PKN2 signalling induces stromal cell protrusions to preserve lymph node structural integrity

Secondary lymphoid tissues develop specialized reticular networks to facilitate immune cell communication and efficient activation of adaptive immunity. This stromal network architecture is robust, maintaining topology throughout extensive remodelling and tissue expansion in response to immune challenge. We have previously reported that cytoskeletal mechanics of the fibroblastic reticular cell (FRC) networks determine tissue tension, and that increased tension initiates stromal proliferation for lymph node growth. However, it is not known how FRCs maintain stromal network connectivity and what cellular mechanisms reinforce stromal cell-cell and cell-matrix interactions. Here, we present a signalling mechanism which coordinates reduced FRC contractility and induction of stromal cell protrusions. RhoA/C GTPase activity is blocked in FRCs to inhibit actomyosin contractility through contact with dendritic cells (DCs) and binding between podoplanin and the C-type lectin CLEC-2. We now find that an additional Rho GTPase target, the PKC family kinase PKN2, regulates the function of myristoylated alanine-rich protein kinase C-substrate (MARCKS). FRCs generate cell protrusions via MARCKS in response to DC contact, which reinforces stromal cell connectivity. In vivo, we found that PKN2 knock-out lymph nodes are unable to regulate MARCKS and show severely disrupted stromal architecture. These results reveal a mechanism of stromal/immune cell crosstalk which actively induces stromal protrusions – an essential component of lymph node remodelling to maintain tissue integrity during an adaptive immune response.