In vivoCRISPR screening identifies Fli1 as a transcriptional safeguard that restrains effector CD8 T cell differentiation during infection and cancer

Abstract

Improving effector activity of antigen specific T cells is a major goal in cancer immunotherapy. Despite the identification of several effector T cell (T EFF )-driving transcription factors (TF), the transcriptional coordination of T EFF biology remains poorly understood. We developed an in vivo T cell CRISPR screening platform and identified a novel mechanism restraining T EFF biology through the ETS family TF, Fli1. Genetic deletion of Fli1 enhanced T EFF responses without compromising memory or exhaustion precursors. Fli1 restrained T EFF lineage differentiation by binding to cis-regulatory elements of effector-associated genes. Loss of Fli1 increased chromatin accessibility at ETS:RUNX motifs allowing more efficient Runx3-driven T EFF biology. CD8 T cells lacking Fli1 provided substantially better protection against multiple infections and tumors. These data indicate that Fli1 safeguards the developing CD8 T cell transcriptional landscape from excessive ETS:RUNX-driven T EFF cell differentiation. Moreover, genetic deletion of Fli1 improves T EFF differentiation and protective immunity in infections and cancer. Utzschneider et al., 2016; Wu et al., 2016; Zhou et al., 2010) TCF-1 represses (directly or indirectly) T EFF -driving TF such as T-bet and Blimp-1(Tiemessen et al., 2014), and may also have a role in enabling epigenetic changes (Xing et al., 2016). Moreover, a second HMG TF, Tox, is essential for the development of the T EX cell fate by promoting T EX differentiation and repressing of the T EFF lineage differentiation(Alfei et al., 2019; Khan et al., 2019; Scott et al., 2019; Seo et al., 2019; Yao et al., 2019). Tox reprograms the open chromatin landscape, inducing global T EX -specific epigenetic changes and repressing chromatin accessibility associated with T EFF differentiation, in part by recruiting epigenetic modifiers including the lysine acetyltransferase Kat7 (Khan et al., 2019). Despite this work, mechanisms that safeguard against commitment to T EFF differentiation remain poorly understood. Such information could be of considerable utility for immunotherapies focused on enhancing anti-tumor or antiviral activity. However, whereas inactivating pathways like TCF-1 or Tox that would de-repress the entire program of T EFF differentiation are of interest, such approaches result in terminal T EFF and may have limited therapeutic benefit because such cells cannot sustain durable responses. Thus, the discovery of mechanisms that selectively de-repress key aspects of T EFF differentiation, particularly those involved in control of numerical expansion and/or protective immunity would be of considerable interest.Here, we used in vivo CRISPR-Cas9 screening in antigen-specific CD8 T cells responding to acute or chronic viral infection to identify key regulators of T EFF and T EX . In particular, we were interested in identifying genes that resulted by gain-of-function, improving T EFF differentiation (i.e. an "Up" screen (Kaelin, 2017)). The CRISPR-Cas9 system has been used to interrogate the cancer-imm...

Type I Interferon Transcriptional Network Regulates Expression of Coinhibitory Receptors in Human T cells

“…These data indicate that T cell metabolic activation, cell cycle regulation, and transcriptional activation are promoted by IFN-β treatment 29,30 . Interestingly, FLI1, which is novel as a IFN-I downstream TF and was recently reported to control effector response in T cells 31 , also dominantly regulated the early transcriptional wave. In the intermediate regulatory network, MYC , MAF, IRF1, AFF1, ATF3, and TBX21 were among the most dominant up-regulated TFs for this transcriptional wave.…”

“…These data indicate that T cell metabolic activation, cell cycle regulation, and transcriptional activation are promoted by IFNβ treatment 29,30 . Interestingly, FLI1, which is novel as a IFN-I downstream TF and was recently reported to control effector response in T cells 31 To further study the relationships between the DETFs, we generated hierarchical backbone networks in order to represent their relationships (Figure 4b, bottom). Interestingly, the top TFs in all transcriptional time waves were down-regulated in response to IFN-β, while TFs lower in the network hierarchy were more up-regulated.…”

Longitudinal single cell transcriptional and epigenetic mapping of effector, memory, and exhausted CD8 T cells reveals shared biological circuits across distinct cell fates

“…6o ), suggesting a prominent role for TCR signaling in shaping the Pre-Exh epigenetic landscape and/or TCR-dependent TFs operating in this ACR landscape. In contrast, MP were enriched in accessibility for ETS family TFs, including motifs for Fli1, a TF that may help restrain CD8 T cell activation (43). Altogether, these data reveal distinct paths of Tcf7 -expressing cells early during acutely-resolved versus chronic infection and identify different biological modules that can be present in TCF1+ “stem” or “progenitor”-like CD8 T cells.…”

Type I Interferon Transcriptional Network Regulates Expression of Coinhibitory Receptors in Human T cells

“…These data indicate that T cell metabolic activation, cell cycle regulation, and transcriptional activation are promoted by IFN-b treatment 29,30 . Interestingly, FLI1, which is novel as a IFN-I downstream TF and was recently reported to control effector response in T cells 31 , also dominantly regulated the early transcriptional wave. In the intermediate regulatory network, MYC, MAF, IRF1, AFF1, ATF3, and TBX21 were among the most dominant up-regulated TFs for this transcriptional wave.…”