ACTIN MUTATIONS THAT SHOW SUPPRESSION WITH FIMBRIN MUTATIONS IDENTIFYA LIKELY FIMBRIN-BINDING SITE ON ACTIN

Actin interacts with a large number of different proteins that modulat e its assembly and mediate its functions. One such protein is the yeas t actin-binding protein Sac6p, which is homologous to vertebrate fimbr in (Adams, A. E. M., D. Botstein, and D. G. Drubin. 1991. Nature (Lond .). 354:404-408.). Sac6p was originally identified both genetically (A dams, A. E. M., and D. Botstein. 1989. Genetics. 121:675-683.) by domi nant, reciprocal suppression of a temperature-sensitive yeast actin mu tation (act1-1), as well as biochemically (Drubin, D. G., K. G. Miller , and D. Botstein. 1988. J. Cell Biol. 107: 2551-2561.). To identify t he region on actin that interacts with Sac6p, we have analyzed eight d ifferent act1 mutations that show suppression with sac6 mutant alleles , and have asked whether (a) these mutations occur in a small defined region on the crystal structure of actin; and (b) the mutant actins ar e defective in their interaction with Sac6p in vitro. Sequence analysi s indicates that all of these mutations change residues that cluster i n the small domain of the actin crystal structure, suggesting that thi s region is an important part of the Sac6p-binding domain. Biochemical analysis reveals defects in the ability of several of the mutant acti ns to bind Sac6p, and a reduction in Sac6p-induced cross-linking of mu tant actin filaments. Together, these observations identify a likely s ite of interaction of fimbrin on actin.