Suling Li

Although T cells can develop into an exhausted state in the tumour microenvironment, tumour infiltrating T cells (TILs) are important to control tumour growth. By analysing single cell RNA-sequencing data from human tumours, we found that the transcription factors Early Growth Response 2 (EGR2) and 3 were highly induced in TILs, but not peripheral CD8 + T cells, in multiple patient cohorts. We found that deficiency of Egr2 and 3 in T cells resulted in enhanced tumour growth and fewer TILs in mouse models. Egr2 is highly expressed together with checkpoint molecules in a proportion of CD8 + TILs and Egr2high cells exhibit better survival and proliferation than Egr2^-/-Egr3^-/- and Egr2low TILs. Anti-PD-1 treatment increases Egr2 expression in CD8 + TILs and reduces tumour growth, while anti-PD-1 efficacy is abrogated in the absence of Egr2 and 3. Thus, Egr2 and 3 are important for maintaining anti-tumour responses of exhausted CD8 + TILs.

The transcription factors Egr2 and 3 are essential for controlling inflammatory autoimmune responses of memory phenotype (MP) CD4 T cells. However, the mechanism is still unclear. We have now found that the Egr2⁺subset (PD-1^highMP) of MP CD4 T cells expresses high levels of checkpoint molecules (PD-1 and Lag3) and also markers of effector T cells (CXCR3 and ICAM-1). Egr2/3 are not required for PD-1^highMP CD4 cell development but mediate a unique transcriptional programme that effectively controls their inflammatory responses, while promoting homeostatic proliferation and adaptive responses. Egr2 negative PD-1^highMP CD4 T cells are impaired in homeostatic proliferation and adaptive responses against viral infection but display inflammatory responses to innate stimulation such as IL-12. PD-1^highMP CD4 T cells have recently been implicated in rheumatoid arthritis pathogenesis, and we have now found that Egr2 expression is reduced in PD-1^highMP CD4 T cells from patients with active rheumatoid arthritis compared with healthy controls. These findings demonstrate that Egr2/3 control the inflammatory responses of PD-1^highMP CD4 T cells and maintain their adaptive immune fitness.

Introduction: Impaired proliferation and production of IL2 are the hallmarks of experimental T cell tolerance. However, in most autoimmune diseases, auto‐reactive T cells do not display hyper proliferation, but inflammatory phenotypes.Methods: We have now demonstrated that the transcription factors Egr2 and 3 are important for the control of inflammatory cytokine production by tolerant T cells, but not for tolerance induction.Results: In the absence of Egr2 and 3, T cell tolerance, as measured by impaired proliferation and production of IL2, can still be induced, but tolerant T cells produced high levels of inflammatory cytokines. Egr2 and 3 regulate expression of differentiation repressors and directly inhibit T‐bet function in T cells. Indeed, decreased expression of differentiation repressors, such as Id3 and Tcf1, and increased expression of inflammatory transcription factors, such as RORγt and Bhlhe40 were found in Egr2/3 deficient T cells under tolerogenic conditions. In addition, T‐bet was co‐expressed with Egr2 in tolerant T cells and Egr2/3 defects leads to production of high levels of IFNγ in tolerant T cells.Conclusions: Our findings demonstrated that despite impaired proliferation and IL2 production, tolerant T cells can display inflammatory responses in response to antigen stimulation and this is controlled at least partly by Egr2 and 3.

T-bet is important for differentiation of cytotoxic CD8 and Th1 CD4 T cells. We have discovered that Egr2 and 3 are potent inhibitors of T-bet function in CD4 and CD8 effector T cells. Egr2 and 3 were essential to suppress Th1 differentiation in Th2 and Th17 conditions in vitro and also to control IFN-γ–producing CD4 and CD8 T cells in response to virus infection. Together with Egr2 and 3, T-bet is induced in naive T cells by Ag stimulation, but Egr2 and 3 expression was inhibited by Th1–inducing cytokines. We found that Egr2 and 3 physically interact with the T-box domain of T-bet, blocking T-bet DNA binding and inhibiting T-bet–mediated production of IFN-γ. Thus, Egr2 and 3 are antagonists of T-bet function in effector T cells and are important for the control of inflammatory responses of T cells.

Egr2 and 3 are important for maintaining immune homeostasis. Here we define a fundamental function of Egr2 and 3 operating as a checkpoint that controls the transition between clonal expansion and differentiation of effector T cells. Egr2 and 3 deficiency resulted in defective clonal expansion but hyperactivation and excessive differentiation of T cells in response to viral infection. Conversely, sustained Egr2 expression enhanced expansion but severely impaired effector differentiation. Egr2 bound to and controlled the expression of genes regulating proliferation (Myc and Myb) and differentiation repressors (Bcl6, Id3), while repressing transcription factors required for effector function (Zeb2, RORa, RORc, and Bhlhe40). Egr2 and 3 expression in T cells was regulated reciprocally by antigen and IFNγ, providing a mechanism for adjusting proliferation and differentiation of individual T cells. Thus, Egr2 and 3 are upstream regulators of effector CD4 and CD8 T cells that are essential for optimal responses with limited immunopathology.

Paper-based generators are essential elements for building all paper-based systems. To obtain robust paper-based generators with outstanding high power outputs, this paper introduced a new type of double-folding paper-based generator by folding two paper components together. The output performance levels of the double-folding generator were twice higher than that of the single-folding and parallel-plate generators. A peak power of ~3.24 mW was achieved under a stimulating frequency of 3 Hz. Furthermore, 47 light-emitting diodes (LEDs) were lit directly by a double-folding paper-based generator assembled to the crack of a door that opens and closes. This finding indicated the potential applications of the double-folding generator in the production of door ornaments or for security in places where doors frequently open and close.

Understanding of how transcription factors are involved in lymphocyte development still remains a challenge. It has been shown that Egr-2 deficiency results in impaired NKT cell development and defective positive selection of T cells. Here we investigated the development of T, B and NKT cells in Egr-2 transgenic mice and the roles in the regulation of distinct stages of B and T cell development. The expression of Egr1, 2 and 3 were analysed at different stages of T and B cell development by RT-PCT and results showed that the expression was strictly regulated at different stages. Forced expression of Egr-2 in CD2⁺ lymphocytes resulted in a severe reduction of CD4⁺CD8⁺ (DP) cells in thymus and pro-B cells in bone marrow, which was associated with reduced expression of Notch1 in ISP thymocytes and Pax5 in pro-B cells, suggesting that retraction of Egr-2 at the ISP and pro-B cell stages is important for the activation of lineage differentiation programs. In contrast to reduction of DP and pro-B cells, Egr-2 enhanced the maturation of DP cells into single positive (SP) T and NKT cells in thymus, and immature B cells into mature B cells in bone marrow. Our results demonstrate that Egr-2 expressed in restricted stages of lymphocyte development plays a dynamic, but similar role for the development of T, NKT and B cells.

Although matured DC are capable of inducing effective primary and secondary immune responses in vivo, it is difficult to control the maturation and antigen loading in vitro. In this study, we show that ER‐enriched microsomal membranes (microsomes) isolated from DC contain more peptide‐receptive MHC I and II molecules than, and a similar level of costimulatory molecules to, their parental DC. After loading with defined antigenic peptides, the microsomes deliver antigenic peptide–MHC complexes (pMHC) to both CD4 and CD8 T cells effectively in vivo. The peptide‐loaded microsomes accumulate in peripheral lymphoid organs and induce stronger immune responses than peptide‐pulsed DC. The microsomal vaccines protect against acute viral infection. Our data demonstrate that peptide–MHC complexes armed microsomes from DC can be an important alternative to DC‐based vaccines for protection from viral infection.

Microarray technology has had a significant impact in the field of systems biology involving the investigation into the biological systems that regulate human life. Identifying genes of significant interest within any given disease on an individual basis is no doubt time consuming and inefficient when considering the complexity of the human genome. Thus, the genetic profiling of the entire human genome in a single experiment has resulted in microarray technology becoming a widely used experimental tool. However, without the use of tools for several aspects of microarray data analysis the technology is limited. To date, no such tool has been developed that allows the integration of numerous microarray results from different research laboratories as well as the design of customised gene chips in a cost-effective manner. In light of this, we have designed the first integrated and automated software called Chip Integration, Design and Annotation (CIDA) for the cross comparison, design and functional annotation of microarray gene chips. The software provides molecular biologists with the control to cross compare the biological signatures generated from multiple microarray studies, design custom microarray gene chips based on their research requirements and lastly characterise microarray data in the context of immunogenomics. Through the relative comparison of related microarray experiments we have identified 258 genes with common gene expression profiles that are not only upregulated in anergic T cells, but also in cells over-expressing the transcription factor Egr2, that has been identified to play a role in T cell anergy. Using the gene chip design aspect of CIDA we have designed and subsequently fabricate immuno-tolerance gene chips consisting of 1758 genes for further research. The software and database schema is freely available at ftp://ftp.brunel.ac.uk/cspgssk/CIDA/. Additional material is available online at http://www.brunel.ac.uk/about/acad/health/healthres/researchgroups/mi/publications/supplementary/cida