The vertebrate vasculature is formed by populations of endothelial and mural cells that arrange into functionally and molecularly distinct arterial, venous and capillary beds. Although a number of signalling pathways and transcriptional regulators have been implicated in these processes of vascular differentiation, a clear picture of how arterial-specific gene regulation is achieved is yet to emerge. In this study I have investigated the transcriptional regulation of arterial identity from two different directions: characterisation of enhancers to identify the transcription factors that bind and direct arterial specification; and direct study of the function of one particular transcription factor expressed specifically in the arterial vasculature. I have identified a novel gene enhancer that directs arterial-specific expression of Flk1 (Vegfr2) in transgenic mouse and zebrafish models. Dissection of inputs from individual transcription factor binding sites within this enhancer shows a requirement for Gata factors for enhancer function in endothelial cells, whereas arterial-specification is directed by Rbpj-mediated repression of enhancer activity in veins. This work demonstrates that Flk1 expression in arterial endothelial cells is downstream of the Notch/Rbpj pathway, and also describes a novel transcriptional mechanism of arterial differentiation. In parallel, I have uncovered a novel role for the transcription factor Tbx2 in the regulation of arterial mural cell identity. Histological analysis demonstrates the previously unreported expression of Tbx2 exclusively in mural cells of peripheral arteries and microvessels, and genetic deletion experiments in mice suggest a role for Tbx2 in mural cell recruitment, survival, proliferation, and differentiation upstream of Notch3 and Pdgfrβ. Together, these results contribute valuable insights into our understanding of the establishment of vascular identity by identifying novel transcriptional regulators of arterial fate in both endothelial and mural cells.