UCSF

Clathrin is a coat protein that mediates membrane traffic by capturing and transporting cargo between different cellular compartments. It is composed of heavy and light chain subunits, which coat membranes of budding vesicles involved in receptor-mediated endocytosis and organelle biogenesis. Clathrin-mediated endocytosis (CME) of many cargoes can occur in the absence of CLCs, however recent studies have demonstrated that they are required for specialized clathrin functions that require actin assembly through binding the actin-organizing huntingtin-interacting proteins (Hip). Since cell migration requires both clathrin and actin, we investigated the role of the CLCs in this pathway. Inhibition of CLC function by siRNA-mediated depletion and by expression of a dominant negative CLC decrease cell migration and recycling of inactive β1 integrin. These CLC functions occur through Hip binding. CLC depletion and expression of a dominant negative CLC also reduce the appearance of gyrating (G)-clathrin structures, which mediate rapid recycling of transferrin receptor, implicating G-clathrin in these pathways. These results demonstrate that CLCs’ ability to link clathrin to actin is necessary for important cellular functions like cell migration and led us to ask which CLC functions are physiologically relevant in vivo. We addressed this question by genetically engineering mice lacking the CLCa isoform. Murine tissue analyses indicated that lymphoid tissues express predominantly CLCa, which prompted a detailed investigation of these cell types in the CLCa mutant mice. Immunoglobulin class switch recombination between IgA and IgG is impaired in the germinal center B cells of the CLCa-deficient mice, which stems from CLCs’ role in mediating TGFβR2 endocytosis. CLCs are also necessary for CME of CXCR4 and the delta opioid receptor but not the β2 agonist receptor or CXCR5, indicating that CLCs exhibit differential cargo selectivity. Thus, our results reveal that CLCs play an important role in CME for a subset of signaling receptors in vivo. Moreover, these studies demonstrate that CLCs influence a broad range of physiological functions including cell migration and adaptive immune responses through their regulation of clathrin coat dynamics.