Real-time AA and MeOH emission measurements
Experimental details of the leaf, branch, and ecosystem gas exchange
methods to determine AA and MeOH emissions, as well as the detached
stem, detached leaf, and hydrated AIR temperature response curves can be
found in the supplementary methods. Briefly, emission rates of MeOH and
AA were quantified in real-time (roughly 2.5 measurements per minute)
using a high sensitivity quadrupole proton transfer reaction mass
spectrometer (PTR-MS, Ionicon,
Innsbruck Austria, with a QMZ 422 quadrupole, Balzers, Switzerland). The
PTR-MS was regularly calibrated to a primary standard by dynamic
dilution (Supplementary Figure S1 ). AA and MeOH emissions were
determined using PTR-MS at the leaf level using an environmentally
controlled leaf photosynthesis system (Model 6800, Licor Biosciences,
USA), branch level using a custom 5.0 L transparent Tedlar gas exchange
enclosure with artificial lighting, and from a temperature-controlled
chamber used for detached leaf, stem, and hydrated AIR AA and MeOH
emission studies (Model 150 Dynacalibrator, +/- 0.01 C temperature
accuracy, Vici Metronics, USA). Together with air temperature,
continuous above canopy ambient AA and MeOH concentrations during the
growing season were made at a
poplar
plantation in Belgium (Portillo-Estrada et al., 2018), a mixed
hardwood forest in Alabama
(Suet al., 2016), and above a citrus grove in California
(Parket al., 2013). Vertical ecosystem fluxes of MeOH and AA were
estimated at the Belgium field site using the technique of eddy
covariance employing high frequency vertical wind and MeOH and AA
concentration measurements
(Portillo-Estradaet al., 2018). While ecosystem concentration and flux
measurements MeOH were collected at all three sites using eddy
covariance with PTR-TOF-MS, only the Belgium poplar plantation reported
ecosystem scale AA flux data. At the Alabama mixed forest site, AA
fluxes were not reported
(Suet al., 2016) and at the citrus grove in California, AA fluxes
were reported to suffer from inlet dampening of high frequency
concentration variations
(Parket al., 2013). This loss of AA flux signal was explained by
dampening of fast fluctuations in the sample tube due to stickiness of
AA with respect to the inert tubing walls. Therefore, at the Alabama and
California sites, diurnal ambient concentrations of MeOH and AA were
analyzed instead of fluxes as a function of air temperature.
Long-distance13C2-acetatetransport in the transpiration stream and leaf cell wall
O-acetylation interactions
In order to evaluate the possibility of long-distance metabolic
interactions between plant tissues mediated by acetate in the
transpiration stream, including influencing O -acetylation
dynamics of cell walls, 13C2-acetate
labeling studies were carried out on individual P. trichocarpatrees transferred from the greenhouse to the laboratory.13C2-acetate delivery to leaves was
accomplished using detached branches (N = 3 branches, 1
branch/individual) placed in a 10 mM solution of sodium13C2-acetate (Sigma-Aldrich, USA) for
2 days inside an environmentally controlled growth chamber (Percival
Intellus Control System, USA) maintained at 27.5 °C daytime temperature
(6:00–20:00; 30% light) and 23 °C nighttime temperature (20:00-6:00).
After 2 days, the branches took up roughly 30-40 ml of the13C2-acetate -acetate solution. In
addition, a single individual of 2.1 m height was placed in the
laboratory under automated daytime lighting with continuous daytime (150
µl min-1) and nighttime (70 µl
min-1) xylem injection at the base of the stem with a
10 mM sodium 13C2-acetate acetate
solution (1,176 ml injected over 7 days using a flow controlled M6 Pump,
Valco Instruments Co. Inc., USA). Following the13C2-acetate -acetate labeling period
(branch: 2 day, tree: 7 day), a mature leaf was removed and flash frozen
under liquid nitrogen and stored at -80 °C before isolating whole leaf
cell walls through the generation of AIR. Leaf AIR samples were also
prepared from detached branches fed with water and 10 mM acetate with
natural 13C/12C abundance as
controls. Experimental details of the AIR saponification followed by13C-labeling analysis of the released acetate can be
found in the supplementary methods.