A Coupling Of Clathrin-dependent Endocytosis And Exocytosis In Arabidopsis

Cells transport substances away or to the plasma membrane(PM)through endocytosis and exocytosis,including some PM proteins with important biological functions.Therefore,the coupling between endocytosis and exocytosis is essential to maintain the integrity of PM and control the abundance and activity of PM proteins.Clathrin,an evolutionarily conserved vesicle coat protein,plays a critical role in vesicle trafficking.In plants,clathrin locates at the PM and trans-Golgi network/early endosome(TGN/EE),and mediates endocytosis by interacting with adaptor protein complexes AP-2 and TPC at the PM,but its function at the TGN/EE is still unclear.In this study,we demonstrated that the recruitment of Arabidopsis clathrin to the TGN/EE was mediated by adaptor protein complex 1(AP-1),and defective for clathrin or AP-1 function impaired the secretion and recycling pathway,suggesting that clathrin-mediated exocytosis(CMX)exists in plants.Furthermore,we found that defects in AP-1-dependent CMX mutants significantly reduced recruitment of clathrin,AP-2,and TPC subunits to the PM,thereby inhibiting CME(clathrin-mediated endocytosis,CME).In addition,destruction of exocytosis by pharmacological treatment also inhibited clathrin,AP-2,and TPC recruited to the PM and slowed down the endocytosis.Likewise,in mutants with defective AP-2-/TPC-dependent CME,the recruitment of clathrin and AP-1 to the TGN/EE was found to be significantly reduced,thereby inhibiting CMX.Collectively these findings indicate that the association of clathrin and its adaptor protein with the PM and TGN/EE is coordinately regulated,thereby coupling the processes of endocytosis and exocytosis in plant cells.Furthermore,this study showed that the internalization of SNARE protein VAMP721/722 depend on CME,while the association of clathrin with the PM and TGN/EE in vamp721 vamp722 mutant was significantly reduced,indicating that VAMP721/722 is involved in the coupling of CME and CMX.Together,these results suggest that CME and CMX are coupled tightly in plant cells,which is a new mechanism for cells to maintain the balance between endocytosis and exocytosis,and VAMP721/722 may play an important role in this coupling regulation.