Studies on the Mechanism of Regulation of the mRNA Level for a Soybean Storage Protein Subunit by Exogenous l-Methionine

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Abstract
In previous studies (GL Creason et al. 1983 Biochem Biophys Res Commun 117: 658-662; LP Holowach et al. 1984 Plant Physiol 74: 576-583), we have shown that when soybean (Glycine max L. Merrill cv Provar) cotyledons are cultured in medium supplemented with l-methionine, the β-subunit of 7S protein and β-mRNA are absent. We have carried out further studies on the mechanism of the methionine action. In one experiment, cotyledons were cultured for 16 days with or without methionine. After 4 days, some cotyledons were transferred from methionine-supplemented to basal (no methionine) medium and vice versa. In basal medium, β-subunit was detected at 4 days whereas in methionine-supplemented medium, no β-subunit was present. When cotyledons were transferred from basal to methionine-supplemented medium, the β-subunit increased within a 4 day period and then remained constant (on a per cotyledon basis). This result indicated that methionine was not acting by accelerating the degradation of the β-subunit. Four days after transfer from supplemented to basal medium cotyledons contained β-subunit, thus demonstrating that the inhibition was reversible. During this time, the uncombined methionine declined from 7 to 1.5 μmoles methionine per gram fresh weight. When β-mRNA was measured by in vitro translation, functional β-mRNA was absent in tissue that was not accumulating β-subunit. The messenger RNA for the β-subunit had a half-life of about 1 day in the presence of methionine. Hybridization of cotyledon mRNA with cDNA complementary to β-mRNA revealed that the 1700 nucleotide β-mRNA was not present in supplemented cotyledons. Thus, expression of the β-subunit gene is controlled at the level of transcription, RNA processing, or RNA turnover, rather than at the level of translation.