单细胞 RNA 测序揭示了 G 蛋白信号转导调节剂 1 作为调节性 T 细胞在肝癌中的作用靶点。

ScRNA-seq revealed targeting regulator of G protein signaling 1 to mediate regulatory T cells in Hepatocellular carcinoma.

机构信息

Institute of Translational Medicine, Hunan Provincial People's Hospital, First Affiliated Hospital of Hunan Normal University, Changsha, Hunan, China.

Department of Anatomy and Cell Biology, Carver College of Medicine, University of Iowa, Iowa City, IA, USA.

出版信息

Cancer Biomark. 2023;36(4):299-311. doi: 10.3233/CBM-220226.

DOI:10.3233/CBM-220226

PMID:36938729

Abstract

BACKGROUND

Regulatory T cells (Tregs) are central to determine immune response, thus targeting Tregs for immunotherapy is a promising strategy against tumor development and metastasis.

OBJECTIVES

The objective of this study was to identify genes for targeting Tregs to improve the outcome of HCC.

METHODS

We integrated expression data from different samples to remove batch effects and further applied embedding function in Scanpy to conduct sub-clustering of CD4+ T cells in HCC for each of two independent scRNA-seq data. The activity of transcription factors (TFs) was inferred by DoRothEA. Gene expression network analysis was performed in WGCNA R package. We finally used R packages (survminer and survival) to conduct survival analysis. Multiplex immunofluorescence analysis was performed to validate the result from bioinformatic analyses.

RESULTS

We found that regulator of G protein signaling 1 (RGS1) expression was significantly elevated in Tregs compared to other CD4+ T cells in two independent public scRNA-seq datasets, and increased RGS1 predicted inferior clinical outcome of HCC patients. Multiplex immunofluorescence analysis supported that the higher expression of RGS1 in HCC Tregs in tumor tissue compared to it in adjacent tissue. Moreover, RGS1 expression in Tregs was positively correlated with the expression of marker genes of Tregs, C-X-C chemokine receptor 4 (CXCR4), and three CXCR4-dependent genes in both scRNA-seq and bulk RNA-seq data. We further identified that these three genes were selectively expressed in Tregs as compared to other CD4+ T cells. The activities of two transcription factors, recombination signal binding protein for immunoglobulin kappa J region (RBPJ) and yin yang 1 (YY1), were significantly different in HCC Tregs with RGS1 high and RGS1 low.

CONCLUSIONS

Our findings suggested that RGS1 may regulate Treg function possibly through CXCR4 signaling and RGS1 could be a potential target to improve responses for immunotherapy in HCC.

摘要

背景

调节性 T 细胞（Tregs）是决定免疫反应的关键，因此针对 Tregs 进行免疫治疗是对抗肿瘤发展和转移的一种有前途的策略。

目的

本研究的目的是确定针对 Tregs 的靶向基因，以改善 HCC 的治疗效果。

方法

我们整合了来自不同样本的表达数据，以去除批次效应，并进一步应用 Scanpy 中的嵌入功能对来自两个独立的 scRNA-seq 数据的 HCC 中的 CD4+T 细胞进行亚群聚类。通过 DoRothEA 推断转录因子（TFs）的活性。在 WGCNA R 包中进行基因表达网络分析。最后，我们使用 R 包（survminer 和 survival）进行生存分析。进行多重免疫荧光分析以验证生物信息学分析的结果。

结果

我们发现，在两个独立的公共 scRNA-seq 数据集，与其他 CD4+T 细胞相比，G 蛋白信号调节因子 1（RGS1）在 Tregs 中的表达显著升高，并且 RGS1 的增加预示着 HCC 患者的临床预后较差。多重免疫荧光分析支持在肿瘤组织中 HCC Tregs 中的 RGS1 表达高于相邻组织。此外，在 scRNA-seq 和批量 RNA-seq 数据中，Tregs 中 RGS1 的表达与 Tregs 的标记基因 C-X-C 趋化因子受体 4（CXCR4）和三个 CXCR4 依赖性基因的表达呈正相关。我们进一步发现，与其他 CD4+T 细胞相比，这三个基因在 Tregs 中特异性表达。在 RGS1 高和 RGS1 低的 HCC Tregs 中，两个转录因子，免疫球蛋白 k 区重组信号结合蛋白（RBPJ）和阴阳 1（YY1）的活性有显著差异。