Trait measurements
We measured 27 traits (Table 2) which together describe plant
morphology, leaf structure, plant gas exchange and its temperature
dependence, and plant water use strategy. For the thirteen of these
traits which we measured at more than one elevation, we calculated
plasticity indices. We made these trait measurements on 16 target
species (Table S1), chosen to represent several plant families, to be
widely present in the turfs in most treatments, and to have leaves which
fitted into the gas analyzer.
Traits describing plant morphology and leaf structure were measured at
the source elevation (2000 m) in August 2019 and 2020. To measure leaf
area (LA), leaf mass per area (LMA), leaf thickness (LT), and leaf dry
matter content (LDMC), we collected leaves of the target species,
scanned them and measured their fresh and oven-dried weight. LT was
measured with calipers from the fresh leaves. LA was estimated from the
leaf scans with the software ImageJ (Rasband 1997). Canopy shape index
(CSI) was measured as the ratio of mean canopy width to height (ranging
from near zero for tall plants to >1 for prostrate plants).
Rooting depth (Rootd) and width (Rootw)
were measured from excavated root systems.
To characterize plant gas exchange, we measured photosynthesis (A),
nighttime respiration (R), stomatal conductance during day and night
(GS, GSnight), transpiration during day and night (E,
Enight; for the importance of night-time water flux, see
Snyder 2003; Daley & Phillips 2006), instantaneous water use efficiency
(WUEinst) and intrinsic water use efficiency
(WUEint) with an infra-red gas analyser (CIRAS-2,
PPSystems, Hitchin, UK). To quantify how photosynthesis or respiration
varied after transplantation to lower elevation, measurements were taken
from two sites: +3°C (1400 m) and +0°C (2000 m). Temperature in the leaf
chamber was kept constant at either 24°C (day measurements) or 14°C
(night measurements). Gas exchange measurements were taken in 2019 and
2020, 3-4 years after the transplantation.
To characterize the instantaneous response of gas exchange to
temperature, we measured photosynthesis and respiration response to
temperature at 2000 m (+0°C) using both a CIRAS-2 and Licor-6800 (LICOR,
Lincoln, Nebraska, USA). Photosynthesis was measured at 14°C – 34°C, at
4°C intervals, and respiration at 5°C – 30°C, at 5°C intervals. After,
the leaves were scanned, oven-dried, weighted and milled to measure
carbon (C) and nitrogen (N) content either with combustion analysis (for
large samples) or with an elemental analyzer coupled to a mass
spectrometer (for small samples, Werner et al. 1999; Werner &
Brand 2001). Leaf scans and weights were used to estimate change in LMA
and LA resulting from transplantation. Photosynthetic nitrogen use
efficiency (PNUE) was calculated by dividing mass-based photosynthetic
rate (A/LMA) with the N content of the same leaf.
Plant water use strategy was characterized by measuring stable isotope
ratios in 13C/12C and18O/16O (‰) from leaf cellulose to
determine long-term stomatal conductance and water use efficiency
(δ13C and δ18O; Scheidegger et
al. 2000; Moreno‐Gutiérrez et al. 2012). δ13C
is a proxy for intrinsic water use efficiency (A/GS) and
δ18O correlates negatively with stomatal conductance
(GS), both integrated over the leaf life span. Leaf samples collected at
each site were oven-dried and cut into small pieces for cellulose
extraction. Measurements of δ13C and
δ18O from the extracted cellulose were performed using
a PYROCUBE (Elementar, Hanau, Germany) connected to a Delta Plus XP mass
spectrometer (Thermo Finnigann, Bremen, Germany) at the laboratory of
stable isotopes at the Swiss Federal Institute of Forest, Snow and
Landscape (WSL), following Weigt et al. (2015).
We measured the depth of water uptake by comparing the
δ18O of root crown water to the δ18O
of water from different soil depths (Barnard et al. 2006;
Moreno‐Gutiérrez et al. 2012). Root crowns were collected into
airtight vials and pooled together per species and site to ensure enough
water per sample. Soil was collected from each site with soil corers
(max 35 cm depth) and separated every 5 cm into airtight glass vials.
Water was extracted from samples in the lab by using a water bath and
cooling traps. δ18O was analysed from the extracted
water with a High Temperature Combustion Elemental Analyzer (TC/EA;
Thermo Finnigan, Bremen, Germany) and a mass spectrometer (Delta plus
XP; Thermo Finnigan) (Saurer et al. 2016). Since each site was
sampled on only one occasion (and the measurements could be influenced
by weather), we used the average depth across all four sites as the
species-level trait in the analyses. As source water
δ18O can drive variation in leaf cellulose
δ18O (Roden & Farquhar 2012), we used
δ18O enrichment (δ18O leaf cellulose
– δ18O root crown water) as the estimate of long-term
stomatal conductance (Scheidegger et al. 2000; Moreno‐Gutiérrezet al. 2012). See Supporting Information Table S2 for sample
sizes for all traits.