Endothelial cells (ECs) are plastic cells that can switch between growth states with different bioenergetic and biosynthetic requirements1. Although quiescent in most healthy tissues, ECs divide and migrate rapidly upon proangiogenic stimulation2,3. Adjusting endothelial metabolism to growth state is central to normal vessel growth and function1,4, yet poorly understood at the molecular level. Here we report that the forkhead box O (FOXO) transcription factor FOXO1 is an essential regulator of vascular growth that couples metabolic and proliferative activities in ECs. Endothelial-restricted deletion of FOXO1 in mice induces a profound increase in EC proliferation that interferes with coordinated sprouting, thereby causing hyperplasia and vessel enlargement. Conversely, forced expression of FOXO1 restricts vascular expansion and leads to vessel thinning and hypobranching. We find that FOXO1 acts as a gatekeeper of endothelial quiescence, which decelerates metabolic activity by reducing glycolysis and mitochondrial respiration. Mechanistically, FOXO1 suppresses signalling by c-MYC (termed MYC hereafter), a powerful driver of anabolic metabolism and growth5,6. MYC ablation impairs glycolysis, mitochondrial function and proliferation of ECs while its EC-specific overexpression fuels these processes. Moreover, restoration of MYC signalling in FOXO1-overexpressing endothelium normalises metabolic activity and branching behaviour. Our findings identify FOXO1 as a critical rheostat of vascular expansion and define the FOXO1 – MYC transcriptional network as a novel metabolic checkpoint during endothelial growth and proliferation.

Blood vessels form the first organ in the developing embryo and build extensive networks that supply all cells with nutrients and oxygen throughout life. As blood vessels get older, they often become abnormal in structure and function, thereby contributing to numerous age-associated diseases including ischemic heart and brain disease, neurodegeneration, or cancer.

Yersinia enterocolitica is a common cause of food-borne gastrointestinal disease leading to self-limiting diarrhea and mesenteric lymphadenitis. Occasionally, focal abscess formation in the livers and spleens of certain predisposed patients (those with iron overload states such as hemochromatosis) is observed. In the mouse oral infection model, yersiniae produce a similar disease involving the replication of yersiniae in the small intestine, the invasion of Peyer's patches, and dissemination to the liver and spleen. In these tissues and organs, yersiniae are known to replicate predominately extracellularly and to form microcolonies. By infecting mice orally with a mixture of equal amounts of green-and red-fluorescing yersiniae (yersiniae expressing green or red fluorescent protein), we were able to show for the first time that yersiniae produce exclusively monoclonal microcolonies in Peyer's patches, the liver, and the spleen, indicating that a single bacterium is sufficient to induce microcolony and microabscess formation in vivo. Furthermore, we present evidence for the clonal invasion of Peyer's patches from the small intestine. The finding that only very few yersiniae are required to establish microcolonies in Peyer's patches is due to both Yersinia-specific and host-specific factors. We demonstrate that yersiniae growing in the small intestinal lumen show strongly reduced levels of invasin, the most important factor for the early invasion of Peyer's patches. Furthermore, we show that the host severely restricts sequential microcolony formation in previously infected Peyer's patches.

Key Points• Integrin signaling promotes proliferative signals in AML cells that are mediated by the kinase Syk and the transcription factors STAT3 and STAT5.Spleen tyrosine kinase (Syk) induces cell survival and proliferation in a high proportion of acute myeloid leukemia (AML) blasts, but the underlying molecular events of Syk signaling have not been investigated. Proteomic techniques have allowed us to identify the multiprotein complex that is nucleated by constitutively active Syk in AML cells. This complex differs from the B-lymphoid Syk interactome with respect to several proteins, especially the integrin receptor Mac-1, the Fc-g receptor I (FcgRI), and the transcription factors STAT3 and STAT5. We show in several AML cell line models that tonic signals derived from the Fc-g chain lead to Syk-dependent activation of STAT3 and STAT5, which in turn induces cell survival and proliferation. Moreover, stimulation of Mac-1 or FcgRI intensifies the constitutive Syk-mediated STAT3/5 activation in AML cells, a scenario likely to take place in the bone marrow niche. In accordance with these findings, we observed that b 2 integrins, including Mac-1, trigger proliferation of AML cells in an AML cell/stroma coculture model. Taken together, we identified an oncogenic integrin/Syk/STAT3/5 signaling axis that might serve as a therapeutic target of AML in the future. (Blood. 2013;121(19):3889-3899)

Key Points• Two novel transducer modules consisting of BTK in combination with either FLT3-ITD or TLR9 induce distinct oncogenic signaling programs.• This study suggests subtypespecific treatment strategies, including BTK/FLT3 inhibitor combinations, and shows how TLR9 affects AML biology.Acute myeloid leukemia (AML) is driven by niche-derived and cell-autonomous stimuli. Although many cell-autonomous disease drivers are known, niche-dependent signaling in the context of the genetic disease heterogeneity has been difficult to investigate. Here, we analyzed the role of Bruton tyrosine kinase (BTK) in AML. BTK was frequently expressed, and its inhibition strongly impaired the proliferation and survival of AML cells also in the presence of bone marrow stroma. By interactome analysis, (phospho)proteomics, and transcriptome sequencing, we characterized BTK signaling networks. We show that BTK-dependent signaling is highly context dependent. In Fms-like tyrosine kinase 3 internal tandem duplication (FLT3-ITD)-positive AML, BTK mediates FLT3-ITDdependent Myc and STAT5 activation, and combined targeting of FLT3-ITD and BTK showed additive effects. In Fms-like tyrosine kinase 3 internal tandem duplication (FLT3-ITD)-negative AML, BTK couples Toll-like receptor 9 (TLR9) activation to nuclear factor kΒ and STAT5. Both BTK-dependent transcriptional programs were relevant for cell cycle progression and apoptosis regulation. Thus, we identify context-dependent oncogenic driver events that may guide subtype-specific treatment strategies and, for the first time, point to a role of TLR9 in AML.