Department of Plant Pathology and Institute of Plant Breeding, Genetics, and Genomics, University of Georgia, Athens, GA 30602, U.S.A.
Complex Carbohydrate Research Center, University of Georgia, Athens, GA 30602, U.S.A.
Mol Plant Microbe Interact. 2023 Jun;36(6):372-380. doi: 10.1094/MPMI-11-22-0232-R. Epub 2023 Jul 27.
Root-knot nematodes (RKN) ( spp.) represent one of the most damaging groups of plant-parasitic nematodes. They secrete effector proteins through a protrusible stylet to manipulate host cells for their benefit. Stylet-secreted effector proteins are produced within specialized secretory esophageal gland cells, one dorsal gland (DG) and two subventral glands (SvG), whose activity differ throughout the nematode life cycle. Previous gland transcriptomic profiling studies identified dozens of candidate RKN effectors but were focused on the juvenile stages of the nematode, when the SvGs are most active. We developed a new approach to enrich for the active DGs of adult female RKN for RNA and protein extraction. Female heads were manually cut from the body, and a combination of sonication and vortexing was used to dislodge contents inside the heads. DG-enriched fractions were collected by filtering, using cell strainers. Comparative transcriptome profiling of pre-parasitic second-stage juveniles, female heads, and DG-enriched samples was conducted using RNA sequencing. Application of an established effector mining pipeline led to the identification of 83 candidate effector genes upregulated in DG-enriched samples of adult females that code for proteins with a predicted signal peptide but lack transmembrane domains or homology to proteins in the free-living nematode . In situ hybridization resulted in the identification of 14 new DG-specific candidate effectors expressed in adult females. Taken together, we have identified novel candidate effector genes that may have essential roles during later stages of parasitism. [Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.
根结线虫（RKN）（ spp.）是最具破坏性的植物寄生线虫之一。它们通过可伸出的口针分泌效应蛋白，以操纵宿主细胞为自身谋取利益。口针分泌的效应蛋白是在专门的分泌食管腺细胞中产生的，包括一个背腺（DG）和两个亚腹腺（SvG），它们的活性在整个线虫生命周期中有所不同。先前的腺体转录组分析研究确定了数十种候选 RKN 效应子，但这些研究主要集中在幼年线虫阶段，此时 SvG 最为活跃。我们开发了一种新的方法来富集成熟雌性 RKN 的活性 DG 进行 RNA 和蛋白质提取。手动从虫体上切下雌性头部，然后使用超声处理和涡旋混合将头部内部的内容物分离出来。通过过滤，使用细胞筛收集富含 DG 的部分。使用 RNA 测序对未寄生的第二龄幼虫、雌性头部和富含 DG 的样品进行比较转录组分析。应用已建立的效应子挖掘管道，鉴定出 83 个候选效应子基因在富含 DG 的成年雌性样本中上调，这些基因编码的蛋白质具有预测的信号肽，但缺乏跨膜结构域或与自由生活线虫中的蛋白质同源。原位杂交鉴定出 14 个在成年雌性中表达的新的 DG 特异性候选效应子。总之，我们已经确定了新的候选效应子基因，它们可能在寄生后期阶段发挥重要作用。[公式：见文本] 版权所有 © 2023 作者。这是一个 CC BY-NC-ND 4.0 国际许可下的开放获取文章。
