Huq E, Tepperman J M, Quail P H
Department of Plant and Microbial Biology, University of California, Berkeley 94720, USA.
Proc Natl Acad Sci U S A. 2000 Aug 15;97(17):9789-94. doi: 10.1073/pnas.170283997.
In a genetic screen of available T-DNA-mutagenized Arabidopsis populations for loci potentially involved in phytochrome (phy) signaling, we identified a mutant that displayed reduced seedling deetiolation under continuous red light, but little if any change in responsiveness to continuous far-red light. This behavior suggests disruption of phyB, but not phyA signaling. We have cloned the mutant locus by using the T-DNA insertion and found that the disrupted gene is identical to the recently described GIGANTEA (GI) gene identified as being involved in control of flowering time. The encoded GI polypeptide has no sequence similarity to any known proteins in the database. However, by using beta-glucuronidase-GI and green fluorescent protein-GI fusion constructs, we have shown that GI is constitutively targeted to the nucleus in transient transfection assays. Optical sectioning by using the green fluorescent protein-GI fusion protein showed green fluorescence throughout the nucleoplasm. Thus, contrary to previous computer-based predictions that GI would be an integral plasma membrane-localized polypeptide, the data here indicate that it is a nucleoplasmically localized protein. This result is consistent with the proposed role in phyB signaling, given recent evidence that early phy signaling events are nuclear localized.
在对现有的经T-DNA诱变的拟南芥群体进行的基因筛选中,以寻找可能参与光敏色素(phy)信号传导的基因座,我们鉴定出一个突变体,该突变体在持续红光下幼苗去黄化作用减弱,但对持续远红光的反应性几乎没有变化(如果有变化也是极小的)。这种表现表明phyB信号传导受到破坏,但phyA信号传导未受影响。我们利用T-DNA插入克隆了该突变基因座,发现被破坏的基因与最近描述的、被确定参与开花时间控制的GIGANTEA(GI)基因相同。编码的GI多肽与数据库中任何已知蛋白质均无序列相似性。然而,通过使用β-葡萄糖醛酸酶-GI和绿色荧光蛋白-GI融合构建体,我们发现在瞬时转染试验中,GI始终定位于细胞核。利用绿色荧光蛋白-GI融合蛋白进行光学切片显示,整个核质中均有绿色荧光。因此,与之前基于计算机的预测(即GI将是一种整合的质膜定位多肽)相反,此处的数据表明它是一种定位于核质的蛋白质。鉴于最近有证据表明早期phy信号传导事件发生在细胞核中,这一结果与GI在phyB信号传导中所提出的作用是一致的。