“Low-cost” initial burst of root development in whole Fagus crenata seedlings: The key to survival?

Terrestrial plants are rooted in one place, and therefore their metabolism must be flexible to adapt to continuously changing environments. This flexibility is probably influenced by the divergent metabolic traits of plant organs. However, direct measurements on organ-specific metabolic rates are particularly scarce and little is known about their roles in determining whole-individual meatabolism. To reveal this on seedlings of Fagus crenata, which is one of the most widespread dominant genus in temperate deciduous broad leaf forests in the circum-polar Northern Hemisphere, we measured respiration, fresh mass and surface area for total leaves, stems and roots of 55 individuals in two years from germination and analyzed their relationships with individual metabolism. Proportion of roots to whole plant in mass increased from approximately 17% to 74%, and that in surface area increased from about 11% to 82% in the two years. Nonetheless, the increment of the proportion of root respiration to whole-plant respiration was from 9.2% to only 40%, revealing that the increment in mass and surface area of roots was much larger than the increment in energetic cost. As a result, only the roots showed a substantial decline in both respiration/surface area and respiration/mass among the three organs; roots had about 90% decline in their respiration/surface area, and 84% decline in their respiration/mass, while those in leaves and stems were relatively constant. The low-cost and rapid root development is specific to the two years after germination and would be effective for avoiding water and nutrient deficit, and possibly helps seedling survival. This drastic shift in structure and function with efficient energy use in developmental change from seeds to seedlings may underpin the establishment of F. crenata forests. We discuss significance of lowering energetic cost for various individual organisms to effectively acquire resources from a wide perspective of view.


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Individual metabolism is a fundamental process that transforms energy and materials to 3 50 support various biological processes as a base for adaptation to changing environments [1,2]. 51 Therefore, the metabolic rate has profound physiological, ecological and evolutionary implications 52 [3,4], which would be a key to understand and predict the effects of climate change on organisms 53 and ecosystems [1,2]. 54 In general, the metabolic rate (i.e. respiration rate, R) of individual organisms scales with body 55 size (X), and is usually described as the simple power function of body mass: where a is a normalization constant, and b is the scaling exponent (slope on the log-log coordinates) 58 [5][6][7]. The equation (1) represents the emergent outcomes of the metabolism of individuals under 59 various constraints [2,8,9]. Therefore, to obtain a mechanistic insight into the regulation of scaling of 60 metabolic rate, we need empirical evidence of whole-organism measurements. However, little is 61 known about the relationships between metabolic rate and body size with the reliable data from small 62 to giant individuals [10]. This is because most of the studies on metabolic scaling have been based on 63 indirect evidence and aimed to construct theoretical models to explain the exponent b, which has 64 widely been assumed to be 3/4 as suggested by the WBE model [5][6][7][8][9]. The size scaling of individual 65 organisms results from the sum of differentiated organs with distinctive functions and structure, each 66 one showing contrasting responses under changing environments [11][12][13]. Therefore, evaluating 67 metabolic rate of each organ is crucial to understand the scaling of metabolism associating with body 68 size, namely mass or surface area. 69 Terrestrial plants are supposed to adapt to various environments by adjusting the biomass 70 partitioning among organs, as typically shown in the root/shoot ratio [11][12][13][14][15][16][17]. To date, the optimal 71 partitioning theory has mainly evaluated allocation between shoot and roots in mass. It suggests that 72 plants should allocate more biomass to the shoot when limiting resource is carbon and to the roots 73 when limiting resource is water or nutrient [11,17]. However, in spite of the significant implication 74 of metabolic rate, few studies have compared metabolic rate of shoot (leaves + stems) and roots at 4 75 the whole-plant level [18]. The comparison of respiration rate between shoot and roots, with 76 measurement of organ-specific respiration at the individual level, would provide a new insight into 77 the energy partitioning and would progress our understanding about whole-plant adaptation for 78 resource acquisition.

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The purpose of this study is to understand the processes of establishment of individual individual. To test this assumption, we need size-scaling of respiration that cover seedlings in 88 current-year of germination and 1-year old.

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Here, we show the respiration rates of total leaves, stems, and roots of various sized seedlings

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Ethics statement 100 Our study included fieldwork activities for collecting F. crenata seeds and seedlings, and were 101 conducted in Japanese National Forest. The field work was permitted by the Shonai District Forest

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Office and did not involve any endangered or protected species.  The pots were filled with commercially available Kanuma pumice mixed with leaf mold, and 111 kept in a place with sufficient natural light and well-watered. We conducted every measurement at 112 the whole-plant level, using total leaves (including cotyledon), stems, and roots. The whole-plant 113 fresh mass ranged between 41.2×10 −5 and 23.5×10 −3 (kg) from the smallest current-year seedling to 114 the largest 1-year-old individual, as compiled in Table 1.   122 We separated the seedlings into leaves, stems, and roots, and enclosed them separately in 123 custom-made chambers (80 or 160 cm 3 ), promoting air circulation within the chamber using a DC 124 fan. We confirmed that the separation did not have an effect on the measured values of whole-plant 125 respiration, as reported by Mori et al. [10]. Increment rates of CO 2 concentrations within the closed 126 air-circulation system were measured every 5 seconds using an infrared CO 2 analyzer (GMP343,

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Vaisala, Helsinki, Finland), and normalized to 20℃ assuming a standard Q 10 = 2. During the 128 measurements, we kept the plant materials wrapped in wet paper to prevent transpiration, and the 129 measurements were taken within 20 minutes of the excavation for each seedling. Data analysis 138 We fitted the respiration-fresh mass and respiration-area scaling relationships using a 139 simple-power function on log-log coordinates, based on reduced major axis regression (RMA) [28] 140 of the log transformed version of equation (1), using all the measurement data of the 55 seedlings 141 from current-year to 1-year-old. We also analyzed the size-scaling values for surface area in relation   Table 2. Scaling of respiration rate (µmol sec −1 ) of whole plant, roots, leaves, and stems with 166 their fresh mass (kg) and surface area (m 2 ) fitted by equation (1). 167 In all regression, number of observation = 55, using RMA on log-log coordinates (for all cases, P < 168 0.001).

Respiration of organs 171
To see how organ-specific respiration contribute to the size-scaling of R shown in Fig. 1, we   172 analyzed the size-scaling values for area in relation to mass (Fig. 2), and evaluated the relationships 173 between respiration of roots, leaves, and stems (R R , R L , R S ; μmol s −1 ) and their fresh mass (M R , M L , M S ; kg) and surface area (S R , S L , S S ; m 2 ) separately, at the whole-organ level (Fig. 3, Table 2). values was not significant, the exponent b of R R to S R was relatively lower than that of R R to M R , 195 indicating that the increase in S R was more efficient than that in M R . This seems to represent an  Table 3. The scaling of whole-plant respiration is determined by 1) the relative contribution of each 239 organ to total mass and surface area, and 2) the organ-specific respiration per unit mass and surface 240 area. Table 1 shows the maximum and minimum values of respiration, mass, and surface area of revealing that the increment in the proportion of roots to whole plant is much more larger in mass 247 and surface area than in energetic cost.

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These results revealed that the similarity between the R-M (Fig. 1a) and R-S (Fig. 1b) 249 relationships in their scaling exponents (Table 2)  Ontogenetic shift in root/shoot ratio from seedlings to mature trees 268 Our study suggests the need for further work at the whole-plant level up to mature trees, to 269 clarify the role of low-cost rapid root development, beyond the initial seedling stage. Figure  individuals, respectively that were obtained from our prior work.

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As to M R /M Shoot , the rapid increment is specific of the seedling stage, but after that, it gradually 285 declines with size (and presumably age) in both pot-and field-grown individuals. and nutrient deficit [11,17], and reduce mortality of seedlings. the underlying processes to effectively improve whole-plant energetic efficiency. In that case, it may 317 be an energy-saving process of seedlings that is comparable to that in animal locomotion for resource 318 acquisition [35][36][37][38]. In the present study, the measurement at the whole-plant level revealed the 319 drastic reduction in energetic cost for rapid root development that would underpin the population