Figure S1

A)Relative gene expression of each PIP1 isoform in the stems of each transgenic line analyzed. B) Relative gene expression of each PIP2 isoform in the stems of each transgenic line analyzed. Letters denote homogeneous groups based on the Fisher LSD corrected for the multiple comparisons. Anova analysis revealed the presence of significant differences for all PIP1 genes tested (p < 0.001), while no differences were founded for PIP2 genes.

Figure S2

Changes of xylem sap osmotic potentials resulting from sugar (A) and ion (B) concentrations in response to stem water potential (wt, light red; transgenic, light green) subjected to increased water stress (xylem pressure). The estimation of osmotic potential from sugar was calculated as the equivalent of glucose content; the estimation of osmotic potential from ions was calculated as the equivalent of K+ ion concentration. Red (wt) and green (transgenic) circles represent the averages for well-watered, moderately and severely stressed plants. A one way Anova test suggests significant differences between treatments and lines (P <0.001). Letters denote homogeneous groups based on Fisher LSD method.

Figure. S3

Sugar (A) and ion (B) osmotic potential collected from xylem sap of well-watered, stressed and recovered plants. A one way Anova test suggests significant differences between treatments in both wt and transgenic plants (p< 0.001). Letters denote homogeneous groups based on the Fisher LSD method (lower-case letter, wt; upper-case letter, transgenic lines). Data are mean values and bars are SD.

PxEC50PLC / Wild -type / Transgenic lines
Initial P xylem [MPa] / -1.316±0.085 / -1.087±0.057
Re-watered P xylem [MPa] / -0.394±0.047 / -0.310±0.029
PLC recovery [%] / 95.45 *±23.81 / 43.69±9.06
PxEC50PLC
Initial P xylem [MPa] / -1.993±0.111 / -1.914±0.111
Re-watered P xylem [MPa] / -0.989±0.128 / -1.037±0.127
PLC recovery [%] / 76.40±26.42 / 64.39±11.25

Table. S1

Mean values (± SD) of xylem pressure in stressed and re-watered plants and % of PLC recoverywt and transgenic plants. Data are presented for moderately (PxEC50PLC)and severely stressed plants (PxEC50PLC).Wild-type poplars showed significantly different recovery of PLCfrom transgenic plants (t-test; t = 2.1150, df = 17, p < 0.05) while recovering from moderate stress, but not from severe stress (where both groups did not recover PLC despite significant reduction of tension.

Gene / Primer Sequences 5’-3’
Ubiquitin / For / CAGCTTGAAGATGGGAGGAC
Rev / CAATGGTGTCTGAGCTCTCG
Actin / For / GCAACTGGGATGATATGGAGA
Rev / TACGACCACTGGCATACAGG
PIP1.1 / For / CAAGCCCAGTTTGTTCCATT
Rev / CAGCCAAACCCCTCAAACTA
PIP1.2 / For / TTTGGCTACCATCCCCATAA
Rev / GGCGACGAAATTAAGCTCTG
PIP1.3 / For / GTGATGGAGGGCAAAGAAGA
Rev / ACAAGAAGGTGGCCATGAAC
PIP1.4 / For / CAAGAGGCAAGTGAGACCAA
Rev / GCAGCTGTTCCTATGGGTTG
PIP1.5 / For / AGCCAGAAGTCTTGGTGCAG
Rev / ACATCCATTGATTGGGAACG
PIP2.1 / For / TTTGGTGCTGCTGTCATCAT
Rev / GGGGTGGATAAAGACAAGCTC
PIP2.2 / For / CCGCCAACTAAAGAGGAAAA
Rev / TGGGCAAAAGAAGAAAGGTC
PIP2.3 / For / GGATTTTCTGGGTTGGACCT
Rev / TGGCGTACGTTCCATAACTG
PIP2.4 / For / CTCTCTCGCTCAAACCTTCG
Rev / GGTGGTGGGTCATGGTAGTC
PIP2.5 / For / AAGACAAGGCATGGGATGAC
Rev / AAGGACCCAAGGGCTTTAAC
PIP2.6 / For / ATTGGGCGCTGAGATTATTG
Rev / CAAATCCAATTGGGAGAGGA
PIP2.7 / For / ATGACCATTGGCTCTTCTGG
Rev / CCCATGAGAAGGCAAAGAGA

Table. S2

Sequences of primers used for quantitative real time PCR