Analysis of signal transduction pathway during seed germination

点击翻译按钮获取中文摘要

1.The expression pattern of the putative alpha subunit of a heterotrimeric G protein in rice seeds was investigated during germination and flowering stages. The amount of the transcript for the alpha subunit was observed to increase rapidly from 2nd to 5th day after germination, followed by a gradual increase during growth. The cells expressing the mRNA for RGA1 were analyzed by in situ hybridization with digoxigenin-labeled RNA probes prepared with parts of RGA1 cDNA as the templates. The mRNA was detected in the root tip, scutellum, phloem but not in the root cap and xylem during germination. It was also detected in rachilla, nucellus, and paricape during rapening stage. These results suggest that RGA1 may take part in cell division and/or material transport.2.A recombinant protein with the cDNA that encodes the putative alpha subunit of G protein was synthesized in Escherichia coli. The purified recombinant protein (rGricealpha) showed an apparent molecular mass of 45 kDa and bound guanosine 5'-(3-O-thio) triphosphate with an apparent association constant (k_<app>) of 0.36. The rGricealpha was ADP-ribosylated by activated cholera toxin. Monoclonal antibodies raised against rGricealpha reacted with a 45-kDa polypeptide localized in the plasma membrane of rice seedlings. A 45-kDa polypeptide in the plasma membrane, as well as rGricealpha, was ADP-ribosylated by activated cholera toxin. The properties observed are similar to those reported for the alpha subunits of heterotrimeric G proteins in animals, suggesting that the putative alpha subunit is truly the alpha subunit itself.