Aberrant neovascularization contributes to diseases such as cancer, blindness and atherosclerosis, and is the consequence of inappropriate angiogenic signalling. Although many regulators of pathogenic angiogenesis have been identified, our understanding of this process is incomplete. Here we explore the transcriptome of retinal microvessels isolated from mouse models of retinal disease that exhibit vascular pathology, and uncover an upregulated gene, leucine-rich alpha-2-glycoprotein 1 (Lrg1), of previously unknown function. We show that in the presence of transforming growth factor-β1 (TGF-β1), LRG1 is mitogenic to endothelial cells and promotes angiogenesis. Mice lacking Lrg1 develop a mild retinal vascular phenotype but exhibit a significant reduction in pathological ocular angiogenesis. LRG1 binds directly to the TGF-β accessory receptor endoglin, which, in the presence of TGF-β1, results in promotion of the pro-angiogenic Smad1/5/8 signalling pathway. LRG1 antibody blockade inhibits this switch and attenuates angiogenesis. These studies reveal a new regulator of angiogenesis that mediates its effect by modulating TGF-β signalling.

Nesprins form a novel class of nuclear envelope-anchored spectrin-repeat proteins. We show that a direct association of their highly conserved C-terminal luminal domain with the inner nuclear membrane protein Sun1 mediates their nuclear envelope localisation. In Nesprin-1 and Nesprin-2 the conserved C-terminal amino acids PPPX are essential for the interaction with a C-terminal region in Sun1. In fact, Sun1 is required for the proper nuclear envelope localisation of Nesprin-2 as shown using dominant-negative mutants and by knockdown of Sun1 expression. Sun1 itself does not require functional A-type lamins for its localisation at the inner nuclear membrane in mammalian cells. Our findings propose a conserved nuclear anchorage mechanism between Caenorhabditis elegans and mammals and suggest a model in which Sun1 serves as a `structural bridge' connecting the nuclear interior with the actin cytoskeleton.

The vertebrate proteins Nesprin-1 and Nesprin-2 (also referred to as Enaptin and NUANCE) together with ANC-1 of Caenorhabditis elegans and MSP-300 of Drosophila melanogaster belong to a novel family of α-actinin type actin-binding proteins residing at the nuclear membrane. Using biochemical techniques, we demonstrate that Nesprin-2 binds directly to emerin and the C-terminal common region of lamin A/C. Selective disruption of the lamin A/C network in COS7 cells, using a dominant negative lamin B mutant, resulted in the redistribution of Nesprin-2. Furthermore, using lamin A/C knockout fibroblasts we show that lamin A/C is necessary for the nuclear envelope localization of Nesprin-2. In normal skin where lamin A/C is differentially expressed, strong Nesprin-2 expression was found in all epidermal layers, including the basal layer where only lamin C is present. This indicates that lamin C is sufficient for proper Nesprin-2 localization at the nuclear envelope. Expression of dominant negative Nesprin-2 constructs and knockdown studies in COS7 cells revealed that the presence of Nesprin-2 at the nuclear envelope is necessary for the proper localization of emerin. Our data imply a scaffolding function of Nesprin-2 at the nuclear membrane and suggest a potential involvement of this multi-isomeric protein in human disease.

Growth, maturation, and integrity of the blood vessel network require extensive communication between the endothelial cells, which line the vascular lumen, and associated mural cells, namely vascular smooth muscle cells and pericytes. Pericytes extend long processes, make direct contact with the capillary endothelium, and promote vascular quiescence by suppressing angiogenic sprouting. Vascular smooth muscle cells are highly contractile, extracellular matrix–secreting cells that cover arteries and veins and provide them with mechanical stability and elasticity. In the damaged blood vessel wall, for example in atherosclerotic lesions, vascular smooth muscle cells lose their differentiated state and acquire a highly mitotic, so-called “synthetic” phenotype, which is thought to promote pathogenesis. Among other factors, extracellular matrix molecules and integrin family cell–matrix receptors may regulate this phenotypic transition. Here we show that the inactivation of the gene encoding the integrin β1 subunit ( Itgb1 ) with a Cre-loxP approach in mice leads to mural cell defects and postnatal lethality. Integrin β1–deficient vascular smooth muscle cells display several hallmarks of the synthetic phenotype: Cell proliferation is enhanced, whereas differentiation and their ability to support blood vessels are compromised. Similarly, mutant pericytes are poorly spread but present in larger numbers. Our analysis of this mutant model shows that integrin β1–mediated cell–matrix adhesion is a major determinant of the mural cell phenotype.