title: Tropomyosin-dependent filament formation by a polymerization-defective mutant yeast actin (V266G,L267G).
abstract: A major function of tropomyosin (TPM) in nonmuscle cells may be stabilization of F-actin by binding longitudinally along the actin filament axis. However, no clear evidence exists in vitro that TPM can significantly affect the critical concentration of actin. We previously made a polymerization-defective mutant actin, GG (V266G, L267G). This actin will not polymerize alone at 25 degrees C but will in the presence of phalloidin or beryllium fluoride. With beryllium fluoride, but not phalloidin, this polymerization rescue is cold-sensitive. We show here that GG-actin polymerizability was restored by cardiac tropomyosin and yeast TPM1 and TPM2 at 25 degrees C with rescue efficiency inversely proportional to TPM length (TPM2 > TPM1 > cardiac tropomyosin), indicating the importance of the ends in polymerization rescue. In the presence of TPM, the apparent critical concentration of actin is 5.5 microm, 10-15-fold higher than that of wild type actin but well below that of the GG-actin alone (>20 microm). Non N-acetylated TPMs did not rescue GG-actin polymerization. The TPMs did not prevent cold-induced depolymerization of GG F-actin. TPM-dependent GG-actin polymerization did not occur at temperatures below 20 degrees C. Polymerization rescue may depend initially on the capture of unstable GG-F-actin oligomers by the TPM, resulting in the strengthening of actin monomer-monomer contacts along the filament axis.
authors.affiliation: Department of Biochemistry, University of Iowa College of Medicine, Iowa City, Iowa 52242, USA.
authors.forename: K K
authors.initials: KK
authors.lastname: Wen
authors.forename: B
authors.initials: B
authors.lastname: Kuang
authors.forename: P A
authors.initials: PA
authors.lastname: Rubenstein
mesh_terms: D000199:Actins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001704:Biopolymers / Q000737:chemistry / Q000378:metabolism*; D008274:Magnesium / Q000378:metabolism; D008854:Microscopy, Electron; D009154:Mutation; D014335:Tropomyosin / Q000378:metabolism*
chemical_list: D000199:Actins; D001704:Biopolymers; D014335:Tropomyosin; D008274:Magnesium