Lymphatic endothelial cells (LECs) line lymphatic vessels, which are critical in health through the control of fluid homeostasis, lipid transport and inflammation. In contrast to angiogenesis, it is not known whether metabolic pathways may regulate lymphangiogenesis. The prospero homeobox transcription factor 1 (PROX1) is critical for the differentiation of LECs from venous endothelial cells (VECs) in vivo, and overexpression of PROX1 in VECs induces differentiation to LECs in vitro. To date, only genetic signals have been shown to determine the VEC-to-LEC switch, and a role for metabolism in LEC differentiation has not been considered. We demonstrate that LEC-specific loss of carnitine palmitoyltransferase 1A (CPT1A), a rate-controlling step of fatty acid ß-oxidation (FAO), impairs lymphatic development. LECs require FAO for lymphatic sprouting and lymphangiogenesis through the epigenetic regulation of lymphatic differentiation marker expression. Mechanistically, PROX1 directly upregulates CPT1A expression, thereby increasing FAO-dependent acetyl-CoA production, which is used by the histone acetyltransferase p300 to specifically acetylate histones at lymphangiogenic genes. Unbiased, whole-genome ChIP-seq experiments demonstrate that interactions between PROX1 and the histone acetyltransferase p300 facilitates preferential histone acetylation at PROX1-target genes. Thus, the increased FAO flux during LEC differentiation mediates epigenetic changes that promote lymphatic development and lymphangiogenesis. Notably, pharmacological CPT1 blockade inhibits pathological lymphangiogenesis, while replenishing acetyl-CoA by supplementing acetate rescues histone acetylation and lymphangiogenesis in vitro and in vivo.