A Chimeric Arabinogalactan Protein Promotes Somatic Embryogenesis in Cotton Cell Culture
A Chimeric Arabinogalactan Protein Promotes Somatic Embryogenesis in Cotton Cell Culture
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DOI：
10.1104/pp.112.203075
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发表时间：
2012-10-01
期刊：
影响因子：
7.4
通讯作者：
Clarke, Adrienne Elizabeth
中科院分区：
文献类型：
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作者：
Poon, Simon;Heath, Robyn Louise;Clarke, Adrienne Elizabeth
阿拉伯半乳聚糖蛋白（AGPs）是一类植物胞外蛋白聚糖，参与植物生长发育的许多方面，包括体外体细胞胚胎发生（SE）。我们发现，棉花（陆地棉）愈伤组织进行SE产生的特定的AGP，当这些AGP被分离出来，并纳入组织培养基，棉花SE促进。当AGPs部分或完全去糖基化时，SE促进活性并没有减弱。通过反相高效液相色谱法分离的AGP的测试显示，SE促进活性驻留在疏水部分。我们克隆了全长互补DNA（棉花类植物蓝蛋白阿拉伯半乳聚糖蛋白1 [GhPLA1]），其编码活性组分中AGP的蛋白质骨架。它具有包含氨基末端信号序列、植物蓝蛋白样结构域、AGP样结构域和疏水羧基末端结构域的嵌合结构。在烟草（烟草）细胞中重组生产GhPLA1使我们能够纯化和分析一个单一的糖基化AGP，并证明这种嵌合AGP促进棉花SE。此外，从GhPLA1，这是细菌产生的非糖基化的植物蓝蛋白样结构域，也促进SE，表明糖基化的AGP结构域是不必要的体外活性。
Arabinogalactan proteins (AGPs) are a family of extracellular plant proteoglycans implicated in many aspects of plant growth and development, including in vitro somatic embryogenesis (SE). We found that specific AGPs were produced by cotton (Gossypium hirsutum) calli undergoing SE and that when these AGPs were isolated and incorporated into tissue culture medium, cotton SE was promoted. When the AGPs were partly or fully deglycosylated, SE-promoting activity was not diminished. Testing of AGPs separated by reverse-phase high-performance liquid chromatography revealed that the SE-promoting activity resided in a hydrophobic fraction. We cloned a full-length complementary DNA (cotton PHYTOCYANINLIKE ARABINOGALACTAN-PROTEIN1 [GhPLA1]) that encoded the protein backbone of an AGP in the active fraction. It has a chimeric structure comprising an amino-terminal signal sequence, a phytocyanin-like domain, an AGP-like domain, and a hydrophobic carboxyl-terminal domain. Recombinant production of GhPLA1 in tobacco (Nicotiana tabacum) cells enabled us to purify and analyze a single glycosylated AGP and to demonstrate that this chimeric AGP promotes cotton SE. Furthermore, the nonglycosylated phytocyanin-like domain from GhPLA1, which was bacterially produced, also promoted SE, indicating that the glycosylated AGP domain was unnecessary for in vitro activity.