Autocatalytic C2H4 biosynthesis in preclimacteric apple fruit (Malus domestica Borkh. `Golden Delicious') was prevented by storage in atmospheres of 20% CO2-enriched air (17% O2 + 63% N2) or 0.25% O2 (balance N2). In preclimacteric fruit, both treatments inhibited C2H2 biosynthesis by suppressing expression of ACC synthase (ACC-S) at the mRNA level. ACC oxidase (ACC-O) mRNA abundance and in vitro enzyme activity also were impaired by these treatments. However, the conversion of ACC to C2H4 never became the rate limiting step in C2H4 biosynthesis. C2H4 biosynthesis also was effectively inhibited in climacteric apple fruit kept in air + 20% CO2 or 0.25% O2. Climacteric apples also exhibited suppressed expression of ACC-S at the mRNA level, while ACC-O transcript abundance, enzyme activity, and protein abundance were reduced only slightly. ACC-S is the key regulatory enzyme of C2H4 biosynthesis and is the major site at which elevated CO2 and reduced O2 atmospheres inhibit C2H4 biosynthesis, irrespective of fruit physiological maturity. Chemical names used: 1-aminocyclopropane-1-carboxcylic acid (ACC).