AtFim1 is an actin filament crosslinking protein from Arabidopsis thaliana.
ATFIM1 is a widely expressed gene in Arabidopsis thaliana that encodes a putative actin filament-crosslinking protein, AtFim1, belonging to the fimbrin/plastin class of actin-binding proteins. In this report we have used bacterially expressed AtFim1 and actin isolated from Zea mays pollen to demonstrate that AtFim1 functions as an actin filament-crosslinking protein. AtFim1 binds pollen actin filaments (F-actin) in a calcium-independent manner, with an average dissociation constant (Kd) of 0.55+/-0.21 microM and with a stoichiometry at saturation of 1:4 (mol AtFim1 : mol actin monomer). AtFim1 also crosslinks pollen F-actin by a calcium-independent mechanism, in contrast to crosslinking of plant actin by human T-plastin, a known calcium-sensitive actin-crosslinking protein. When micro-injected at high concentration into living Tradescantia virginiana stamen hair cells, AtFim1 caused cessation of both cytoplasmic streaming and transvacuolar strand dynamics within 2-4 min. Using the 'nuclear displacement assay' as a measure of the integrity of the actin cytoskeleton in living stamen hair cells, we demonstrated that AtFim1 protects actin filaments in these cells from Z. mays profilin (ZmPRO5)-induced depolymerization, in a dose-dependent manner. The apparent ability of AtFim1 to protect actin filaments in vivo from profilin-mediated depolymerization was confirmed by in vitro sedimentation assays. Our results indicate that AtFim1 is a calcium-independent, actin filament-crosslinking protein that interacts with the actin cytoskeleton in living plant cells.
Kovar
D R
DR
Department of Biological Sciences, Purdue University, West Lafayette, IN 47907-1392, USA.
Staiger
C J
CJ
Weaver
E A
EA
McCurdy
D W
DW
D000199:Actins / Q000378:metabolism; D017360:Arabidopsis / Q000235:genetics / Q000378:metabolism*; D029681:Arabidopsis Proteins; D001667:Binding, Competitive; D002118:Calcium / Q000494:pharmacology; D003432:Cross-Linking Reagents; D004274:DNA, Recombinant; D008840:Microfilament Proteins / Q000378:metabolism*; D059828:Plant Cells; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D010944:Plants / Q000187:drug effects / Q000378:metabolism; D011058:Pollen / Q000737:chemistry; D011485:Protein Binding / Q000187:drug effects
C110898:ATFIM1 protein, Arabidopsis; D000199:Actins; D029681:Arabidopsis Proteins; D003432:Cross-Linking Reagents; D004274:DNA, Recombinant; D008840:Microfilament Proteins; D010940:Plant Proteins; D002118:Calcium