Heath-Heckman Elizabeth, Yoo Shinja, Winchell Christopher, Pellegrino Maurizio, Angstadt James, Lammardo Veronica B, Bautista Diana, De-Miguel Francisco F, Weisblat David
Department of Molecular & Cell Biology, University of California, Berkeley, Berkeley, CA, USA.
Current address: Department of Integrative Biology, Michigan State University, East Lansing, MI, USA.
BMC Genomics. 2021 Mar 25;22(1):215. doi: 10.1186/s12864-021-07526-0.
While leeches in the genus Hirudo have long been models for neurobiology, the molecular underpinnings of nervous system structure and function in this group remain largely unknown. To begin to bridge this gap, we performed RNASeq on pools of identified neurons of the central nervous system (CNS): sensory T (touch), P (pressure) and N (nociception) neurons; neurosecretory Retzius cells; and ganglia from which these four cell types had been removed.
Bioinformatic analyses identified 3565 putative genes whose expression differed significantly among the samples. These genes clustered into 9 groups which could be associated with one or more of the identified cell types. We verified predicted expression patterns through in situ hybridization on whole CNS ganglia, and found that orthologous genes were for the most part similarly expressed in a divergent leech genus, suggesting evolutionarily conserved roles for these genes. Transcriptional profiling allowed us to identify candidate phenotype-defining genes from expanded gene families. Thus, we identified one of eight hyperpolarization-activated cyclic-nucleotide gated (HCN) channels as a candidate for mediating the prominent sag current in P neurons, and found that one of five inositol triphosphate receptors (IP3Rs), representing a sub-family of IP3Rs absent from vertebrate genomes, is expressed with high specificity in T cells. We also identified one of two piezo genes, two of ~ 65 deg/enac genes, and one of at least 16 transient receptor potential (trp) genes as prime candidates for involvement in sensory transduction in the three distinct classes of leech mechanosensory neurons.
Our study defines distinct transcriptional profiles for four different neuronal types within the leech CNS, in addition to providing a second ganglionic transcriptome for the species. From these data we identified five gene families that may facilitate the sensory capabilities of these neurons, thus laying the basis for future work leveraging the strengths of the leech system to investigate the molecular processes underlying and linking mechanosensation, cell type specification, and behavior.
虽然医蛭属的水蛭长期以来一直是神经生物学的模型，但该类群中神经系统结构和功能的分子基础在很大程度上仍不为人知。为了开始弥合这一差距，我们对中枢神经系统（CNS）中已鉴定的神经元群体进行了RNA测序：感觉T（触觉）、P（压力）和N（伤害感受）神经元；神经分泌性雷丘斯细胞；以及去除了这四种细胞类型的神经节。
生物信息学分析确定了3565个推定基因，其表达在样本之间存在显著差异。这些基因聚集成9组，可与一种或多种已鉴定的细胞类型相关联。我们通过对整个CNS神经节进行原位杂交验证了预测的表达模式，发现直系同源基因在不同的水蛭属中大部分以相似的方式表达，表明这些基因具有进化上保守的作用。转录谱分析使我们能够从扩展的基因家族中鉴定出候选的表型定义基因。因此，我们确定八个超极化激活的环核苷酸门控（HCN）通道之一是介导P神经元中显著下陷电流的候选基因，并发现五个肌醇三磷酸受体（IP3R）之一，代表脊椎动物基因组中不存在的IP3R亚家族，在T细胞中具有高度特异性表达。我们还确定了两个piezo基因之一、约65个deg/enac基因中的两个以及至少16个瞬时受体电位（trp）基因之一是参与三种不同类别的水蛭机械感觉神经元感觉转导的主要候选基因。
我们的研究定义了水蛭CNS内四种不同神经元类型的独特转录谱，此外还为该物种提供了第二个神经节转录组。从这些数据中，我们确定了五个基因家族，它们可能促进这些神经元的感觉能力，从而为未来利用水蛭系统的优势来研究机械感觉、细胞类型特化和行为背后及相互联系的分子过程奠定基础。
