Fig. 5

SRMT positively and directly regulates PtoRD26 which is epistatic to SRMT. Temporal expression patterns of PtoRD26 and SRMT in response to salt stress and ABA treatment. A The distribution of SRMT binding elements (ME) in the 1.5 kb promoter region of PtoRD26.1, named ME1-ME6 (red lines). The promoter regions (R1–R3) show the amplified fragments of ChIP-qPCR. B ChIP-qPCR detection of SRMT binding regions in the PtoRD26.1 promoter in vivo. There were significant enrichments in the R1 and R3 regions. C EMSA shows the GST-SRMT.3-fused protein binds to ME2, ME4, and ME5 in vitro. These bindings could be competitive by cold probes; the mutant probe could not be bound by the GST-SRMT.3-fused protein. D Dual-luciferase assay indicating that SRMT.3 bound to fourfold repeats of ME2, ME4 and ME5 to activate the expression of LUC gene in tobacco leaves. However, the mutated ME2, ME4 and ME5 could not be bound by SRMT.3 and thus LUC could not be activated. EPtoRD26 was epistatic to SRMT. This genetic relationship was determined by assessing the salt tolerance of the double transgenic poplars (L3 and L5 of PtoRD26-RNAi/SRMT.3-OE) in which PtoRD26 was knocked down in the background of SRMT.3-OE-L11, in comparison to PtoRD26-RNAi and SRMT.3 overexpression poplars. F The MDA content and EL in PtoRD26-RNAi and SRMT.3-OE and PtoRD26-RNAi/SRMT.3-OE poplar leaves after salt treatment. The means ± s.d. of all data from at least three biological replicates are shown. Asterisks indicate statistically significant differences (two-sided Student’s t test, **P < 0.01, *P < 0.05, ns = not significant). G The temporal expression levels of PtoRD26 and SRMT at 0, 1, 2, 4, 8, 16, 24, and 36 h after NaCl treatment. H The temporal expression levels of PtoRD26 and SRMT at 0, 1, 2, 4, 8, 16, 24, and 36 h after ABA treatment. I The comparison of the temporal expression pattern of PtoRD26 in responsive to NaCl treatment in the background of WT and SRMT-RNAi poplars, respectively. J The comparison of the temporal expression pattern of PtoRD26 in response to ABA treatment in the background of WT and SRMT-RNAi poplars