Precisely controlled gene regulatory networks are required during embryonic development to give rise to various structures, including those of the cardiovascular system. Long non-coding RNA (lncRNA) ...loci are known to be important regulators of these genetic programs. We have identified a novel and essential lncRNA locus Handsdown (Hdn), active in early heart cells, and show by genetic inactivation that it is essential for murine development. Hdn displays haploinsufficiency for cardiac development as Hdn-heterozygous adult mice exhibit hyperplasia in the right ventricular wall. Transcriptional activity of the Hdn locus, independent of its RNA, suppresses its neighboring gene Hand2. We reveal a switch in a topologically associated domain in differentiation of the cardiac lineage, allowing the Hdn locus to directly interact with regulatory elements of the Hand2 locus.
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•The essential lncRNA Handsdown is haploinsufficient for embryonic development•The Handsdown RNA transcript is dispensable for cardiac gene regulation•The TAD association of Handsdown shifts during cardiac differentiation•Transcription of Handsdown regulates the cis-located Hand2 gene
Ritter et al. characterize an essential lncRNA locus, Handsdown, which regulates development by controlling cardiac gene programs during early vertebrate embryo formation. The Handsdown locus physically interacts with the Hand2 gene during cardiac differentiation and genetic ablation of Handsdown leads to elevated Hand2 levels.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Abstract
After myocardial infarction in the adult heart the remaining, non-infarcted tissue adapts to compensate the loss of functional tissue. This adaptation requires changes in gene expression ...networks, which are mostly controlled by transcription regulating proteins. Long non-coding transcripts (lncRNAs) are taking part in fine-tuning such gene programs. We describe and characterize the cardiomyocyte specific lncRNA
Sweetheart RNA
(
Swhtr
), an approximately 10 kb long transcript divergently expressed from the cardiac core transcription factor coding gene
Nkx2-5
. We show that
Swhtr
is dispensable for normal heart development and function but becomes essential for the tissue adaptation process after myocardial infarction in murine males. Re-expressing
Swhtr
from an exogenous locus rescues the
Swhtr null
phenotype. Genes that depend on
Swhtr
after cardiac stress are significantly occupied and therefore most likely regulated by NKX2-5. The
Swhtr
transcript interacts with NKX2-5 and disperses upon hypoxic stress in cardiomyocytes, indicating an auxiliary role of
Swhtr
for NKX2-5 function in tissue adaptation after myocardial injury.
Summary
We have recently identified endothelial cell-secreted developmental endothelial locus-1 (Del-1) as an endogenous inhibitor of β2-integrin–dependent leukocyte infiltration. Del-1 was ...previously also implicated in angiogenesis. Here, we addressed the role of endogenously produced Del-1 in ischaemia-related angiogenesis. Intriguingly, Del-1–deficient mice displayed increased neovascularisation in two independent ischaemic models (retinopathy of prematurity and hind-limb ischaemia), as compared to Del-1–proficient mice. On the contrary, angiogenic sprouting
in vitro
or
ex vivo
(aortic ring assay) and physiological developmental retina angiogenesis were not affected by Del-1 deficiency. Mechanistically, the enhanced ischaemic neovascularisation in Del-1-deficiency was linked to higher infiltration of the ischaemic tissue by CD45+ haematopoietic and immune cells. Moreover, Del-1-deficiency promoted β2-integrin–dependent adhesion of haematopoietic cells to endothelial cells
in vitro
, and the homing of hematopoietic progenitor cells and of immune cell populations to ischaemic muscles
in vivo
. Consistently, the increased hind limb ischaemia-related angiogenesis in Del-1 deficiency was completely reversed in mice lacking both Del-1 and the β2-integrin LFA-1. Additionally, enhanced retinopathy-associated neovascularisation in Del-1-deficient mice was reversed by LFA-1 blockade. Our data reveal a hitherto unrecognised function of endogenous Del-1 as a local inhibitor of ischaemia-induced angiogenesis by restraining LFA-1–dependent homing of pro-angiogenic haematopoietic cells to ischaemic tissues. Our findings are relevant for the optimisation of therapeutic approaches in the context of ischaemic diseases.
Supplementary Material to this article is available online at www.thrombosis-online.com.
Congenital lower urinary-tract obstruction (LUTO) is caused by anatomical blockage of the bladder outflow tract or by functional impairment of urinary voiding. About three out of 10,000 pregnancies ...are affected. Although several monogenic causes of functional obstruction have been defined, it is unknown whether congenital LUTO caused by anatomical blockage has a monogenic cause. Exome sequencing in a family with four affected individuals with anatomical blockage of the urethra identified a rare nonsense variant (c.2557C>T p.Arg853∗) in BNC2, encoding basonuclin 2, tracking with LUTO over three generations. Re-sequencing BNC2 in 697 individuals with LUTO revealed three further independent missense variants in three unrelated families. In human and mouse embryogenesis, basonuclin 2 was detected in lower urinary-tract rudiments. In zebrafish embryos, bnc2 was expressed in the pronephric duct and cloaca, analogs of the mammalian lower urinary tract. Experimental knockdown of Bnc2 in zebrafish caused pronephric-outlet obstruction and cloacal dilatation, phenocopying human congenital LUTO. Collectively, these results support the conclusion that variants in BNC2 are strongly implicated in LUTO etiology as a result of anatomical blockage.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
We have recently identified endothelial cell-secreted developmental endothelial locus-1 (Del-1) as an endogenous inhibitor of β
2
-integrin–dependent leukocyte infiltration. Del-1 was previously also ...implicated in angiogenesis. Here, we addressed the role of endogenously produced Del-1 in ischemia-related angiogenesis. Intriguingly, Del-1–deficient mice displayed increased neovascularization in two independent ischemic models (retinopathy of prematurity and hind-limb ischemia), as compared to Del-1–proficient mice. On the contrary, angiogenic sprouting
in vitro
or
ex vivo
(aortic ring assay) and physiological developmental retina angiogenesis were not affected by Del-1 deficiency. Mechanistically, the enhanced ischemic neovascularization in Del-1-deficiency was linked to higher infiltration of the ischemic tissue by CD45
+
hematopoietic and immune cells. Moreover, Del-1-deficiency promoted β
2
-integrin–dependent adhesion of hematopoietic cells to endothelial cells
in vitro
, and the homing of hematopoietic progenitor cells and of immune cell populations to ischemic muscles
in vivo
. Consistently, the increased hind limb ischemia-related angiogenesis in Del-1 deficiency was completely reversed in mice lacking both Del-1 and the β
2
-integrin LFA-1. Additionally, enhanced retinopathy-associated neovascularization in Del-deficient mice was reversed by LFA-1 blockade. Our data reveal a hitherto unrecognized function of endogenous Del-1 as a local inhibitor of ischemia-induced angiogenesis by restraining LFA-1–dependent homing of pro-angiogenic hematopoietic cells to ischemic tissues. Our findings are relevant for the optimization of therapeutic approaches in the context of ischemic diseases.
β1-integrins are mediating endothelial cell (EC) adhesion to extracellular matrix proteins and are critical mediators of neovascularization. In addition, intracellular signaling stimulating integrin ...affinity promotes angiogenesis. However, the role of intracellular integrin affinity inhibitors in angiogenesis remains elusive. SHARPIN is a negative regulator of integrin affinity in fibroblasts and leukocytes by binding to several α-integrin subunits. Nevertheless, the role of SHARPIN in EC biology and its contribution to angiogenesis is yet unknown. According to our findings, EC express SHARPIN. Silencing of SHARPIN with siRNA significantly reduced angiogenic sprouting of EC spheroids in collagen gels and inhibited tube formation in matrigel assays (by 41 ± 4 %) in comparison to scrambled siRNA-transfected EC. An essential angiogenic function of EC is chemotactic migration. In real-time video-microscopy chemotaxis assays, SHARPIN silencing almost abolished the ability of EC to follow chemotactic VEGF-gradients in comparison to scrambled siRNA-transfected EC. Furthermore, EC migration is dependent on integrin-mediated adhesion. Silencing of SHARPIN significantly increased β1-integrin-dependent adhesion of EC on fibronectin and collagen. In line with these results, silencing of SHARPIN reduced the abundance of the inactive conformation of β1-integrins on the EC surface as assessed by flow cytometry, while the total protein expression levels of β1-integrins remained unaltered. Interestingly, inhibition of β1-integrins with a neutralizing antibody (at low concentration) restored the ability of SHARPIN siRNA-transfected EC to follow chemotactic VEGF-gradients and significantly enhanced angiogenic sprouting in comparison to SHARPIN siRNA-transfected EC treated with the isotype control antibody. To address the in vivo relevance of our findings, we employed the murine model of post-natal retinal angiogenesis. SHARPIN-deficient mice displayed significantly lower vessel density and less vessel branching points in the retinal plexus when compared to the wild type mice. In conclusion, SHARPIN is a novel mediator of angiogenesis by promoting VEGF-induced EC chemotactic migration by downregulating β1-integrin-activity.
Neun Fachbeiträge dokumentieren die Ergebnisse der aktuellen Projekt- und Forschungsarbeit des Landesamtes in den Themenbereichen Grundwasser, Altlasten und Boden.
Neun Fachbeiträge dokumentieren die Ergebnisse der aktuellen Projekt- und Forschungsarbeit des Landesamtes in den Themenbereichen Grundwasser, Altlasten und Boden.
Among tool-using animals 1–4, none are known to adaptively change the hydrodynamic properties of a free jet of water—a task considered difficult in human technology 5–7. Hunting archerfish can strike ...their targets with precisely aimed water jets (e.g., 8, 9), but they are also presently thought to be unable to actively control the hydrodynamics of their jets 8–13. By using specifically trained fish, we were able to monitor several aspects of jet production and propagation as the fish fired at targets over a much wider range of distances than previously explored 10, 13. We show that jets that have to travel farther also live longer. Furthermore, the time needed until water assembles at the jet tip is not fixed. Rather, it is adjusted so that maximum focusing occurs just before impact. Surprisingly, the fish achieve this by modulating the dynamics of changes in the cross-section of their mouth opening, a mechanism that seems to not have been applied yet in human-built nozzles. The timing adjustments archerfish make in order to powerfully hit targets over an extended range strikingly parallel the situation in the “uniquely human” ability of powerful throwing 14–18. Based on the key role throwing played in human encephalization and cognitive evolution 14–20, skillfully “throwing” water should similarly have led to the correlated rapid evolution of cognitive skills in this animal.
•Archerfish shape the hydrodynamics of their jets to adjust them to target distance•Shaping involves adjustments in the dynamics of mouth opening and closing•As in human throwing, an extended striking range requires timing control
Archerfish shoot down target prey with precisely aimed water jets. Gerullis and Schuster show that by using their mouths as active nozzles that continuously change width, archerfish are able to actively shape the hydrodynamics of their jets so that they can powerfully strike prey over an extended range.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
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