Single-cell RNA-sequencing identifies the developmental trajectory of C-Myc-dependent NK1.1\u2212 T-bet+ intraepithelial lymphocyte precursors

Jonas F Hummel,Jonas Fixemer,Patrice Zeis,Yakup Tanriver,Sebastian J Arnold,Christian Schachtrup,Philip Konrad,Dominic Grün,Karolina Ebert

doi:10.1038/s41385-019-0220-y

Abstract

Natural intraepithelial lymphocytes (IELs) are thymus-derived adaptive immune cells, which are important contributors to intestinal immune homeostasis. Similar to other innate-like T cells, they are induced in the thymus through high-avidity interaction that would otherwise lead to clonal deletion in conventional CD4 and CD8 T cells. By applying single-cell RNA-sequencing (scRNA-seq) on a heterogeneous population of thymic CD4−CD8αβ−TCRαβ+NK1.1− IEL precursors (NK1.1− IELPs), we define a developmental trajectory that can be tracked based on the sequential expression of CD122 and T-bet. Moreover, we identify the Id proteins Id2 and Id3 as a novel regulator of IELP development and show that all NK1.1− IELPs progress through a PD-1 stage that precedes the induction of T-bet. The transition from PD-1 to T-bet is regulated by the transcription factor C-Myc, which has far reaching effects on cell cycle, energy metabolism, and the translational machinery during IELP development. In summary, our results provide a high-resolution molecular framework for thymic IEL development of NK1.1− IELPs and deepen our understanding of this still elusive cell type.

Highlights

Intraepithelial lymphocytes (IELs) are an important component of the epithelial barrier that constitutes the boundary between the body and the environment
Single-cell transcriptomics reveal thymic differentiation pathway of NK1.1− IELPs Historically, IELPs have been defined as DN TCRαβ+NK1.1− thymocytes.[22]
We mapped the phenotype of our index sorted cells individually onto those clusters to test whether the development of IELPs can be tracked based on the expression of CD122 and T-bet

Summary

Introduction

Intraepithelial lymphocytes (IELs) are an important component of the epithelial barrier that constitutes the boundary between the body and the environment. The intestine consists of various innate and adaptive immune cells that execute specific functions to maintain epithelial integrity and intestinal immune homeostasis.[1] Here adaptive immune cells can be broadly divided into induced and natural IELs.[2] Natural IELs comprise both T cell receptor (TCR) γδ+ and TCRαβ+ T cells, which lack the classical coreceptor CD4 or CD8αβ (double negative (DN)) but instead largely express the homodimer CD8αα. Natural TCRαβ+ IELs are selected and fate-determined in the thymus through high affinity TCR interaction with self-peptide major compatibility complex (MHC) in a process termed “agonist selection.”[3,4] This pathway is not unique to natural TCRαβ+ IELs as other lineages, e.g., invariant natural killer T (NKT) cells and thymic regulatory T cells, require strong TCR interactions for their development.[5,6] In contrast, such strong interaction would result in the clonal deletion of conventional CD4 and CD8αβ single-positive (SP) T cells, which are selected by low affinity TCR stimulation.[7]

Methods

Results

Conclusion