Protein Targeting and Vesicular Trafficking
Fuji, Kentaro , Shimada, Tomoo , Takahashi, Hideyuki , Tamura, Kentaro , Koumoto, Yasuko , Nishizawa, Keito , Maruyama, Nobuyuki , Utsumi, Shigeru , Hara-Nishimura, Ikuko .
Vacuolar-Sorting Mutants (green-fluorescent seed) Can Be Efficiently Identified by Secretion of Vacuole-Targeted GFP in Their Seeds of Arabidopsis.
Two Arabidopsis genes have been so far demonstrated in planta to function in vacuolar sorting of seed storage proteins: a vacuolar sorting receptor, AtVSR1, and a homolog of the yeast retromer component, MAG1/AtVPS29. Here we show an efficient and simple method for isolating vacuolar sorting mutants of Arabidopsis. The method was based on two findings in this study. First, AtVSR1 functioned as a sorting receptor for ?-conglycinin, a soybean seed storage protein, by recognizing the vacuolar targeting signal. Secondly, when GFP fusion with the targeting signal (GFP-CT24) was expressed in atvsr1, mag1/atvps29, and wild-type seeds, both atvsr1 and mag1/atvps29 gave strongly fluorescent seeds and the wild-type did not, suggesting that a defect in vacuolar sorting provided fluorescent seeds due to secretion of GFP-CT24 out of the cells. 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, all of which caused mis-sorting of storage proteins. We mapped gfs1 to AtVSR1, gfs2 to KAM2/GRV2 and the others to different loci. The method should provide valuable insights into the complex molecular mechanisms underlying vacuolar sorting of storage proteins.
1) Fuji et al., Plant Cell, 19: 597-609 (2007).
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1 - Kyoto University, Department of Botany, Graduate School of Science, Kitashirakawa-Oiwakecho, Sakyo-ku, Kyoto, Kyoto, 606-8234, Japan
2 - Kyoto University, Department of Botany, Graduate School of Science
3 - Kyoto University, Graduate School of Agriculture
Protein storage vacuole
seed storage protein
vacuolar protein sorting
Presentation Type: Plant Biology Abstract
Location: Exhibit Hall (Northeast, Southwest & Southeast)/Hilton
Date: Sunday, July 8th, 2007
Time: 8:00 AM