Abstract
Two Arabidopsis genes have been so far demonstrated in planta to function in vacuolar sorting of seed storage proteins: AtVSR1 and MAG1/AtVPS29. We developed an efficient and simple method for isolating vacuolar sorting mutants of Arabidopsis. When GFP fusion with the signal (GFP-CT24) was expressed in atvsr1, mag1/atvps29, both atvsr1 and mag1/atvps29 gave strongly fluorescent seeds, suggesting that a defect in vacuolar sorting provided fluorescent seeds due to secretion of GFP-CT24. We mutagenized transformant seeds expressing GFP-CT24. From ~3,000,000 lines of M2 seeds, we obtained more than 100 fluorescent seeds and designated them as green-fluorescent seed (gfs) mutants. We mapped gfs1 to AtVSR1, gfs2 to KAM2/GRV2, gfs10 to the At4g35870 gene encoding a novel membrane protein and the others to different loci. The method should provide valuable insights into the complex molecular mechanisms underlying vacuolar sorting of storage proteins.
