Table_2_Variegated Transcription of the WC1 Hybrid PRR/Co-Receptor Genes by Individual γδ T Cells and Correlation With Pathogen Responsiveness.DOCX

γδ T cells have broad reactivity and actively participate in protective immunity against tumors and infectious disease-causing organisms. In γδ-high species such as ruminants and other artiodactyls many γδ T cells bear the lineage-specific markers known as WC1. WC1 molecules are scavenger receptors coded for by a multigenic array and are closely related to SCART found on murine γδ T cells and CD163 found on a variety of cells. We have previously shown that WC1 molecules are hybrid pattern recognition receptors thereby binding pathogens as well as signaling co-receptors for the γδ T cell receptor. WC1+ γδ T cells can be divided into two major subpopulations differentiated by the WC1 genes they express and the pathogens to which they respond. Therefore, we hypothesize that optimal γδ T cell responses are contingent on pathogen binding to WC1 molecules, especially since we have shown that silencing WC1 results in an inability of γδ T cells from primed animals to respond to the pathogen Leptospira, a model system we have employed extensively. Despite this knowledge about the crucial role WC1 plays in γδ T cell biology, the pattern of WC1 gene expression by individual γδ T cells was not known but is critical to devise methods to engage γδ T cells for responses to specific pathogens. To address this gap, we generated 78 γδ T cell clones. qRT-PCR evaluation showed that approximately 75% of the clones had one to three WC1 genes transcribed but up to six per cell occurred. The co-transcription of WC1 genes by clones showed many combinations and some WC1 genes were transcribed by both subpopulations although there were differences in the overall pattern of WC1 genes transcription. Despite this overlap, Leptospira-responsive WC1+ memory γδ T cell clones were shown to have a significantly higher propensity to express WC1 molecules that are known to bind to the pathogen.

γδ T细胞（γδ T cell）具有广泛的反应性，可积极参与对抗肿瘤与致病微生物的保护性免疫应答。在反刍动物及其他偶蹄动物这类γδ T细胞富集的物种中，多数γδ T细胞表达名为WC1的谱系特异性标志物。WC1分子是由多基因阵列编码的清道夫受体，与小鼠γδ T细胞表面的SCART以及多种细胞表面的CD163具有密切的同源关系。我们此前的研究证实，WC1分子属于混合型模式识别受体，既可结合病原体，同时可作为γδ T细胞受体的信号共受体。表达WC1的γδ T细胞可根据其所表达的WC1基因以及所响应的病原体，分为两大主要亚群。据此我们提出假说：最优的γδ T细胞应答依赖于病原体与WC1分子的结合，尤其因为我们此前证实，沉默WC1基因会使经免疫致敏的动物的γδ T细胞无法响应钩端螺旋体（Leptospira）——这也是我们已广泛应用的模型系统。尽管已知WC1在γδ T细胞生物学中发挥关键作用，但此前尚不明确单个γδ T细胞的WC1基因表达模式，而该信息对于设计策略以激活γδ T细胞对抗特定病原体至关重要。为填补这一研究空白，我们构建了78株γδ T细胞克隆。实时定量逆转录聚合酶链反应（quantitative real-time reverse transcription polymerase chain reaction，qRT-PCR）检测显示，约75%的克隆可转录1至3个WC1基因，而单个细胞最多可转录6个WC1基因。各克隆的WC1基因共转录存在多种组合模式，部分WC1基因在两个亚群中均有转录，尽管整体的WC1基因转录模式存在差异。尽管存在这种表达重叠，响应钩端螺旋体的WC1阳性记忆性γδ T细胞克隆，其表达已知可结合该病原体的WC1分子的概率显著更高。