Ethylene controls multiple physiological processes in plants, including cell elongation. Consequently, ethylene synthesis is regulated by internal and external signals. We show that a light-entrained circadian clock regulates ethylene release from unstressed, wild-type Arabidopsis (Arabidopsis thaliana) seedlings, with a peak in the mid-subjective day. The circadian clock drives the expression of multiple ACC SYNTHASE genes, resulting in peak RNA levels at the phase of maximal ethylene synthesis. Ethylene production levels are tightly correlated with ACC SYNTHASE 8 steady-state transcript levels. The expression of this gene is controlled by light, by the circadian clock, and by negative feedback regulation through ethylene signaling. In addition, ethylene production is controlled by the TIMING OF CAB EXPRESSION 1 and CIRCADIAN CLOCK ASSOCIATED 1 genes, which are critical for all circadian rhythms yet tested in Arabidopsis. Mutation of ethylene signaling pathways did not alter the phase or period of circadian rhythms. Mutants with altered ethylene production or signaling also retained normal rhythmicity of leaf movement. We conclude that circadian rhythms of ethylene production are not critical for rhythmic growth.Since the discovery of ethylene production in plants in the 1930s, researchers have tried to elucidate mechanisms governing ethylene formation. A major breakthrough was the completion of the enzymatic pathway for ethylene biosynthesis 50 years later (for review, see Yang and Hoffman, 1984). Shortly thereafter, the first genes encoding ethylene biosynthetic enzymes were cloned (Sato and Theologis, 1989;Van Der Straeten et al., 1990;Hamilton et al., 1991;Spanu et al., 1991). With the use of tomato (Lycopersicon esculentum) and especially Arabidopsis (Arabidopsis thaliana) as model plants, molecular biological and genetic analysis has shed light on many physiological processes involving ethylene (Abeles et al., 1992;Somerville and Meyerowitz, 2002). In higher plants, the enzymes for ethylene biosynthesis are encoded by gene families. The members of these families are differentially responsive to various ethylene-inducing factors, including wounding, fruit ripening, pathogen infections, auxins, and cytokinins (for review, see Fluhr and Mattoo, 1996).In Arabidopsis, there are 12 genes in the family of enzymes that produces the ethylene precursor 1-amino-cyclopropane-1-carboxylic acid (ACC), one of which, ACC SYNTHASE 3 (ACS3), is a pseudogene (Yamagami et al., 2003;Tsuchisaka and Theologis, 2004). ACS1 is not functional as an ACS (Liang et al., 1992). ACS10 and ACS12 do not function as ACSs either, but as aminotransferases (Yamagami et al., 2003). Many of the ACS genes are regulated on the transcriptional level. ACS2 transcription in leaves is switched off when tissues mature (Rodrigues-Pousada et al., 1993; in this paper the gene was designated ACS1). ACS4 can be induced by auxins (Abel et al., 1995). ACS5 is regulated by cytokinins that cause stabilization of the protein (Chae et al., 2003). ACS6 is induce...