TaUBC24 interacted physically with TaICE1
According to the high ICE1 content of OEmiR399 transgenic Arabidopsis and CsUBC24 interacting with CsICE1 in previous study (R. Wang et al., 2020), we supposed TaUBC24 might interact with TaICE1. To confirm the protein-protein interaction between poplar TaUBC24 and TaICE1, the full-length CDS of TaICE1 fused to AD (AD- TaICE1) was tested interaction with the full-length CDS of TaUBC24 fused to BD (BD- TaUBC24). As shown in Fig. 13A, an interaction between the TaUBC24 and TaICE1 was detected in the Y2H assay.
To verify the protein interaction between TaUBC24 and TaICE1 in plant cell, the bimolecular fluorescence complementation (BiFC) was performed. Coding sequences of TaUBC24 were fused to the N-terminal fragment of green fluorescent protein (GFP) (TaUBC24 -GFPN), while TaICE1 was fused to the C-terminal fragment of GFP (TaICE1 -GFPC). GFP fluorescence was detected in the nucleus of N. benthamiana epidermal cells co-transfected with plasmids that harbored either theTaUBC24 -GFPN andTaICE1 -GFPC (Fig. 13B). These results showed that TaUBC24 was capable of interacting with TaICE1 in vivo.