阿拉伯芥中GH3-like基因家族的功能性研究與GH3-10在光訊息調控中的功能探討

Abstract

Abstract Light is one of the most important environmental factors that affect the growth of plants. Three types of photoreceptors identified so far are phytochromes, cryptochromes, and phototropins. Each photoreceptor uses different downstream signal transduction components to transmit light signals. One of these components, identified as FIN219 gene, is involved in phytochrome A-mediated signaling pathway and encodes a GH3-like protein with 575 amino acids. Intriguingly, there are at least 20 GH3-like genes present in Arabidopsis genome. Sequence comparison among them indicates that there is a 35%~47% identity at amino acid level, in which some conserved regions are also found. However, their functions remain unknown. To understand further to see if other GH3-like genes in addition to FIN219 are also involved in light signaling pathway, T-DNA insertion lines and other light signaling mutants in Arabidopsis were used as samples to analyze the possible phenotypes and regulatory relationship under dark, blue, far-red, and red light. The results showed that the hypocotyl lengths of all T-DNA insertion lines are significantly longer than wild type under red and far-red light conditions. This indicated that most of the GH3-like genes are involved in red- and far-red-mediated inhibition of hypocotyl elongation in Arabidopsis. Besides, RT-PCR was ultilized to investigate the expression patterns of different GH3-like genes in different light signaling mutants under dark, white and far-red light. The results indicated that many photoreceptor mutants influence the expression of GH3-like genes. In particular, FIN219, GH3-7 and GH3-10 may be involved in more than one light signaling pathways. COP1 can repress gene expression of some of the GH3-like genes under far-red light. Furthermore, since it was reported that one of the GH3-like genes, GH3-10, can be up-regulated under far-red light and this induction was abolished in phyA mutant. GH3-10 was also most close to FIN219 at amino acid level, leading us to investigate the actual function of GH3-10 in light signaling. The transgenic plants overexpressing and antisensing the GH3-10 in wild type displayed a longer hypocotyl phenotype in red and far-red light than that in wild-type. This implies that GH3-10 participates in both red and far-red light signal transduction pathways and may form a functional complex with a dosage effect to carry out its physiological role in Arabidopsis. Meanwhile, it was also found that GH3-10 was functional only in the presence of phyA and phyB. However, the transgenic plants overexpressing the N-terminal 232 amino-acid portion of GH3-10 did not show any obvious phenotype under all light conditions, implying that the C-terminus of GH3-10 may play more important roles to trigger light signaling. Expression of GH3-10 protein in onion cells using particle bombardment showed that it was localized in cytosol and this subcellular localization is light-independent. At last, the crossing test between GH3-10 and FIN219 suggested that both are nonallelic noncomplementational genetic relationship and may interact with each other or work in parallel with each other in phyA-mediated signal transduction pathway.