Photorespiration response and biomass production
The two most important factors explaining the variation in the
D6S /D6R ratio (range:
0.85-0.94) were WT and atmospheric CO2 concentration,
which respectively accounted for 48% (P< 0.001) and
14% (P< 0.001) of the total variance (Fig. 2A, Table
1). In addition, temperature and interactions between WT and
CO2 both explained 7% each (P= 0.004 andP= 0.005, respectively) of the variance in this ratio. Increasing
atmospheric CO2 from 280 to 400 ppm resulted in a 0.03
decrease in the
D6S /D6R ratio at low
WT, but had no significant effect on it at high WT (Fig. 2A, Table S1).
Together with the observed interaction between CO2 and
WT, this indicates WT-dependent suppression of photorespiration at the
high CO2 level. Raising the WT from -20 to
~0 cm resulted in a significant (0.01-0.05) increase in
the D6S /D6R ratio,
indicating that the high WT increased the
photorespiration/photosynthesis ratio (Fig. 2A). Increasing the
day/night temperatures from 12˚C/7˚C to 17˚C/12˚C caused a small
(~0.01) increase in the
D6S /D6R ratio at low
WT, but increasing the light intensity from 250 to 500 µmol
m-2 s-1 had no significant effect on
it (Fig. 2A, Table 1).
Theoretically, the suppression of
photorespiration at high CO2 and low WT should have been
accompanied by increases in CO2 assimilation rates, and
thus biomass production, but no significant increase in biomass
accumulation associated with the high CO2 level was
observed under any test conditions (Fig. 2B). The variation in biomass
production (0.2-3.6 g m-2 d-1) was
mostly explained by temperature (27%, P< 0.001), WT
(52%, P< 0.001), and to a smaller degree light
intensity (4%, P= 0.003) and the interaction between temperature
and WT (4%, P= 0.005; Table 1). Increasing the temperature caused
a massive 2.6- to 4.5-fold increase in biomass production, whereas
raising the WT strongly reduced biomass production, by 53-74% (Fig.
2B). Increasing the light intensity caused a 1.1- to 1.5-fold increase
in biomass. No major between-differences in C content of the biomass
(48.1 ± 0.11%, SE) were detected (Fig. S3). Thus, the observed changes
in biomass production reflect proportional variations in C accumulation.
Whole-tissue δ13C and chloroplastic to ambient
CO2 concentration
To further investigate physiological effects of increasing atmospheric
CO2 from 280 to 400 ppm, we analyzed13C discrimination by measuring whole-tissue
δ13C signatures (Fig. 3A). Variation in
δ13C (which ranged from -30.5 to -25.7‰) was explained
by changes in WT (48%, P< 0.001), atmospheric
CO2 (13%, P< 0.001), temperature (10%,P< 0.001), light intensity (3%, P= 0.02), and the
interaction between CO2 and WT (10%,P< 0.001; Table 1). Increasing atmospheric
CO2 consistently decreased δ13C (by
1.4 to 1.7 ‰) at low WT, but had no significant effect at high WT (Fig.
3A, Table S1). Concomitantly, raising the WT resulted in a 0.5-2.5‰
increase in δ13C. Increasing the temperature caused a
significant, 0.5-1.0 ‰, increase in δ13C. Increasing
the light intensity resulted in a small 0.3-0.6‰ increase in
δ13C at low WT.
The δ13C data allowed estimation of the chloroplastic
CO2 concentration (c c) and
subsequently the chloroplastic to ambient CO2 ratio
(c c/c a, Flanagan &
Farquhar, 2014), which is a key determinant of metabolic C fluxes.
Variation in c c/c a(0.63-0.80, Fig. 3B) was explained by WT (57%,P< 0.001), light intensity (4%, P= 0.022),
temperature (3%, P= 0.035), atmospheric CO2 (3%,P= 0.046) and the interaction between CO2 and WT
(13%, P< 0.001; Table 1). Increasing atmospheric
CO2 significantly increasedc c/c a, by 0.03-0.05, at
low WT but had no significant effect at high WT (Fig. 3B, Table S1).
Raising the WT caused a 0.02-0.09 decrease inc c/c a, and increasing the
temperature decreasedc c/c a by 0.02 at low WT.
Altogether, this indicates that increases in atmospheric
CO2 increase c c particularly at
low WT, whereas raising the WT reduces c c. A
strong negative correlation was detected betweenc c/c a and the
D6S /D6R ratio at low
WT (R2=0.85, P< 0.001), suggesting
that an increase in c c caused the decrease in
photorespiration/photosynthesis ratio, i.e. suppression of
photorespiration at low WT. No significant relationship in these
variables was observed at high WT (R2=0.10,P =0.208).
Variation in the
CO2 diffusion gradient (72-150 ppm) from the atmosphere
to the chloroplasts
(c a-c c) was mostly
explained by CO2 (53%, P< 0.001), WT
(26%, P< 0.001) and the interaction of
CO2 and WT (9%, P< 0.001). Increasing
atmospheric CO2 resulted in increases inca-cc of 15-52 ppm (Fig. S3,
Table S2). This indicates that increases in atmospheric
CO2 increased CO2 assimilation.