Mechanisms of actin polymerization in T lymphocyte nuclei

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While the roles of the actin cytoskeleton in the cytoplasm of mammalian cells are well studied, the functional relevance of nuclear actin dynamics has only recently begun to emerge. An increasing number of studies report that nuclear actin filaments play important roles in regulating interactions of mammalian cells with their environment and that distinct actin nucleators govern the formation of nuclear actin filaments in individual pathways. At the beginning of the first funding period, information was lacking on the role of nuclear actin filaments in immune cells in general and in T Cell Receptor (TCR) signaling specifically. During the first funding period we established a transient burst of nuclear actin polymerization as essential effector function of CD4 T cell activation that is required for expression of a select set of target genes, ample cytokine expression, and thus efficient helper function. We also defined Arp2/3 complex as an essential actin nucleator for this process, whose activity is regulated by nuclear Ca2+ transients. While nuclear Ca2+ may act as general regulator of nuclear actin dynamics, different actin nucleators are involved in the formation of nuclear actin networks in a cell type and stimulus dependent manner. Our preliminary results suggest that in CD4 T cells, the specific involvement of the Arp2/3 isoforms C5L and C5 govern the compartmentalization of actin polymerization following TCR engagement and selectively drive nuclear and cytoplasmic actin dynamics, respectively. In the second funding period we will now aim at defining in more detail the compartment specific regulation of nuclear actin polymerization and dissect how nuclear actin dynamics is coupled to selective target gene expression (specific aim 1). In a second work package we will make use of a new mouse model we generated to express a fluorescent reporter for nuclear F-actin to visualize nuclear actin dynamics by intravital microscopy in vivo, dissect how nuclear actin dynamics drives mounting of humoral immune responses, and assess how conserved triggering of nuclear dynamics is in various immune cells. Together, these analyses will provide important insight in mechanism and relevance of nuclear actin dynamics in CD4 T cells and other immune cells and establish principles of the functional impact of nuclear F-actin in general.