Nuclear localization of Arabidopsis HD-Zip IV transcription factor GLABRA2 is driven by importin α.

IF 5.6 2区 生物学 Q1 PLANT SCIENCES Journal of Experimental Botany Pub Date : 2024-10-30 DOI:10.1093/jxb/erae326
Bilal Ahmad, Ruben Lerma-Reyes, Thiya Mukherjee, Hieu V Nguyen, Audra L Weber, Emily E Cummings, Waltraud X Schulze, Jeffrey R Comer, Kathrin Schrick
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Abstract

GLABRA2 (GL2), a class IV homeodomain leucine-zipper (HD-Zip IV) transcription factor from Arabidopsis, is a developmental regulator of specialized cell types in the epidermis. GL2 contains a monopartite nuclear localization sequence (NLS) that is conserved in most HD-Zip IV members across the plants. We demonstrate that NLS mutations affect nuclear transport and result in a loss-of-function phenotypes. NLS fusions to enhanced yellow fluorescent protein (EYFP) show that it is sufficient for nuclear localization in roots and trichomes. Despite partial overlap of the NLS with the homeodomain, genetic dissection indicates that nuclear localization and DNA binding are separable functions. Affinity purification of GL2 from plants followed by MS-based proteomics identified importin α (IMPα) isoforms as potential GL2 interactors. NLS structural prediction and molecular docking studies with IMPα-3 revealed major interacting residues. Cytosolic yeast two-hybrid assays and co-immunoprecipitation experiments with recombinant proteins verified NLS-dependent interactions between GL2 and several IMPα isoforms. IMPα triple mutants (impα-1,2,3) exhibit abnormal trichome formation and defects in GL2 nuclear localization in trichomes, consistent with tissue-specific and redundant functions of IMPα isoforms. Taken together, our findings provide mechanistic evidence for IMPα-dependent nuclear localization of GL2 in Arabidopsis, a process that is critical for cell type differentiation of the epidermis.

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拟南芥 HD-Zip IV 转录因子 GLABRA 的核定位由导入素 α 驱动。
GLABRA2(GL2)是拟南芥中的第四类同源结构域亮氨酸-拉链(HD-Zip IV)转录因子(TF),是表皮中特化细胞类型的发育调节因子。GL2 包含一个单部分核定位序列(NLS),该序列在大多数 HD-Zip IV 植物成员中都是保守的。我们证明,NLS突变会影响核运输并导致功能缺失表型。NLS 与 EYFP 的融合表明,它足以在根和毛状体中进行核定位。尽管 NLS 与同源结构域部分重叠,但基因分析表明核定位和 DNA 结合是可分离的功能。对植物中的 GL2 进行亲和纯化,然后进行基于 MS 的蛋白质组学研究,发现导入蛋白 α(IMPα)异构体是潜在的 GL2 相互作用体。与 IMPα-3 的 NLS 结构预测和分子对接研究揭示了主要的相互作用残基。细胞质酵母双杂交实验和重组蛋白的共免疫沉淀实验验证了 GL2 与几种 IMPα 异构体之间的 NLS 依赖性相互作用。IMPα 三重突变体(impα-1,2,3)表现出毛状体形成异常和 GL2 在毛状体中核定位缺陷,这与 IMPα 同工酶的组织特异性和冗余功能一致。总之,我们的研究结果为拟南芥中 GL2 依赖于 IMPα 的核定位提供了机理证据,这一过程对表皮的细胞类型分化至关重要。
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来源期刊
Journal of Experimental Botany
Journal of Experimental Botany 生物-植物科学
CiteScore
12.30
自引率
4.30%
发文量
450
审稿时长
1.9 months
期刊介绍: The Journal of Experimental Botany publishes high-quality primary research and review papers in the plant sciences. These papers cover a range of disciplines from molecular and cellular physiology and biochemistry through whole plant physiology to community physiology. Full-length primary papers should contribute to our understanding of how plants develop and function, and should provide new insights into biological processes. The journal will not publish purely descriptive papers or papers that report a well-known process in a species in which the process has not been identified previously. Articles should be concise and generally limited to 10 printed pages.
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