Investigation of FIN219 gene expression by promoter fusion assay
Date Issued
2004
Date
2004
Author(s)
Chung, Shu-Hsing
DOI
zh-TW
Abstract
fin219 (far-red insensitive 219) mutant is derived from the screening of extragenic suppressors of the cop1-6 mutant in Arabidopsis. That abnormal DNA methylation was found in the promoter region of the fin219 mutant results in longer hypocotyl phenotype in far-red (FR) light than that in wild type. FIN219 belongs to a GH3-like gene family, is induced rapidly by auxin, and is involved in phytochrome A-mediated light signaling. However, its actual function and role in plant development and light signal transductions are not clear yet. So, the objective of my thesis is to study the promoter activity, gene expression patterns and possible regulatory mechanism of the FIN219 gene by transcriptional fusion with GUS.
According to the results of GUS staining carried out on different developmental stages of the T2 transgenic plants containing the P219::GUS fusion construct, FIN219 gene was highly expressed in the germinating seeds, the hydathod of cotyledons, the base of trichomes, the emerging point of lateral roots, and reproductive tissues, especially in the stigma, pollens as well as the funiculus. All these GUS staining patterns indicated that FIN219 gene expression was highly correlated to the site of auxin production and also consistent with the result reported previously that FIN219 transcripts were induced by auxin.
Besides, we also utilized the transgenic plants containing the P219::GUS fusion construct to study regulatory mechanisms of FIN219 expression regulated by light. The result from light switch experiments indicated that GUS activity was much reduced after transferring 2 day-old of dark-grown seedlings to FR light, when compared to that of dark-grown seedlings, implying that FR seems to inhibit FIN219 gene expression. Furthermore, this reduction of GUS activity in FR can be reversed by the addition of auxin, jasmonate or methyl jasmonate, suggesting that FIN219 may play an important role to integrate the cross talk between light and hormone signalings.
In addition, the transgenic plants containing the promoter and FIN219 coding region fused with GUS in wild type background exhibit a hypersensitive phenotype in cFR and their adult plants also show partial sterility, which was consistent with the result of FIN219 overexpressor in cFR and white light conditions. On the other hand, when we investigated the endogenous FIN219 expression in these transgenic plants by Western and Northern blot analyses, it was found that there were two endogenous FIN219 transcripts in transgenic plants, but only one in wild type and fin219 mutant. Further, FIN219-GUS fusion protein in transgenic plants was greatly reduced compared with the endogenous FIN219 protein, suggesting that the transgene expression was regulated at translational or posttranslational level, which resulted in the reduction of FIN219 fusion protein and then failed to rescue the fin219 mutant efficiently.
Subjects
光訊息傳遞
FIN219
Type
other
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