Summary
High-speed supernatant obtained from the lysate of cultured BHK cells has been chromatographed on Sepharose-4B, DEAE-cellulose and hydroxyapatite columns, and a fraction has been identified with characteristics similar to an actin depolymerizing factor (ADF), a small protein previously isolated from embryonic chick brain. Using a rabbit antibody against the chick brain protein, two immunoreactive forms were identified: a 19 kDa form co-migrating in SDS-polyacrylamide gels with embryonic chick brain ADF, and a 20 kDa form. The two species could be separated on a hydroxyapatite or green A dye matrix columns and only the 20 kDa protein was active when assayed for effects on pyrene-G-actin assembly. It enhanced the rate of F-actin assembly, but only after an initial lag phase, and decreased the final proportion of actin in filamentous form. These effects were calcium-independent. Actin depolymerizing factor constituted at least 0.5% of the total protein in the cytoplasmic fraction. A Triton extract of plasma membrane-enriched fraction from BHK cells was fractionated on a Sepharose-4B column and again, a fraction was found which had an ADF-like activity and also contained the two immuno-cross-reactive forms, 19 kDa and 20 kDa. These results suggest a novel regulation of the microfilament system in eukaryotic cells via the control of the ADF activity.
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Research performed while on sabbatical leave from the Department of Biochemistry, Colorado State University, Fort Collins, CO 80523, U.S.A.
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Koffer, A., Edgar, A.J. & Bamburg, J.R. Identification of two species of actin depolymerizing factor in cultures of BHK cells. J Muscle Res Cell Motil 9, 320–328 (1988). https://doi.org/10.1007/BF01773875
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DOI: https://doi.org/10.1007/BF01773875