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MBC in Press, published online ahead of print February 9, 2005
Mol. Biol. Cell 10.1091/mbc.E04-07-0562

A more recent version of this article appeared on April 1, 2005
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Submitted on July 7, 2004
Revised on January 28, 2005
Accepted on January 30, 2005

Ectopic Expression of an Activated RAC in Arabidopsis Disrupts Membrane Cycling

Daria Bloch,* Meirav Lavy,* Yael Efrat,* Idan Efroni,* Keren Bracha-Drori,* Mohamad Abu-Abied,{dagger} Einat Sadot,{dagger} and Shaul Yalovsky*

*Department of Plant Sciences, Tel Aviv University, Tel Aviv 69978, Israel; {dagger}Department of Ornamental Horticulture, Volcani Center, Bet Dagan 50250, Israel

Monitoring Editor: Anne Ridley

Rho GTPases regulate the actin cytoskeleton, exocytosis, endocytosis and other signaling cascades. Rhos are subdivided into 4 subfamilies designated Rho, Racs, Cdc42, and a plant specific group designated RACs/Rops. This research demonstrates that ectopic expression of a constitutive active Arabidopsis RAC, AtRAC10, disrupts actin cytoskeleton organization and membrane cycling. We created transgenic plants expressing either wild-type or constitutive active AtRAC10 fused to the green fluorescent protein. The activated AtRAC10 induced deformation of root hairs and leaf epidermal cells and was primarily localized in TritonX-100 insoluble fractions of the plasma membrane. Actin cytoskeleton reorganization was revealed by creating double transgenic plants expressing activated AtRAC10 and the actin marker YFP-Talin. Plants were further analyzed by membrane staining with FM4-64 under different treatments including the protein trafficking inhibitor brefeldin A or the actin depolymeryzing agents latrunculin-B and cytochalasin-D. Following drug treatments, activated AtRAC10 did not accumulate in BFA compartments, but rather reduced their number and colocalized with FM4-64 labeled membranes in large intracellular vesicles. Furthermore, endocytosis was compromised in root hairs of activated AtRAC10 transgenic plants. FM4-64 was endocytosed in nontransgenic root hairs treated with the actin stabilizing drug jasplakinolide. These findings suggest complex regulation of membrane cycling by plant RACs.

Address correspondence to: Shaul Yalovsky (shauly{at}tauex.tau.ac.il)






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