Tbeta 4 is not a simple G-actin sequestering protein and interacts with F-actin at high concentration.
Thymosin beta 4 is acknowledged as a major G-actin binding protein maintaining a pool of unassembled actin in motile vertebrate cells. We have examined the function of Tbeta 4 in actin assembly in the high range of concentrations (up to 300 micron) at which Tbeta 4 is found in highly motile blood cells. Tbeta 4 behaves as a simple G-actin sequestering protein only in a range of low concentrations (<20 micron). As the concentration of Tbeta 4 increases, its ability to depolymerize F-actin decreases, due to its interaction with F-actin. The Tbeta 4-actin can be incorporated, in low molar ratios, into F-actin, and can be cross-linked in F-actin using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide. As a result of the copolymerization of actin and Tbeta 4-actin complex, the critical concentration is the sum of free G-actin and Tbeta 4-G-actin concentrations at steady state, and the partial critical concentration of G-actin is decreased by Tbeta 4-G-actin complex. The incorporation of Tbeta 4-actin in F-actin is associated to a structural change of the filaments and eventually leads to their twisting around each other. In conclusion, Tbeta 4 is not a simple passive actin-sequestering agent, and at high concentrations the ability of Tbeta 4-actin to copolymerize with actin reduces the sequestering activity of G-actin-binding proteins. These results question the evaluation of the unassembled actin in motile cells. They account for observations made on living fibroblasts overexpressing beta-thymosins.
Carlier
M F
MF
Laboratoire d'Enzymologie CNRS, Gif-sur-Yvette, France.
Didry
D
D
Erk
I
I
Lepault
J
J
Van Troys
M L
ML
Vandekerckhove
J
J
Perelroizen
I
I
Yin
H
H
Doi
Y
Y
Pantaloni
D
D
D008841:Actin Cytoskeleton / Q000502:physiology / Q000648:ultrastructure; D000199:Actins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D000818:Animals; D002850:Chromatography, Gel; D003285:Contractile Proteins; D005347:Fibroblasts; D007700:Kinetics; D046911:Macromolecular Substances; D008840:Microfilament Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D008854:Microscopy, Electron; D018482:Muscle, Skeletal / Q000378:metabolism; D051304:Profilins; D011817:Rabbits; D011994:Recombinant Proteins / Q000378:metabolism; D013947:Thymosin / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D014162:Transfection; D000099201:Thymosin beta(4)
D000199:Actins; D003285:Contractile Proteins; D046911:Macromolecular Substances; D008840:Microfilament Proteins; D051304:Profilins; D011994:Recombinant Proteins; D013947:Thymosin; D000099201:Thymosin beta(4)