RNA-seq analysis of the Innate Lymphoid Cells type 2 (ILC2) or their precursors (ILC2p ) sorted from WT and Tgfbr2-/- mice

Purpose: The molecular pathways underlying the development of innate lymphoid cells (ILCs) are mostly unknown. Results: TGF-β signaling programs the development of ILC2s from their progenitors. Specifically, the deficiency of TGF-β receptor II in bone marrow progenitors results in inefficient development of ILC2s, but not ILC1s or ILC3s. We found that hundreds of genes were changed in Tgfbr2-/- ILC2s compared to WT ILC2s (FDR < 0.1 and Fold-Change >1.5). Some ILC2-associated genes including Gata3, Il1rl1 (ST2), Il-5, Atxn1 (Sca1), and Ccr4 were decreased in Tgfbr2-/- ILC2s, according to RNA-seq analysis Conclusions: TGF-β upregulates the expression of the IL-33 receptor gene Il1rl1 (encoding IL-1 receptor-like 1, also known as ST2) in ILC2p and common helper-like innate lymphoid progenitors (CHILP), at least partially through the MEK-dependent pathway. These findings identify a function of TGF-β in the development of ILC2s from their progenitors. Lamina propria CD45+Lin–CD127+Sca–1+CD25+KIRG1+ ILC2 cells were sorted from tamoxifen or oil-treated Tgfbr2f/fER-Cre+ mice. Bone marrow CD45.2+Lin–CD127+α4β7+CD25+ ILC2p cells were sorted from Tgfbr2-/-/45.1 or Control/45.1 chimeras. RNA was extracted using RNeasy Plus Micro Kit (QIAGEN), and RNA-seq experiments were performed.

研究背景：固有淋巴细胞（innate lymphoid cells, ILCs）发育的分子通路目前尚不完全明确。
实验结果：转化生长因子β（TGF-β）信号通路可调控祖细胞向2型固有淋巴细胞（ILC2）的发育过程。具体而言，骨髓祖细胞中转化生长因子βⅡ型受体（TGF-β receptor Ⅱ）的缺陷会导致2型固有淋巴细胞发育效率低下，但对1型固有淋巴细胞（ILC1）和3型固有淋巴细胞（ILC3）的发育无显著影响。转录组测序（RNA sequencing, RNA-seq）分析显示，与野生型（wild type, WT）ILC2相比，Tgfbr2-/- ILC2中有数百个基因的表达发生显著改变（错误发现率false discovery rate, FDR < 0.1，折叠变化fold change, FC >1.5）。部分与2型固有淋巴细胞相关的基因，包括GATA结合蛋白3（Gata3）、白细胞介素1受体样1基因（Il1rl1，又名ST2）、白细胞介素5（Il-5）、共济失调蛋白1（Atxn1，又名Sca1）以及趋化因子受体4（Ccr4），在Tgfbr2-/- ILC2中的表达水平显著下调。
研究结论：转化生长因子β（TGF-β）可在2型固有淋巴细胞祖细胞（ILC2p）与共同辅助样固有淋巴祖细胞（common helper-like innate lymphoid progenitors, CHILP）中上调白细胞介素33受体基因Il1rl1（编码白细胞介素1受体样1，又名ST2）的表达，这一过程至少部分依赖于丝裂原活化蛋白激酶激酶（mitogen-activated protein kinase kinase, MEK）信号通路。本研究明确了转化生长因子β在祖细胞向2型固有淋巴细胞发育过程中的功能。
实验方法：本研究从他莫昔芬或油剂处理的Tgfbr2f/fER-Cre+小鼠中分选得到固有层CD45+Lin–CD127+Sca-1+CD25+KIRG1+ ILC2细胞；从Tgfbr2-/-/45.1或对照/45.1嵌合小鼠中分选得到骨髓CD45.2+Lin–CD127+α4β7+CD25+ ILC2p细胞。采用RNeasy Plus微型试剂盒（QIAGEN）提取RNA，并开展转录组测序实验。