Rasip1 controls lymphatic vessel lumen maintenance by regulating endothelial cell junctions

Although major progress in our understanding of the genes and mechanisms regulating lymphatic vasculature development has been made, we still do not know how lumen formation and maintenance occurs. Here we identify the Ras interacting protein Rasip1 as a key player in this process. We show that lymphatic endothelial cells-specific Rasip1 deficient embryos exhibit enlarged and blood-filled lymphatics at E14.5. These vessels have patent lumens with disorganized junctions. Later on, as those vessels become fragmented and lumens collapse, cell junctions become irregular. In addition, Rasip1 deletion at later stages impairs lymphatic valve formation. We determined that Rasip1 is essential for lymphatic lumen maintenance during embryonic development by regulating junction integrity, as Rasip1 loss results in reduced levels of junction molecules and defective cytoskeleton organization in vitro and in vivo. We determined that Rasip1 regulates Cdc42 activity as deletion of Cdc42 shows similar phenotypes to that of Rasip1 loss. Furthermore, ectopic Cdc42 expression rescues the phenotypes in Rasip1 deficient LECs, further supporting that Rasip1 regulates Cdc42 activity to regulate cell junctions and cytoskeleton organization, activities required for lymphatic lumen maintenance.