T cells play a vital role in both vaccine immunity and tumor immunotherapy. However, whether it is vaccine development or tumor immunotherapy, there are still many deficiencies, so sorting out the molecular mechanisms of T cells' immune memory and secondary immune response is crucial for the subsequent development of vaccines with higher protection levels and more perfect tumor immunotherapy.
On February 21, 2022, the team of Professor Haihui Xue of Hackensack University Medical Center and the team of Professor Chongzhi Zang of the University of Virginia jointly published a conference entitled Tcf1 preprograms the mobilization of Nature Immunology Glycolysis in central memory CD8+ T cells during recall responses research paper. The transcription factor Tcf1 is clearly illustrated by preprogramming the mobilization of glycolysis in CD8+ TCM cells to meet the energy requirements associated with rapid proliferation of secondary response CD8+ T cells.
Screenshot of the paper
TCM cells functioning relies on the transcription factor Tcf1
TCM cells (CD8+ central memory T cells) are a subset of CD8+ memory T cells that differentiate from CD8+ T cells. CD8+ TCM cells are abundant in lymphatic organs and have a strong proliferative capacity during secondary stimulation. Tcf1 (transcription factor T cell factor 1, encoded by the gene Tcf7) is abundantly expressed in T cells, and its regulatory role in T cells is the basis for the body to produce memory T cells and immune memories.
In the study, the team first clarified the dependence of secondary immunity on Tcf1 in TCM cells. By comparing the secondary response of Tcm cells in Tcf7 knockout mice with wild-type mice, the researchers found that Tcf7 gene-missing mice had significantly fewer secondary response TCM cells than wild-type mice, and could not effectively remove pathogens.
Tcf7 gene-deficient mice had significantly fewer secondary response TCM cells than wild-type mice and were unable to effectively clear pathogens | References[1]
In addition, the team found 623 "Tcf1-dependent, recall-induced genes," which are genes that are normally induced in secondary immunity in wild-type mice and abnormal in knockout mice by comparing transcriptome data after secondary immunity occurred in Tcf7 mice and wild-type mice. Annotated, most of the genes were found to be associated with cell cycles, DNA replication, and ribosomal protein/protein translation, which corresponded to the rapid proliferation of CD8+ cells in secondary responses, while other genes were associated with mitochondria, oxidative phosphorylation, and glycolysis, which was consistent with the large amount of energy required for secondary immune responses.
Most genes are associated with cell cycles, DNA replication, and ribosomal protein/protein translation, while other genes are associated with mitochondria, oxidative phosphorylation, and glycolysis | References[1]
Further, with the help of CHIP-seq technology, the researchers sequenced the binding sites with Tcf1 in chromatin and found that Tcf1 mostly bound to key gene loci in CD8+ TCM cells at rest. It is speculated that Tcf1 predetermines transcriptional activation of these key genes at the time of secondary stimulation.
Innovative ideas and high-standard technology
In order to better understand the regulatory mechanism of Tcf1 in the secondary response of CD8+ TCM cells, the research team designed and used a series of high-standard experimental techniques. First, the researchers used ATAC-seq to analyze the effect of Tcf1 deletion on chromatin accessibility. The analysis found that the change in chromosomal accessibility caused by stimulation was smaller in the knockout group compared with the wild type, and the specific manifestation was that Tcf1 deletion led to a lower degree of chromatin openness in the stimulated state.
In the stimulated state, Tcf1 deletion results in a lower degree of chromatin opening | References[1]
Combined with the transcriptome data, the researchers found that this change is closely related to the transcriptome, as follows: the dominant chrAcc (chromatin accessibility) site in the wild type can act as a stabilizing enhancer in the resting CD8+ TCM cells, while those co-enriched with Tcf1 chrAcc sites can promote the co-activation of key target genes in the secondary response of CD8+ TCM cells.
According to previous research, Tcf1 can regulate the three-dimensional genome in the initial CD8+ T cells, so the researchers obtained three-dimensional structural information of chromatin through Hi-C technology (High-through chromosome conformation capture) and correlated with the previously discovered chrAcc site for correlation analysis. The results show that Tcf1-mediated chromatin interaction provides an ordered structural environment that allows stable enhancers to spatially approach target genes in resting TCM cells, thereby promoting the activation of these elements and their associated target genes in secondary responses, i.e., pre-programmed expression of related genes.
To determine the importance of this pre-programmed gene for secondary response function, the researchers first analyzed through GSEA (gene set enrichment analysis) and found that glycolysis-related genes are among the most enriched genes. Analysis combining the transcriptome with the chrAcc locus found that these glycolysis-related genes were not expressed differentially in resting wild-type and Tcf1-deficient CD8+ TCM cells, but showed induced expression abnormalities and reduced chromatin openness in stimulated Tcf1-deficient cells compared to wild-type CD8+ TCM cells, suggesting that Tcf1 was pre-programmed for induced expression of glycolysis genes in secondary-response CD8+ TCM cells.
Next, the researchers designed an experiment to see if there was a Tcf1 deficiency on glycolysis in CD8+ TCM cells by performing ECAR (Seahorse Extracellular Flux Analyzer to track extracellular acidification rate) and OCR (oxygen consumption) on infected cells rate) analysis to obtain glycolysis data, the results found that Tcf1 deficiency CD8+ TCM cells have reduced glycolysis capacity compared to the wild type.
The research team also verified through technologies such as gene cloning and transgenes that Tcf1-deficient CD8+ TCM secondary response abnormalities can be recovered by tcf1 preprogrammed genes through ectopic expression. Experimental observations have shown that ectopic expression of the Tcf1 preprogrammed gene enhances immune memory in CD8+ TCM cells with Tcf1 sufficiency and Tcf1 deficiency, in part by enhancing the activation of glycolysis.
Tcf1-deficient CD8+ TCM secondary response abnormalities can be recovered by ectopic expression by the Tcf1 preprogramming gene | References[1]
In this study, the researchers discovered an entirely new secondary immune regulatory mechanism: chromatin interactions that rely on Tcf1 can narrow the spatial distance between enhancers and downstream gene promoters, and by "pre-programming," they enter a "ready- state" ahead of schedule. Related studies have also found that Tcf1 is sensitive to histone deacetylase (HDAC) inhibitors, so this result will inevitably have a positive impact on translational medicine and may enable therapeutic interventions to enhance antibacterial and anti-tumor immunity.
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Author: BorderTown
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Typography: Yin Ningliu
Title image source: "Working Cell"
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