The seed storage protein P-phaseolin of the common bean (Pbaseolus vulgaris L.) was expressed i n the endosperm of transgenic rice (Oryza sativa L.) plants. The 5.1 -or 1.8-kb promoter fragment of the rice seed storage protein glutelin C t l gene was fused transcriptionally to either the genomic or cDNA coding sequence of the P-phaseolin gene. The highest quantity of phaseolin estimated by enzyme-linked immunosorbent assay was 4.0% of the total endosperm protein in the transgenic rice seeds. The phaseolin trait was segregated as a single dominant trait with a positive gene dosage effect and was stably inherited through three successive generations. 60th phaseolin genomic and cDNA coding sequences were used to synthesize four isoforms of mature phaseolin protein with apparent molecular masses of 51, 48, 47, and 45 kD. Enzyme deglycosylation experiments indicated that the 51 -kD form contains high-mannose N-glycans; the 48-and 47-kD forms have further modified N-glycans; and the 45-kD form is a nonglycosylated protein. lmmunolabeling studies using light and electron microscopy demonstrated that phaseolin accumulates primarily in the vacuolar type-I1 protein bodies located at the periphery of the endosperm near the aleurone layer. We discuss the implications of these results on nutritional improvement of rice grains.

The deep-sea yeast Cryptococcus liquefaciens strain N6 possesses high superoxide dismutase (SOD) activity and a high tolerance toward metal ions. To clarify the relationship between metal tolerance and SOD activity in this strain, we cloned the Cu/Zn SOD gene. This gene (Cl-SOD1) consists of 471 bp encoding 157 amino acids; the associated protein had 59.9-76.7% identity with Cu/Zn SOD proteins of other yeast species. The highest identity corresponded to Cryptococcus gattii (76.7%). Cl-SOD1 expression in the sod1 mutant of Saccharomyces cerevisiae revealed that this SOD protein was functional in S. cerevisiae. The Cl-SOD1 protein possessed approximately fourfold greater activity than S. cerevisiae SOD1 (Sc-SOD1) at 30 degrees C. The amount of Cl-SOD1 mRNA in strain N6 increased in the presence of copper ion. However, the level of this transcript was not dependent on an increase in copper ion concentration and did not correlate well with changes in the amount of Cu/Zn SOD protein. This result suggests that strain N6 possesses other Cu/Zn SOD genes induced in a manner different from Cl-SOD1 as found in Candida albicans, or that the Cl-SOD1 gene undergoes posttranscriptional regulation upon increase of copper ion.