Biochemical Characterization of Soluble Acid and Alkaline Invertases from Shoots of Etiolated Pea Seedlings

Soluble invertase was purified from pea （Pisum sativum L.） by sequential procedures entailing ammonium sulfate precipitation, DEAE-Sepharose column, Con-A- and Green 19-Sepharose affinity columns, hydroxyapatite column, ultra-filtration, and Sephacryl 300 gel filtration. The purified soluble acid （SAC） and alkaline （SALK） invertases had a pH optimum of 5.3 and 7.3, respectively. The temperature optimum of two invertases was 37 ℃. The effects of various concentrations of Tris-HCI, HgCI2, and CuSO4 on the activities of the two purified enzymes were examined. Tris-HCI and HgCI2 did not affect SAC activity, whereas 10 mM Tris-HCI and 0.05 mM HgCI2 inhibited SALK activity by about 50%. SAC and SALK were inhibited by 4.8 mM and 0.6 mM CuSO4 by 50%, respectively. The enzymes display typical hyperbolic saturation kinetics for sucrose hydrolysis. The Kms of SAC and SALK were determined to be 1.8 and 38.6 mM, respectively. The molecular masses of SAC shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting were 22 kDa and 45 kDa. The molecular mass of SALK was 30 kDa. Iso-electric points of the SAC and SALK were estimated to be about pH 7.0 and pH 5.7, respectively.

Overexpression of OsPIL15, a phytochrome-interacting factor-like protein gene, represses etiolated seedling growth in rice

Phytochrome-interacting factors （PIFs） regulate an array of developmental responses ranging from seed germi- nation to vegetational architecture in Arabidopsis. However, information regarding the functions of the PIF family in monocots has not been widely reported. Here, we investigate the roles of OsPIL15, a member of the rice （Oryza sativa L. cv. Nipponbare） PIF family, in regulating seedling growth. OsPIL15 encodes a basic helix-loop-helix factor localized in the nucleus. OsPIL15-OX seedlings exhibit an exaggerated shorter above- ground part and undeveloped root system relative to wild-type seedlings, suggesting that OsPIL15 represses seedling growth in the dark. Microarray analysis combined with gene ontology analysis revealed that OsPIL15 represses a set of genes involved in auxin pathways and cell wall organization or biogenesis. Given the important roles of the auxin pathway and cell wall properties in controlling plant growth, we speculate that OsPIL15 represses seedling growth likely by regulating the auxin pathway and suppressing cell wall organization in etiolated rice seedlings. Additionally, exposure to red light or far-red light relieved growth retardation and promoted seedling elongation in the OsPIL15-OX lines, despite higher levels of OsPIL15 transcripts under red light and far-red light than in the dark. These results suggest that light regulation of OsPIL15 expres- sion is probably involved in photomorphogenesis in rice.