Studies on the Mechanism of Regulation of the mRNA Level

In previous studies (GL Creason et al. 1983 Biochem Biophys Res Commun 117: 658-662; LP Holowach et al. 1984 Plant Physiol 74: 576583), we have shown that when soybean (Glycine max L. Merrill cv Provar) cotyledons are cultured in medium supplemented with L-methionine, the ,B-subunit of 7S protein and fl-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, ,-subunit was detected at 4 days whereas in methioninesupplemented medium, no ,B-subunit was present. When cotyledons were transferred from basal to methionine-supplemented medium, the ,-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 ,-subunit. Four days after transfer from supplemented to basal medium cotyledons contained ,B-subunit, thus demonstrating that the inhibition was reversible. During this time, the uncombined methionine declined from 7 to 1.5 Mmoles methionine per gram fresh weight. When fl-mRNA was measured by in vitro translation, functional ,l-mRNA was absent in tissue that was not accumulating ,Bsubunit. The messenger RNA for the ,8-subunit had a half-life of about 1 day in the presence of methionine. Hybridization of cotyledon mRNA with cDNA complementary to fl-mRNA revealed that the 1700 nucleotide ,B-mRNA was not present in supplemented cotyledons. Thus, expression of the li-subunit gene is controlled at the level of transcription, RNA