ASGSB 1999 Annual Meeting Abstracts

THE ACTIN CYTOSKELETON MAY CONTROL THE POLAR DISTRIBUTION OF AN AUXIN TRANSPORT PROTEIN. G.K. Muday¹, S.R. Brady¹, S.Hu1, J. Butler¹, M.W. Dixon¹, S.J. Roux², and G.B. Clark². ¹Dept of Biology, Wake Forest University, Winston-Salem, NC ²Dept of Botany, University of Texas, Austin

The actin cytoskeleton is a filamentous network containing cross-linked polymers of actin filaments (F-actin) assembled from globular actin (G-actin) monomers, and a myriad of associated proteins. The actin cytoskeleton is required for many cellular processes including localization of membrane proteins. One membrane protein complex that may use the actin cytoskeleton for its polar distribution is the auxin efflux carrier. In stems, this protein complex is localized to the basal plasma membrane and thereby controls the polarity of auxin movement. The auxin efflux carrier contains a regulatory subunit that binds inhibitors that block auxin transport, such as naphthylphthalamic acid (NPA). Previous experiments have demonstrated that the NPA binding protein remains with the cytoskeletal during detergent extraction and partitions with actin during in vitro depolymerization and repolymerization experiments. The purpose of this study was to more directly examine the interaction between the NPA binding protein and actin filaments using F-actin affinity chromatography. Zucchini hypocotyl G-actin was purified to electrophoretic homogeneity and was shown to be native and competent for polymerization to actin filaments. Purified plasma membranes were treated to release actin associated proteins, and the resulting extracts were applied to G-actin, F-actin and BSA affinity columns. Two abundant polypeptides eluted from the F-actin column and cross-reacted with antiserum against pea annexins. NPA binding activity was specifically eluted in a single peak from the F-actin column, but was not retained by a BSA column. Drugs that alter the state of actin polymerization also decreased the amount of polar auxin transport. The function of the association of the NPA binding protein with actin filaments may be to localize the auxin efflux carrier complex in the basal plasma membrane and thereby control the polarity of auxin transport.

(Supported by NASA: NAG2-1023 and the NSCORT in Plant Biology at NC State University).

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