Investigation Of Mechanisms Underlying Arabidopsis MAG2-and Homolog Protein MAL-Regulating Polar Auxin Transport And Plant Growth And Development

In higher plants,the seeds store a large amount of storage proteins,which provide nitrogen sources for seed germination,and nutritional needs of the seedlings during early growth.During seed maturation,a large amount of seed storage proteins(SSPs)are synthesized as precursors in s in the rough endoplasmic reticulum(rER),and then transported by vesicles to protein storage vacuoles(PSVs),processed into mature forms and deposited.In previous studies,we found that the Arabidopsis MAG2 complex is involved in vesicle transport between the ER and the Golgi apparatus.When the subunit(s)of the MAG2 complex is(are)depleted,the export of the storage protein precursors from the ER is blocked,resulting in abnormal accumulation of the precursors of 12S globulins and 2S albumins.The transport vesicles are delivered along the cytoskeleton toward the target membrane,and then through tethering,docking and membrane fusion to complete the vesicle trafficking.The initial interaction between the transport vesicles and the target membrane is conducted by tethering factors.The tethering factors capture and recognize the vesicles and then pull them close to the target membrane.This process is called tethering.After tethering,the transport vesicles are anchored to and fused with the target membrane.MAG2 complex is localized on the ER,might regulate Golgi-derived vesicle fusion with the ER membrane.MAG2-like(MAL)is Arabidopsis homolog protein of MAG2.In this study,we used mag2-1 and mal-1 single mutants and mag2-1 mal-1 double mutant,as well as MAG2-and MAL-overexpressing plants as materials,to investigate the effects of MAG2 and MAL depletions on auxin transport,plant growth and development and stress trsponses.1.By observation of the growth and development of the mutants and overexpressing plants,we found that compared with wild type,the seedlings of the mag2-1 mal-1 double mutant have shorter root length,and the plants have shorter height,smaller rosette leaves,smaller seeds and lower thousand grain weight.The seedlings of MAG2-overexpressing plants have longer root length,and the plants have higher height as well.These phenotypes suggest that MAG2 and MAL play important roles in regulation of plant growth and development.Detection of BiP1/2 protein level reveals that BiP1/2 protein level in the single and double mutants are higher than that in wild type,suggesting that loss of MAG2 and MAL result in vesicle transport defects which induce ER stress.2.GUS stain results indicate that compared with wild type,DR5::GUS distribution in mag2-1 single mutant and mag21l mal-1 double mutant show significant differences in the root tips,lateral root primordia and leaves from seedlings,indicating that MAG2 and MAL regulate auxin distribution.Confocal observation reveals that the expression level of PIN 1-GFP and PIN3-GFP in the root tips of the mutants are reduced,indicating that MAG2 and MAL are important regulators of the expression of the auxin transporters,PIN1 and PIN3.By observation of the effects of exogenous auxin and auxin transport inhibitors on the mutants and overexpressing plants,we found that mag2-1 single mutant and mag2-1 mal-1 double mutant were less sensitive to the NAA and NPA treatments than wild type,indicating MAG2 and MAL deficiencies affect auxin transport.All these results suggest that MAG2 and MAL modulate auxin polar transport via regulating PIN protein expression.3.NaCl,mannitol and ABA treatments indicate that mag2-1 and mag2-1 mal-1 are sensitive to salt,osmotic and ABA stresses,suggesting that loss of MAG2 and MAL affect plant stress responses.In addition,yeast two-hybrid analysis revealed that MAG2 and MAL interact with the protein kinase isoforms of SnRK2 which functions in ABA signaling pathway,suggesting that MAG2-and MAL-mediated ER-Golgi transport pathway is closely related to the ABA signaling pathway.