Role of Rasa1 in embryonic development

PROJECT SUMMARY Vascular and lymphatic anomalies comprise a spectrum of defects that originate in embryonic blood or lymphatic vessels and are present at birth. These defects can be congenital or sporadic and nearly 50 genes bearing causative mutations have been identified for both cases. Among these is RASA1,

which encodes a negative regulator of the RAS GTPase that is normally activated by growth factor signaling. Germline autosomal dominant mutations in RASA1 lead to capillary malformation- arteriovenous malformation (CM-AVM) syndrome, in which patients exhibit port wine stains on their skin at birth. CM-AVM patients also harbor AVMs, which are direct artery/vein connections without an

intervening capillary bed, or lymphatic anomalies. AVMs usually occur in the brain where they have a significant negative health impact, while the fast-flow nature of AVMs can lead to cardiac overload. Lymphatic anomalies can perturb fluid clearance, leading to cardiac or pleural effusions, and hydrops

fetalis. Accordingly, AVMs and lymphatic anomalies can contribute to fetal demise or early neonatal lethality and therapeutic options are limited. Mouse models have provided important insights into the role of Rasa1 in embryogenesis, including its requirement for vascular and lymphatic development. However, many defects in Rasa1-deficient mouse embryos are not seen in human CM-AVM lesions

and this model fails to mimic postnatal features of CM-AVM, underscoring the need for additional animal models. We have developed rasa1-deficient zebrafish embryos and show that they exhibit defects in both vascular and lymphatic development, including vascular regression defects not previously noted in other models. Adult zebrafish carriers of mutant rasa1 alleles also show evidence

of presumptive capillary malformations, suggesting that they may exhibit hallmarks of CM-AVM. To study the role of Rasa1 in zebrafish, we have developed numerous genetic tools, including zebrafish bearing a floxed rasa1a allele and endothelial Cre driver lines. Using these tools, we will define the cellular and molecular effects of rasa1 deficiency that give rise to lymphatic defects. In addition, we

will investigate how rasa1 acts to mediate vascular regression and whether defects in this process contribute to AVM formation. Finally, we will characterize presumptive capillary malformations in rasa1 carriers, while assessing this model as a platform for preclinical testing of therapeutic small molecule candidates to alleviate CM-AVM defects. Together, our studies will provide valuable insight

on the embryonic role of Rasa1 in development and how its deficiency contributes to formation of vascular and lymphatic anomalies.