Direct proof that the primary site of action of cytochalasin on cell motility processes is actin.
We have previously described the isolation of a mutant KB cell (Cyt 1 mutant) resistant to the cytotoxic effect of cytochalasin B (CB). The Cyt 1 mutant carries an altered form of beta-actin (beta'-actin) and lacks normal beta-actin (Toyama, S., and S. Toyama. 1984. Cell. 37:609-614). Increased resistance of the Cyt 1 mutant to CB in vivo is reflected in altered properties of beta'-actin in vitro (Toyama, S., and S. Toyama. 1988. J. Cell Biol. 107:1499-1504). Here, we show that the mutation in beta-actin is solely responsible for the cytochalasin-resistant phenotype of the Cyt mutant. We have isolated a cDNA clone encoding beta'-actin from Cyt 1 cells. Sequence analysis reveals two mutations in the coding region that substitute two amino acid residues (Val139----Met and Ala295----Asp). Expression of the beta'-actin cDNA confers cytochalasin resistance upon transformed cytochalasin-sensitive KB cells. Levels of resistance to CB in the transformed cell clones correlate well with amounts of beta'-actin polypeptide. Both of the two mutations in beta'-actin are necessary for the high level expression of cytochalasin resistance. Overall, we conclude that the primary site of action of cytochalasin on cell motility processes in vivo is actin.
Ohmori
H
H
Institute for Virus Research, Kyoto University, Japan.
Toyama
S
S
Toyama
S
S
D008841:Actin Cytoskeleton / Q000187:drug effects / Q000648:ultrastructure; D000199:Actins / Q000187:drug effects* / Q000235:genetics; D000595:Amino Acid Sequence; D001483:Base Sequence; D015139:Blotting, Southern; D002462:Cell Membrane / Q000187:drug effects / Q000648:ultrastructure; D002465:Cell Movement / Q000187:drug effects*; D003571:Cytochalasin B / Q000494:pharmacology*; D004247:DNA / Q000235:genetics; D005822:Genetic Vectors; D006801:Humans; D007624:KB Cells; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D009154:Mutation; D010641:Phenotype; D014162:Transfection
D000199:Actins; D003571:Cytochalasin B; D004247:DNA