Immunohistochemical observation of indole-3-acetic acid at the IAA synthetic maize coleoptile tips
Immunohistochemical observation of indole-3-acetic acid at the IAA synthetic maize coleoptile tips
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DOI：
10.4161/psb.6.12.18080
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发表时间：
2011-12
影响因子：
2.9
通讯作者：
Takeshi Nishimura;K. Toyooka;Mayuko Sato;S. Matsumoto;M. M. Lucas-M.;M. Strnad;F. Baluška;T. Koshiba
中科院分区：
文献类型：
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作者：
Takeshi Nishimura;K. Toyooka;Mayuko Sato;S. Matsumoto;M. M. Lucas-M.;M. Strnad;F. Baluška;T. Koshiba
为了研究玉米胚芽鞘中吲哚乙酸（IAA）及其合成细胞的分布，我们用抗吲哚乙酸单克隆抗体建立了玉米胚芽鞘中IAA的免疫组织化学方法。我们首先通过比较内源性游离和缀合的IAA的量与从IAA抗体获得的IAA信号来确认抗体的特异性。内源性IAA的耗竭显示免疫信号强度的相应降低，并且观察到对IAA相关化合物（包括色氨酸、吲哚-3-乙酰胺和缀合的IAA）的可忽略的交叉反应性。免疫定位结果表明，IAA信号是强烈的，在约1毫米的区域和外表皮约0.5毫米的区域从1-N-萘酞酸处理的胚芽鞘的顶部。相比之下，5-甲基色氨酸处理后，外表皮中的IAA免疫信号几乎消失。免疫金标记的IAA与抗-IAA-N-多克隆抗体在外表皮细胞显示游离IAA的细胞质定位，但没有在细胞壁或液泡。这些结果表明，玉米胚芽鞘顶端0- 2.0mm区域内的IAA由色氨酸合成，其中0.5mm顶端的外表皮细胞是IAA合成最活跃的细胞。
To investigate the distribution of IAA (indole-3-acetic acid) and the IAA synthetic cells in maize coleoptiles, we established immunohistochemistry of IAA using an anti-IAA-C-monoclonal antibody. We first confirmed the specificity of the antibody by comparing the amounts of endogenous free and conjugated IAA to the IAA signal obtained from the IAA antibody. Depletion of endogenous IAA showed a corresponding decrease in immuno-signal intensity and negligible cross-reactivity against IAA-related compounds, including tryptophan, indole-3-acetamide, and conjugated-IAA was observed. Immunolocalization showed that the IAA signal was intense in the approximately 1 mm region and the outer epidermis at the approximately 0.5 mm region from the top of coleoptiles treated with 1-N-naphthylphthalamic acid. By contrast, the IAA immuno-signal in the outer epidermis almost disappeared after 5-methyl-tryptophan treatment. Immunogold labeling of IAA with an anti-IAA-N-polyclonal antibody in the outer-epidermal cells showed cytoplasmic localization of free-IAA, but none in cell walls or vacuoles. These findings indicated that IAA is synthesized in the 0–2.0 mm region of maize coleoptile tips from Trp, in which the outer-epidermal cells of the 0.5 mm tip are the most active IAA synthetic cells.