Endomembrane-biased dimerization of ABCG16 and ABCG25 transporters determines their substrate selectivity in ABAregulated plant growth and stress responses

ATP-binding cassette (ABC) transporters are integral membrane proteins that have been evolved with diverse functions via transport of various substrates. In Arabidopsis, the G subfamily of ABC proteins is particularly abundant and participate in multiple signaling pathways during plant development and stress responses. Here, we revealed two Arabidopsis ABCG transporters, ABCG16 and ABCG25, were engaged in ABA-mediated stress responses and early plant growth through an endomembrane-specific dimerization-coupled transport of ABA and ABA-glucosyl ester (ABA-GE), respectively. We first disclosed that ABCG16 contributed to osmotic stress tolerance via ABA signaling. More specifically, ABCG16 acted in stimulation of cellular ABA efflux in both yeast and plant cells. Combined with FRET (Förster resonance energy transfer) analysis, we showed that ABCG16 formed obligatory homodimers for ABA export activity and that the plasma membrane resided ABCG16 homodimers specifically responded to ABA with notable conformational changes within the homodimers. Furthermore, we demonstrated that ABCG16 heterodimerized with ABCG25 at the ER membrane and facilitated the ER entry of ABA-GE in both Arabidopsis and tobacco cells. The specific responsiveness of the heterodimer ABCG16/ABCG25 to ABA-GE and the superior growth of the double mutant jat1-2abcg25 supported an inhibitory role of the two ABCGs in early seedling establishment via regulation of ABA-GE translocation across the ER membrane. Together, our endomembrane-specific analysis of the FRET signals derived from the homo- or heterodimerized ABCG complexes allowed us to link an endomembrane-biased dimerization partnership to distinct substrate translocation by ABCG transporters, providing a prototypic framework for understanding the omnipotence of ABCG transporters in plant development and stress responses.