DECODING THE IMMUNE SYSTEM’S PROTECTIVE ABILITIES: THE T CELL RESPONSE TO PATHOGENS AND CANCER
The CD8⁺ T cell response to intracellular pathogens such as bacteria, viruses and protozoan parasites is an essential component of host resistance. Our lab utilizes experimental infectious disease models of to address questions about the host CD8⁺ T cell immune response against pathogens potential microbial agents of bioterrorism. In response to infection, many factors co-operate to direct CD8⁺ T cells through their dramatic expansion and differentiation to effector and memory cells. Defining the transcriptional regulation of immunological memory formation directly informs efforts to understand anti-pathogen immunity and improve vaccine efficacy. As members of the Immunological Genome Project (ImmGen), we have analyzed the transcriptomes of antigen-specific CD8⁺ T cells over the course of infection to establish dynamic gene-expression signatures, which allow us to predict memory-precursor potential of effector CD8⁺ T cells. Notably, these gene-expression signatures identify putative transcriptional regulators of co-expressed genes. These results provide a framework for us to explore the role of specific gene-expression modules in the function and differentiation of CD8⁺ T cells during immune responses to infection or malignancy and to experimentally define the factors responsible for their regulation.
Dynamic changes in the expression of transcription factors (TFs) can influence the specification of distinct CD8⁺ T cell fates, but the observation of equivalent expression of TFs among differentially fated precursor cells suggests additional underlying mechanisms. We have thus profiled the genome-wide histone modifications, open chromatin and gene expression of naive, terminal-effector, memory-precursor and memory CD8⁺ T cell populations induced during the in vivo response to bacterial infection. Integration of these data suggest that the expression and binding of TFs contributed to the establishment of subset-specific enhancers during differentiation. Working with UCSD collaborators (Kai Zang and Wei Wang) who developed a new bioinformatics method using the PageRank algorithm we were able to reveal key TFs that influence the generation of effector and memory populations. Our combined approach define the epigenetic landscape of differentiation intermediates and facilitate the identification of TFs with previously unappreciated roles in CD8⁺ T cell differentiation.
Through these studies, we have discovered that E protein transcription-factors and their inhibitors (ID proteins), regulate the CD8⁺ T cell response to intracellular pathogens, which is a novel function for these proteins. We are working to understand at a molecular level how this family of transcriptional regulators influences the activation, proliferation, differentiation and survival of CD8⁺ T cells as they transition from naïve to effector to memory cells. While the E proteins are known to regulate many key developmental checkpoints, lineage commitment, proliferation and survival during hematopoiesis and lymphocyte development, the function of these important proteins has been until recently largely unexplored in the mature T cell. The lab is investing in new tools to investigate transcriptional regulation of T cell immunity to build on these observations and leverage these finding to exploit immunity to fight infection and malignancy.
Recognition of pathogen by the immune system initiates a multi-step transcriptional program which directs CD4⁺ T cell differentiation into distinct T helper populations that coordinate eradication of infection. Effector CD4⁺ T cells secrete inflammatory cytokines and orchestrate the activation of immune cells, while T follicular helper cells cells secrete cytokines and upregulate ligands that induce B cells to form germinal centers, class switch, and generate high-affinity antibodies. Differentiation of CD4⁺ T cells is directed by cytokine-induced activation of STAT proteins and lineage determining transcription factors. We have found that many of the transcription factors that regulate CD8⁺ T cell activation and differentiation provide key functions in CD4⁺ T helper differentiation and memory formation as well, providing a new direction for our studies.
CD8⁺ T lymphocytes are a key element of endogenous cancer immunity and are frequently utilized in immunotherapies for treatments ranging from solid tumors to blood cancers. CD8⁺ T cells remain an attractive option for anti-tumor therapy approaches due to the ability of tumor-specific T cells to directly target and kill malignant cells. T cells exhibit extensive heterogeneity in functional parameters ranging from localization, metabolic profile, proliferative potential and capacity for self-renewal. Currently, the optimal cellular attributes required for maximizing the efficacy of CD8⁺ T cell immunotherapies remain unclear. By decoding the genetic instructions that direct the immune system to remember past encounters with pathogens and provide robust protection from reinfection, we aim to rev up the immune system to eradicate infected and malignant cells by exploiting their knowledge of programming robust T cell responses.
TISSUE-RESIDENT MEMORY T CELLS
During an immune response to microbial infection, a naive CD8⁺ T lymphocyte can give rise to terminal effector cells that provide acute host defense and functionally distinct subsets of memory cells that provide durable immunity. Generation of one such subset, tissue-resident memory T cells, is now recognized to be essential for protection at mucosal and body surfaces. Long-lived memory cells provide protection from reinfection and can serve as endogenous defenders against tumor growth and metastases. Tissue-resident memory T cells provide essential sentinel protection at body surfaces such as the intestinal epithelium, and are now clearly understood to be among the key ‘ first responders’ in many infection settings. Although we know that resident-memory cells are an essential component of immune memory, little is known about the transcriptional pathways regulating their formation, survival and function. By identifying the unique transcriptional pathways and regulators of tissue-resident-memory T cells, we will discover novel targets informing strategic design of therapeutic and protective vaccines, the development of which are of critical importance to human health.
MICROENVIRONMENTAL INFLUENCES ON T CELL RESPONSES AND DIFFERENTIATION
CD8⁺ T cells are required to control many intracellular infections and tumors, accessing tissues to induce the death of infected/malignant cells and limit dissemination of pathogen. The CD8⁺ T cell can encounter marked changes in microenvironment during the course of infection as it moves within and between damaged and infected tissues, yet how alterations in oxygen and nutrient availability impact cytotoxic T cell differentiation and function is poorly understood. Effector CD8⁺ T cells must balance the metabolic requirements of expansion, survival, and cytotoxic function with the limiting nutrient and oxygen availability of infected tissues. While T cell activation induces profound changes in gene expression, relatively little is known about how the microenvironmental milieu impacts the transcriptional network controlling T cell effector function and memory differentiation. We are examining the impact of microenvironment on gene expression in the context of infection.
Reports utilizing the immunosuppressant rapamycin and the diabetes drug metformin have uncovered an mTOR/AMPK-dependent metabolic switch from glycolytic to fatty acid metabolism. These results suggest metabolism may reinforce cell-fate decisions leading to the generation of CD8⁺ T cell memory, and that metabolic activity may be linked to the function and survival of effector and memory cells, yet how microenvironmental cues modulate the metabolism of these cells is unknown. We are exploring possible links between hypoxia or limiting oxygen and transcriptional integration of tissue specific extracellular signals and CD8⁺ T cell differentiation.
In this context, we find that CD8⁺ T cell immunity is controlled by a central transcriptional regulator of hypoxic responses, the von Hippel-Lindau/Hypoxia-Inducible Factor (VHL/HIF) pathway. Elevated HIF activity dramatically enhances CD8⁺ T cell effector function and delays differentiation to both short-lived effector/memory and long-lived memory cell populations. Further, hypoxia modulates CD8⁺ T cell expression of pivotal transcription factors, effector molecules, and inhibitory receptors in a HIF1/2a (HIFa)-dependent fashion. Thus, we have uncovered a key role for HIF-mediated transcription in the adaptive CD8⁺ T cell immune responses. In vivo, CD8⁺ effector cells traffic to sites of infection, where hypoxia from tissue damage coincides with recognition of pathogen-infected cells and, ultimately, cytolytic activity. Thus, the capacity of HIF signaling to locally limit terminal differentiation in these zones, may serve to tie pathogen load and tissue damage with effector function. We hypothesize that this transcriptional response will be particularly important in the immune response to tissue infections where hypoxic zones coincide with infection and studies to explore this possibility are underway in the lab.
REGULATION OF NAIVE AND MEMORY T CELL POPULATIONS
Mature T cells persist within the body for indefinite periods of time, providing protection from a vast array of pathogens. A fraction of naïve T cells survive for decades and some memory T cells survive for a lifetime without re-exposure to antigen. It is not known how the lifespan of T cells is regulated at the individual, cellular level or at the population level. It is the objective of our research to identify the cellular and molecular interactions regulating the survival of both naive and memory T cells.
This information is crucial to understanding T cell homeostasis and immunological memory (the basis of vaccination against both tumors and pathogens). Several lines of study in our lab aim to identify the molecular signals required to maintain T cells, generate a productive immune response and form long-lived protective immunity. It is our hope that our experiments will improve the understanding of how the immune system recovers following treatment- or illness-induced lymphopenia (such as chemotherapy or HIV infection) and aid in the design of vaccines that provide long-lasting protection from infection. Moreover, by expanding our understanding of how lymphocyte populations are regulated, it will be possible to gain insight into how normal survival signals are co-opted and how homeostatic set points are overcome by cancer cells providing possible targets for therapeutic intervention.
Funding for our research comes from
National Institutes of Health • Leukemia and Lymphoma Foundation • Julia Brown Faculty Fellowship
Pew Foundation • California Institute for Regenerative Medicine • Cancer Research Institute
California Breast Cancer Research Program • V Foundation