CD4 T cells orchestrate antiviral adaptive immunity by providing help to CD8 T cells through Th1 cells and by facilitating B cell maturation and generation of antibody-producing plasma cells through follicular T helper (Tfh) cells. In the second funding period, we have longitudinally analysed HEV-specific CD4 T cells together with Neumann-Haefelin, Bengsch, Hofmann, Thimme and Dao Thi and demonstrated that they form stable immune memory that persists antigen-independently over years after viral clearance. In contrast, during chronic infection, their frequencies are low, yet they are maintained antigen-dependently and retain functional capabilities (with Hofmann and Thimme). In the third funding period, we will continue our studies of antiviral CD4 T cell responses in different settings of viral hepatitis with a special focus on the identification of characteristics and trajectories of virus-specific CD4 T cell memory and self-renewal in different phases of acute and chronic viral hepatitis. The overarching theme is the question which virus-specific CD4 T cell populations facilitate maintenance and longevity of CD4 T cell responses, how they change over time and how they differ from resolved infection with memory development and chronic infection.
Specific aim 1 will tackle the question how virus-specific CD4 T cell responses are maintained in the context of chronic HBV and HCV infection (with Thimme and Hofmann). Using state of the art single-cell technology, we aim to identify which precursor populations replenish the pool of circulating virus-specific CD4 T cells (with Z03). Sequencing of the patient specific viral isolates will allow detailed analyses of CD4 T cells targeting conserved or mutated/escaped MHC class II epitopes (with Neumann-Haefelin/Luxenburger). As was observed for CD8 T cells, we anticipate that viral clearance (i.e. through DAA therapy in patients with chronic HCV-infection) will affect the composition of the CD4 T cell compartment and specifically the pool of precursors.
Similarly, as aim 2 and following our work in the second funding period, we aim to identify characteristics of early memory formation in patients with acute-resolving viral hepatitis. We will longitudinally analyse virus-specific CD4 T cells in patients with acute HEV and HCV infection and identify TCR clones within the virus-specific CD4 T cell pool that are maintained from acute infection to years after viral clearance (with Bengsch, Hofmann, Dao Thi and Z03). Through T cell receptor analyses of virus-specific CD4 T cells during acute infection and several years after infection, we aim to identify phenotypic and transcriptional characteristics of memory precursors during acute infection (reverse phenotyping). This will help us to clearly decipher the phenotype of memory precursors, which is of great importance as there is a strong overlap of markers identifying Tfh cells and memory subsets while it remains unclear how far Tfh functionality is preserved in memory CD4 T cells.
In specific aim 3, we want to functionally validate the relevance of the identified memory signa-tures using an in vitro system for T cell-B cell interactions and for secondary expansion after antigen re-exposure in vivo. For subaim 1, we propose to generate CD4 T cell clones from transcriptionally different memory subsets and evaluate B cell helper capacities by co-cultures with CD4 T cell clones and na-ïve/memory B cells with subsequent IgM and IgG measurements (method established together with Bengsch). For part 2, we will use different vaccinations (seasonal influenza, SARS-CoV-2 and hepa-titis B) as model systems. By analysing vaccine-specific CD4 T cells at baseline and after vaccination, we will be able to identify transcriptional profiles and phenotypic characteristics of specific memory TCR clones that have expanded in response to a vaccine (together with Hofmann and Z03).
Collectively, the experiments will provide important insights into the regulation and dynamics of virus-specific CD4 T cell responses.