THE NUCLEUS AND CENTROSOME REGULATE PERIPHERAL ENDOTHELIAL FUNCTIONS DURING VASCULAR DEVELOPMENT

During angiogenic sprouting and vascular remodeling, endothelial cells, which line the blood vessels, undergo changes in polarity, remodel cell-cell junctions, and respond to mechanical signals to form a fully functional vessel. Recent evidence suggests that two organelles, the centrosome and nucleus, are key players in endothelial cell function, but how the nucleus and centrosome regulate angiogenic processes is largely unknown. The centrosome is a centrally located organelle that acts as a microtubule organizing center. The endothelial cells lining tumor vessels often have supernumerary centrosomes and abnormal vessel properties, including increased permeability and decreased perfusion. However, it is unclear how centrosome number contributes to downstream effects on endothelial function. Near to the centrosome is the nucleus, which is tethered to the rest of the cell by the LINC complex. The LINC complex sits in the nuclear envelope and serves as a bridge that allows for communication between the nucleoplasm and cytoplasm through interactions with the cytoskeleton. Complications in the LINC complex and its associated proteins can lead to vascular disease. For example, Hutchinson-Gilford Progeria Syndrome is a premature aging syndrome characterized by extensive athero- and arteriosclerosis and is associated with changes in the LINC complex. Furthermore, evidence suggests that overactivity of the LINC protein SUN1 may contribute to progeria phenotypes. However, it is unknown how LINC proteins such as SUN1 and the related protein SUN2 function within blood vessels.Here, we sought to understand how the nucleus and centrosome function in endothelial cell function during angiogenesis. In chapter 2, we show that supernumerary centrosomes lead to aberrant apico-basal polarity and disrupted endothelial cell-cell junctions. Altered polarity and cell-cell junctions are upstream of impaired lumenization both in vitro and in vivo, highlighting a role for the centrosome in blood vessel formation. Chapters 3 and 4 explore how LINC complex proteins regulate endothelial function. SUN1 plays a critical role in stabilizing angiogenic sprouts and EC cell-cell junctions. Loss of SUN1 results in impaired angiogenic sprouting both in vitro in vivo, accompanied by destabilization of endothelial adherens junctions and increased permeability. Junction destabilization following SUN1 loss is preceded by dysregulated microtubule dynamics, increased actomyosin contractility, and the release of the microtubule-associated GEF-H1 and is dependent on the LINC complex protein nesprin-1. Meanwhile, SUN2 is important for interactions between endothelial cells and their environment. Loss of SUN2 leads to abnormal responses to blood flow alongside altered focal adhesions, migrations, and lumenization. Overall, our studies reveal important roles for both the centrosome and the nucleus, which both reside in the cell center, in peripheral endothelial functions during angiogenesis.