The impact of severe drought (water withholding) on protein synthesis during cotton (Gossypium hirsutum L. var. DPL 51) seed development was examined, using potted plants of identical age grown under glass. Flowers at anthesis were tagged and labelled as to branch and boll position. Embryos used for radiolabeling were excised from ovules in the maturation period of seed development, i.e. 20 to 40 days post anthesis (DPA). Storage of protein during the maturation stage involves gene activation and translation of a number of stage-specific, messenger RNA transcripts. In this study we tested the hypothesis that severe drought directly affects the developmental program for seed ripening. Our experimental plan was designed to determine at what age during the maturation period developing embryos are most sensitive to severe drought stress. Tagged bolls from the same branch and boll position and equivalent chronological age were used for radiolabeling. The impact of 10 days of severe drought on incorporation of methionine into protein of cotyledons and axes was compared to incorporation pattern for controls throughout the maturation period, with samples differing by 5 day intervals, i.e. 20, 25...45 DPA for controls and 20-30, 25-35 ... 40-50 DPA for drought-stressed plants. Loss in soil water potential to a value greater than -15 bars, as measured by gypsum sensor blocks in a medial pot position, occurred within two days, following water withholding. Six to eight ovules from three separate bolls, one each from separate plants, were dissected and separate cotyledon and axis tissues placed in microfuge tubes containing 50 µci Of (35)S methionine in 250 µl water for 45 min labeling at room temperature. This was followed by addition of 250 µl of 25 mM Tris buffer, pH 6.8, containing 2.5% SDS and 3 M urea and placement in a -70 C freezer. Tissue-free aliquots (containing 15,000 cpm) from homogenized and centrifuged samples were separated on 10% SDS gels. After staining with Comassie blue, gels were dried and exposed to x-ray film for 2-3 weeks at minus 70 C. Developed autoradiographs were scanned with a gel scanner and silver densities in each lane recorded. Separate data files for each control and drought-stressed sample were compared to determine differences in appearance or absence of bands for all labelled polypeptides in each lane for a corresponding control or drought-stressed sample of equivalent age. The abundant storage protein bands at 52 and 48 kD provided markers for end product synthesis which were easily identified in autoradiographs of control and drought-stressed embryos from boils aged 25 to 45 DPA. A significant result of 10 days of drought stress was the absence of distinct radiolabeled bands for high molecular weight (72 and 68 kD) precursors of storage proteins in embryos (cotyledons and axes) from bolls being 35 DPA or older at the beginning of drought treatment. Separate analysis of silver-stained gels containing equivalent amounts per lane of the SDS-solubilized protein used for autoradiographs provided corroborating data for the absence of 72 and 68 kD storage protein precursors in the drought-stressed embryos (35 DPA or older). Bolls for the 35-45 and 40-50 DPA drought samples were cracked or fully opened at time of collection for the experiment. We interpret the arrestment of incorporation of (35)S methionine into high molecular weight polypeptides for storage proteins in the drought-stressed samples as possibly resulting in an earlier and major transition of the normal program of the developing seed from storage of protein reserves to that of the post abscission and desiccation programs coincident with earlier boll opening.