Computational Biology Institute, The George Washington University, Washington DC, USA.
Department of Biochemistry and Molecular Medicine, The George Washington University, Washington DC, USA.
Int J Parasitol. 2020 Jul;50(8):603-610. doi: 10.1016/j.ijpara.2020.05.003. Epub 2020 Jun 25.
Free-living nematodes respond to variable and unpredictable environmental stimuli whereas parasitic nematodes exist in a more stable host environment. A positive correlation between the presence of environmental stages in the nematode life cycle and an increasing number of G-protein coupled receptors (GPCRs) reflects this difference in free-living and parasitic lifestyles. As hookworm larvae move from the external environment into a host, they detect uncharacterized host components, initiating a signalling cascade that results in the resumption of development and eventual maturation. Previous studies suggest this process is mediated by GPCRs in amphidial neurons. Here we set out to uncover candidate GPCRs required by a hookworm to recognise its host. First, we identified all potential Ancylostoma ceylanicum GPCRs encoded in the genome. We then used life cycle stage-specific RNA-seq data to identify differentially expressed GPCRs between the free-living infective L3 (iL3) and subsequent parasitic stages to identify receptors involved in the transition to parasitism. We reasoned that GPCRs involved in host recognition and developmental activation would be expressed at higher levels in the environmental iL3 stage than in subsequent stages. Our results support the model that a decrease in GPCR diversity occurs as the larvae develop from the free-living iL3 stage to the parasitic L3 (pL3) in the host over 24-72 h. We find that overall GPCR expression and diversity is highest in the iL3 compared with subsequent parasitic stages. By 72 h, there was an approximately 50% decrease in GPCR richness associated with the moult from the pL3 to the L4. Taken together, our data uncover a negative correlation between GPCR diversity and parasitic development in hookworm. Finally, we demonstrate proof of principal that Caenorhabditis elegans can be used as a heterologous system to examine the expression pattern of candidate host signal chemoreceptors (CRs) from hookworm. We observe expression of candidate host signal CRs in C. elegans, demonstrating that C. elegans can be effectively used as a surrogate to identify expressed hookworm genes. We present several preliminary examples of this strategy and confirm a candidate CR as neuronally expressed.
自由生活的线虫对可变和不可预测的环境刺激作出反应，而寄生线虫则存在于更稳定的宿主环境中。线虫生命周期中环境阶段的存在与越来越多的 G 蛋白偶联受体 (GPCR) 之间存在正相关，这反映了自由生活和寄生生活方式的差异。当钩虫幼虫从外部环境进入宿主时，它们会检测到未被识别的宿主成分，启动信号级联反应，导致发育的恢复和最终的成熟。先前的研究表明，这个过程是由 amphidial 神经元中的 GPCR 介导的。在这里，我们着手揭示钩虫识别宿主所需的候选 GPCR。首先，我们鉴定了在基因组中编码的所有潜在的Ancylostoma ceylanicum GPCR。然后，我们使用生命周期阶段特异性 RNA-seq 数据，在自由生活的感染性 L3 (iL3) 和随后的寄生阶段之间鉴定差异表达的 GPCR，以鉴定参与向寄生过渡的受体。我们推断，参与宿主识别和发育激活的 GPCR 在环境中的 iL3 阶段的表达水平会高于随后的阶段。我们的结果支持这样的模型，即随着幼虫从自由生活的 iL3 阶段发育到宿主中的寄生 L3 (pL3)，GPCR 多样性会降低。我们发现，与随后的寄生阶段相比，iL3 中的 GPCR 表达和多样性总体上最高。到 72 小时时，与从 pL3 到 L4 的蜕皮相关的 GPCR 丰富度下降了约 50%。总的来说，我们的数据揭示了钩虫中 GPCR 多样性与寄生发育之间的负相关关系。最后，我们证明了 Caenorhabditis elegans 可以用作异源系统来检查钩虫候选宿主信号化学感受器 (CR) 的表达模式。我们观察到候选宿主信号 CR 在 C. elegans 中的表达，证明 C. elegans 可以有效地用作替代物来鉴定表达的钩虫基因。我们展示了这种策略的几个初步例子，并证实了一个候选 CR 是神经元表达的。
