OsPRA1 plays a significant role in targeting of OsRab7 into the tonoplast via the prevacuolar compartment during vacuolar trafficking in plant cells.
In yeast and mammals, the Yip/PRA1 family of proteins has been reported to facilitate the delivery of Rab GTPases to the membrane by dissociating the Rab-GDI complex during vesicle trafficking. Recently, we identified OsPRA1, a plant Yip/PRA1 homolog, as an OsRab7-interacting protein that localizes to the prevacuolar compartment, which suggests that it plays a role in vacuolar trafficking of plant cells. Here, we show that OsPRA1 is essential for vacuolar trafficking and that it has molecular properties that are typical of the Yip/PRA1 family of proteins. A trafficking assay using Arabidopsis protoplasts showed that the point mutant OsPRA1((Y94A)) strongly inhibits the vacuolar trafficking of cargo proteins, but has no inhibitory effect on the plasma membrane trafficking of H(+)-ATPase-GFP, suggesting its specific involvement in vacuolar trafficking. Moreover, OsPRA1 was shown to be an integral membrane protein, suggesting that its two hydrophobic domains may mediate membrane integration, and its cytoplasmic N- and C-terminal regions were found to be important for binding to OsRab7. OsPRA1 also interacted with OsVamp3, implying its involvement in vesicle fusion. Finally, we used a yeast expression system to show that OsPRA1 opposes OsGDI2 activity and facilitates the delivery of OsRab7 to the target membrane. Taken together, our results support strongly that OsPRA1 targets OsRab7 to the tonoplast during vacuolar trafficking.
Heo
Jae Bok
JB
Department of Biology, Pennsylvania State University, University Park, PA 16802, USA.
Bang
Woo Young
WY
Kim
Se Won
SW
Hwang
Sung Min
SM
Son
Young Sim
YS
Im
Chak Han
CH
Acharya
Biswa R
BR
Kim
Chul Wook
CW
Kim
Sam Woong
SW
Lee
Byung Hyun
BH
Bahk
Jeong Dong
JD
D017360:Arabidopsis / Q000235:genetics / Q000378:metabolism; D001692:Biological Transport / Q000235:genetics / Q000502:physiology*; D002462:Cell Membrane / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000378:metabolism; D008969:Molecular Sequence Data; D012275:Oryza / Q000235:genetics / Q000378:metabolism*; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism*; D006180:Proton-Translocating ATPases / Q000378:metabolism; D020798:Two-Hybrid System Techniques; D014617:Vacuoles / Q000378:metabolism*
D010940:Plant Proteins; D049452:Green Fluorescent Proteins; D006180:Proton-Translocating ATPases