingly important to understand both the extent and the limits of hydraulic adjustment within plants.
Comparing different species, xylem structural changes have been shown to vary markedly within woody plants (Pockman and Sperry, 2000; Maherali et al., 2004; Pratt et al., 2007; Ducrey et al., 2008; Markesteijn et al., 2011). These studies focused on the adaptive significance of variation and evolutionary trade-offs between traits related to cavitation resistance or water transport capacity. In contrast to the abundance of studies on hydraulic conductivity and cavitation resistance in woody plants, similar hydraulic studies of herbaceous plants are much less prevalent. Only
Abbreviations: δ13C, carbon isotope ratio; gs, stomatal conductance; kh, hydraulic conductivity; kL, leaf specific conductivity; E(LA), leaf area specific whole plant transpiration; E (LM), leaf mass specific whole plant transpiration; P50, cavitation resistance; SD, stomatal density; SLA, specific leaf area; WD, wood density; WUE, water-use efficiency. © The Author 2014. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: [email protected]
Downloaded from http://jxb.oxfordjournals.org/ at Virginia Commonwealth Universtiy on May 28, 2014
Received 31 January 2014; Revised 11 April 2014; Accepted 22 April 2014
Page 2 of 10 | Matzner et al. herbaceous plants to agriculture, understanding the ability and limits of hydraulic adjustment is critical in attempting to breed for adaptive phenotypic plasticity over a range of conditions (Nicotra and Davidson, 2010). To test hydraulic trade-offs in response to light within an herbaceous annual and the importance of hydraulic acclimation, two cultivars of Phaseolus vulgaris were used that differed in both growth and hydraulic strategies (Mencuccini and Comstock, 1999). Othello is Middle American in origin and selected under irrigated conditions; G4523 is Andean in origin and selected under rain-fed conditions. Othello has smaller leaves and higher rates of transpiration compared with G4523. Specific questions addressed include: (i) are there hydraulic trade-offs within a herbaceous species in response to decreasing light, similar to evolutionary tradeoffs observed within interspecific studies of woody species; (ii) are their coordinated phenotypic shifts (plasticity) in hydraulic traits within P. vulgaris in response to light, and is this plasticity also constrained by trade-offs?
Materials and methods Plant material and experimental design In 2007 and 2008, two Phaseolus vulgaris L. (common bean) cultivars, Othello (Durango race) and G4523 (Nueva Grenada race) (Mencuccini and Comstock, 1999), were grown from seeds under three light levels in a GC-20 Plant Growth Chamber (ENCONAIR Ecological Chambers Inc., Winnipeg, Mb., Canada) at Augustana College, Sioux Falls, South Dakota, USA. The experiment was repeated in 2008 to ensure adequate replication because of space limitations. Replication ranged between 7–12 plants for each cultivar and treatment combination (2–8 in 2007; 4–5 in 2008). Plants were germinated in May or June and grown for 6–9 weeks before harvesting. Plants in 2008 were allowed to grow longer owing to low survival rates for low-light plants in 2007. Growth chamber temperatures were set at 25 °C daytime, and 20 °C at night. Humidity fluctuated with outside conditions, averaging 51.6% daytime, and 55.4% at night. Plants were grown in 4.2 l pots and exposed to low (AV=67.1 μmoles m–2 s–1 SE±7.5), medium (AV=311.9 μmoles m–2 s–1 SE±30.1), and high (AV=604.5 μmoles m–2 s–1 SE±82.8) light levels (top of canopy). Treatments were imposed within one week of germination and continued for the duration of the experiment. Shade cloth was used for low-light treatments, a fibreglass screen was used for medium light, and the high-light treatments were uncovered. Soil media was composed of potting soil, black dirt, peat moss, sand, perlite (1:1:1:0.5:0.5, by volume) with a bulk density of 0.70 kg l–1, and a total porosity of 0.74 (calculated assuming a particle density of 2.65 kg l–1). Treatments were watered at least three times per week with a 1/10 strength nutrient solution (Johnson et al. 1957). Supplemental slow-release fertilizer (approximately 100 g) was provided (14, 14, 14 NPK, Scotts-Sierra, Marysville, Ohio, USA). Biomass, leaf, and root measures Upon harvest, leaf and root area were measured using an Epson V700 photo-scanner and analysed using either WinFolia or WinRhizo software (Regent Instruments Inc.). Plant material was dried for at least 24 h at 70 °C; stem, root, and leaf mass were weighed separately and used to calculate root:shoot ratios. Specific leaf area was calculated from the leaf area divided by biomass. In 2008, stomatal density was measured on the ventral surface of two upper canopy leaves. Leaves were coated with liquid bandage (New-Skin®) which was allowed to dry, peeled off, and the stomatal impressions were counted within a 1 cm2 area.
Downloaded from http://jxb.oxfordjournals.org/ at Virginia Commonwealth Universtiy on May 28, 2014
a few comparisons have been made of cavitation resistance between species of herbaceous plants (Kocacinar and Sage, 2003; Rosenthal et al., 2010). Within-species comparisons of cavitation resistance between different genotypes of corn (Li et al., 2009), sugarcane (Neufeld et al., 1992), and rice (Stiller et al., 2003) have also been made. Plasticity in water-related traits has also been studied in herbaceous plants; however, most studies have focused on changes in biomass allocation or water-use efficiency (WUE; Heschel et al., 2004; Agrawal et al., 2008; Franks, 2011). Within herbaceous plants, studies have looked at embolism and refilling in maize and sunflower (McCully et al., 1998; Stiller and Sperry, 2002). Several studies have focused specifically on long-term structural acclimation in hydraulic traits within herbaceous annual plants and have demonstrated the importance of plasticity in hydraulic traits (Li et al., 2005; Holste et al., 2006; Rosenthal et al., 2010; von Arx et al., 2012). Plasticity in hydraulic traits in response to environmental changes within woody species has also received attention (Barigah et al., 2006; Domec et al., 2009; Schoonmaker et al., 2010; Corcuera et al., 2011; Plavcová and Hacke, 2012). For example, studies of acclimation to increased irradiance within woody species have observed that xylem vessel size or leaf specific whole-plant conductance increased (Lemoine et al. 2002a; Barigah et al., 2006) and cavitation resistance was higher (Cochard et al. 1999; Lemoine et al. 2002a; Barigah et al., 2006; Schoonmaker et al., 2010). Recently, Plavcová and Hacke (2012) demonstrated that constraints to phenotypic plasticity (similar to observed evolutionary trade-offs) can occur within a single hybrid Poplar exposed to various environmental treatments. If trade-offs can constrain plasticity within a woody species, these tradeoffs may also constrain plasticity within herbaceous annuals. Several interspecific hydraulic trade-offs have received attention, but perhaps the best studied is the “safety versus efficiency” trade-off with safety usually measured as the xylem pressure causing a 50% reduction in stem conductivity (P50) and “efficiency” measured as vessel size, specific conductivity, leaf specific conductivity (kL), or some other measure of the capacity to supply water to the canopy (Zimmermann, 1983; Ducrey et al., 2008; Poorter et al., 2010; Markesteijn et al., 2011). In our study, the leaf mass specific transpiration rate (E(LM)) averaged over 3–5 days was used as the capacity to supply water to the canopy. A “safety versus mechanical support” trade-off has also been suggested between greater xylem cavitation resistance and the cost of xylem mechanical reinforcement, often measured as wood density (WD) or conduit wall thickness (Hacke and Sperry, 2001; Pratt et al., 2007; Markesteijn et al., 2011). Trade-offs have been suggested between construction costs such as WD and measures of hydraulic efficiency such as kL (Santiago et al., 2004; McCulloh et al., 2011) or measures of growth (Poorter et al., 2010; Sobrado, 2010) or between hydraulic efficiency and WUE (Santiago et al., 2004; Sobrado, 2010). These evolutionary trade-offs constrain hydraulic traits within woody species in interspecific studies and in comparing populations within a species. Similar trade-offs may also constrain hydraulic traits within herbaceous plants owing to genetic differences or through constraints on plasticity. Given the importance of
Constraints to hydraulic acclimation in beans | Page 3 of 10
Vulnerability curves Xylem cavitation was induced on basal, central axis stems (same stems used to measure kh), using the centrifugal method (Alder et al., 1997). Cavitation was quantified by measuring the decline in flow rates after exposing stems to xylem pressures of –0.1, –0.5, –1.0, –1.5, and –2.0 MPa and expressed as a percent loss of conductivity (PLC) from initial hydraulic conductivity (kh) values. Relationship between xylem pressure and PLC were graphed and fitted with a regression line, and P50 (xylem pressure resulting in 50 % loss in conductivity) was calculated from the equation of the line. Wood density Frozen basal, central axis stem sections (2.5 cm) were longitudinally cut and the pith, phloem, and epidermis removed following Hacke et al. (2000) protocols. Samples were degassed under vacuum for 24 h. Fresh volume was determined by Archimedes’ principle; stems were submerged and displacement weight converted to fresh volume. Samples were dried (70 °C) for 48 h and WD (g cm–3) was calculated as the ratio of dry weight to fresh volume. Leaf carbon isotopes (δ13C) Leaf samples were dried, ground (
