Laboratory of Plant Nematology, South China Agricultural University, Guangzhou 510642, China.
Exp Parasitol. 2013 Sep;135(1):15-23. doi: 10.1016/j.exppara.2013.05.012. Epub 2013 Jun 5.
This study describes the molecular and biochemical characterization of the β-1,4-endoglucanase gene (Mj-eng-3) from the root knot nematode Meloidogyne javanica. A 2156-bp genomic DNA sequence of Mj-eng-3 containing six introns was obtained. Mj-eng-3 was localized in the subventral esophageal glands of M. javanica juveniles by in situ hybridization. Real-time RT-PCR assay showed that the highest transcriptional level of Mj-eng-3 occurred in pre-parasitic second-stage juveniles, and this high expression persisted in parasitic second-stage juveniles. Recombinant MJ-ENG-3 degraded carboxymethylcellulose and optimum enzyme activity at 40°C and pH 8.0. EDTA, Mg(2+), Mn(2+), Ca(2+), Co(2+), and Cu(2+) did not affect the activity of MJ-ENG-3; however, Zn(2+) and Fe(2+) inhibited MJ-ENG-3 enzyme activity. In planta Mj-eng-3 RNAi assay displayed a reduction in the number of nematodes and galls in transgenic tobacco roots. These results suggested that MJ-ENG-3 could be secreted by M. javanica to degrade the cellulose of plant cell walls to facilitate its entry and migration during the early stages of parasitism.
本研究描述了根结线虫（Meloidogyne javanica）β-1,4-内切葡聚糖酶基因（Mj-eng-3）的分子和生化特性。获得了包含 6 个内含子的 Mj-eng-3 的 2156bp 基因组 DNA 序列。通过原位杂交，将 Mj-eng-3 定位于 M. javanica 幼虫的亚腹食管腺中。实时 RT-PCR 分析表明，Mj-eng-3 的转录水平在预寄生的 2 龄幼虫中最高，这种高表达在寄生的 2 龄幼虫中持续存在。重组 MJ-ENG-3 可降解羧甲基纤维素，最适酶活性在 40°C 和 pH8.0 时。EDTA、Mg(2+)、Mn(2+)、Ca(2+)、Co(2+)和 Cu(2+)不影响 MJ-ENG-3 的活性；然而，Zn(2+)和 Fe(2+)抑制了 MJ-ENG-3 的酶活性。在植物体内，Mj-eng-3 RNAi 分析显示，转基烟草根中的线虫数量和虫瘿减少。这些结果表明，MJ-ENG-3 可能由 M. javanica 分泌，以降解植物细胞壁的纤维素，以促进其在寄生早期阶段的进入和迁移。
