Highlights d A single-cell EC atlas of healthy murine tissues d This study provides an interactive webtool for comparative analyses and data exploration d Characterization of inter-and intra-tissue EC heterogeneity d Discovery tool for characterization of ECs in other datasets

Mutations in the cytosine-5 RNA methyltransferase NSun2 cause microcephaly and other neurological abnormalities in mice and human. How post-transcriptional methylation contributes to the human disease is currently unknown. By comparing gene expression data with global cytosine-5 RNA methylomes in patient fibroblasts and NSun2-deficient mice, we find that loss of cytosine-5 RNA methylation increases the angiogenin-mediated endonucleolytic cleavage of transfer RNAs (tRNA) leading to an accumulation of 5′ tRNA-derived small RNA fragments. Accumulation of 5′ tRNA fragments in the absence of NSun2 reduces protein translation rates and activates stress pathways leading to reduced cell size and increased apoptosis of cortical, hippocampal and striatal neurons. Mechanistically, we demonstrate that angiogenin binds with higher affinity to tRNAs lacking site-specific NSun2-mediated methylation and that the presence of 5′ tRNA fragments is sufficient and required to trigger cellular stress responses. Furthermore, the enhanced sensitivity of NSun2-deficient brains to oxidative stress can be rescued through inhibition of angiogenin during embryogenesis. In conclusion, failure in NSun2-mediated tRNA methylation contributes to human diseases via stress-induced RNA cleavage.

Summary Abnormal tumor vessels promote metastasis and impair chemotherapy. Hence, tumor vessel normalization (TVN) is emerging as anti-cancer treatment. Here, we show that tumor endothelial cells (ECs) have a hyper-glycolytic metabolism, shunting intermediates to nucleotide synthesis. EC haplo-deficiency or blockade of the glycolytic activator PFKFB3 did not affect tumor growth, but reduced cancer cell invasion, intravasation and metastasis by normalizing tumor vessels, which improved vessel maturation and perfusion. Mechanistically, PFKFB3 inhibition tightened the vascular barrier by reducing VE-cadherin endocytosis in ECs, and rendering pericytes more quiescent and adhesive (via upregulation of N-cadherin) through glycolysis reduction; it also lowered the expression of cancer cell adhesion molecules in ECs by decreasing NF-κB signaling. PFKFB3-blockade treatment also improved chemotherapy of primary and metastatic tumors.

Endothelial cells (ECs) are more than inert blood vessel lining material. Instead, they are active players in the formation of new blood vessels (angiogenesis) both in health and (life-threatening) diseases. Recently, a new concept arose by which EC metabolism drives angiogenesis in parallel to well-established angiogenic growth factors (e.g., vascular endothelial growth factor). 6-Phosphofructo-2-kinase/fructose-2,6-bisphosphatase-3-driven glycolysis generates energy to sustain competitive behavior of the ECs at the tip of a growing vessel sprout, whereas carnitine palmitoyltransferase 1a-controlled fatty acid oxidation regulates nucleotide synthesis and proliferation of ECs in the stalk of the sprout. To maintain vascular homeostasis, ECs rely on an intricate metabolic wiring characterized by intracellular compartmentalization, use metabolites for epigenetic regulation of EC subtype differentiation, crosstalk through metabolite release with other cell types, and exhibit EC subtype-specific metabolic traits. Importantly, maladaptation of EC metabolism contributes to vascular disorders, through EC dysfunction or excess angiogenesis, and presents new opportunities for anti-angiogenic strategies. Here we provide a comprehensive overview of established as well as newly uncovered aspects of EC metabolism.