Cell cycle arrest of Saccharomyces cerevisiae in G(1) by the antimitogen alpha-factor is mediated by activation of a signal transduction pathway that results in inhibition of the cyclin-dependent kinase Cdc28-Cln. The Farl protein is required for cell cycle arrest and associates with the Cdc28-Cln complex. The kinase activity of Cdc28-Cln was directly inhibited by Far1 both in vivo and in vitro, thus demonstrating that Far1 acts at the final step in the alpha-factor response pathway by inhibiting a G(1) cyclin-dependent kinase.