The biosynthesis of gibberellins (GAs) after GA12-aldehyde involves a series of oxidative steps that lead to the formation of bioactive GAs. Previously, a cDNA clone encoding a GA 20-oxidase [gibberellin, 2-oxoglutarate:oxygen oxidoreductase (20-hydroxylating, oxidizing), EC 1.14. point mutation that inserts a translational stop codon in the protein-coding sequence, thus confirming that the GAS locus encodes GA 20-oxidase. Expression of the GAS gene in Arabidopsis leaves was enhanced after plants were transferred from short to long days; it was reduced by GA4 treatment, suggesting end-product repression in the GA biosynthetic pathway.The gibberellins (GAs) are tetracylic diterpenoid compounds that play an important role in many aspects of plant growth and development, such as promotion of cell division and extension, seed germination, stem growth, and fruit set (1). The biosynthesis of GAs has been studied extensively in developing seeds (2, 3), but their function in immature seeds, if any, is unknown. In contrast, relatively little is known about GA biosynthesis in vegetative tissues, because GA levels in these tissues are low compared with levels in immature seeds, and active enzyme preparations for GA conversions are difficult to obtain. It is, therefore, desirable to extend studies on regulation of GA biosynthesis to green plants in which GAs have definite roles. One of these roles is to mediate photoperiodic control of stem elongation in rosette plants (4-6). Therefore, to understandThe publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact. 6640the mechanism by which long days (LD) cause stem elongation, it is necessary to elucidate how daylength regulates GA biosynthesis at the biochemical and molecular levels.In Arabidopsis thaliana, a long-day plant (LDP) suitable for molecular genetic studies (7), a number of GA-responsive dwarf mutants have been isolated (8). It has been demonstrated that one of these mutants carrying thegaS mutation has reduced levels of Cl-GAs, indicating that the product of the GAS gene catalyzes elimination of C-20 at the aldehyde level in the GA biosynthetic pathway. The gaS dwarf mutant also has increased levels of certain C20-GAs, indicating existence of an additional control, possibly hydroxylation of C-20 (9). This suggests that the GAS locus encodes a GA 20-oxidase [gibberellin, 2-oxoglutarate:oxygen oxidoreductase (20-hydroxylating, oxidizing), EC 1.14.11.-]. In pumpkin endosperm, the enzyme activities for the steps between GA53 and GA20 ( Fig. 1) reside on a single polypeptide (10), indicating that GA 20-oxidase is a multifunctional enzyme that catalyzes the oxidation and elimination of C-20 and thus plays a pivotal role in the conversion of C20-to C19-GAs. Recently, cDNA clones encoding GA 20-oxidases that differ by one amino acid have been isolated from developing cotyledons (11) and liquid endosperm (12) of devel...

If neuropilin-2 and the growth factor VEGF-C don’t come together, lymphatic vessels don’t branch apart.

Neural stem cells (NSCs) are slowly dividing astrocytes that are intimately associated with capillary endothelial cells in the subventricular zone (SVZ) of the brain. Functionally, members of the vascular endothelial growth factor (VEGF) family can stimulate neurogenesis as well as angiogenesis, but it has been unclear whether they act directly via VEGF receptors (VEGFRs) expressed by neural cells, or indirectly via the release of growth factors from angiogenic capillaries. Here, we show that VEGFR-3, a receptor required for lymphangiogenesis, is expressed by NSCs and is directly required for neurogenesis. Vegfr3:YFP reporter mice show VEGFR-3 expression in multipotent NSCs, which are capable of self-renewal and are activated by the VEGFR-3 ligand VEGF-C in vitro. Overexpression of VEGF-C stimulates VEGFR-3-expressing NSCs and neurogenesis in the SVZ without affecting angiogenesis. Conversely, conditional deletion of Vegfr3 in neural cells, inducible deletion in subventricular astrocytes, and blocking of VEGFR-3 signaling with antibodies reduce SVZ neurogenesis. Therefore, VEGF-C/VEGFR-3 signaling acts directly on NSCs and regulates adult neurogenesis, opening potential approaches for treatment of neurodegenerative diseases.

Rationale The lymphatic vasculature plays a major role in fluid homeostasis, absorption of dietary lipids, and immune surveillance. Fluid transport depends on the presence of intraluminal valves within lymphatic collectors. Defective formation of lymphatic valves leads to lymphedema, a progressive and debilitating condition for which curative treatments are currently unavailable. How lymphatic valve formation is regulated remains largely unknown. Objective We investigated if the repulsive axon guidance molecule Semaphorin3A (Sema3A) plays a role in lymphatic valve formation. Methods and Results We show that Sema3A mRNA is expressed in lymphatic vessels and that Sema3A protein binds to lymphatic valves expressing the Neuropilin-1 (Nrp1) and PlexinA1 receptors. Using mouse knockout models, we show that Sema3A is selectively required for lymphatic valve formation, via interaction with Nrp1 and PlexinA1. Sema3a−/− mice exhibit defects in lymphatic valve formation, which are not due to abnormal lymphatic patterning or sprouting, and mice carrying a mutation in the Sema3A binding site of Nrp1, or deficient for Plxna1, develop lymphatic valve defects similar to those seen in Sema3a−/− mice. Conclusions Our data demonstrate an essential direct function of Sema3A-Nrp1-PlexinA1 signaling in lymphatic valve formation.

No abstract

Cranial lymphatic vessels (LVs) are involved in the transport of fluids, macromolecules and central nervous system (CNS) immune responses. Little information about spinal LVs is available, because these delicate structures are embedded within vertebral tissues and difficult to visualize using traditional histology. Here we show an extended vertebral column LV network using three-dimensional imaging of decalcified iDISCO+-clarified spine segments. Vertebral LVs connect to peripheral sensory and sympathetic ganglia and form metameric vertebral circuits connecting to lymph nodes and the thoracic duct. They drain the epidural space and the dura mater around the spinal cord and associate with leukocytes. Vertebral LVs remodel extensively after spinal cord injury and VEGF-C-induced vertebral lymphangiogenesis exacerbates the inflammatory responses, T cell infiltration and demyelination following focal spinal cord lesion. Therefore, vertebral LVs add to skull meningeal LVs as gatekeepers of CNS immunity and may be potential targets to improve the maintenance and repair of spinal tissues.

Neutrophils are critically involved in host defense and tissue damage. Intrinsic signal mechanisms controlling neutrophil activities are poorly defined. We found that the expression of wild-type p53-induced phosphatase 1 (Wip1) in mouse and human neutrophils was downregulated quickly after neutrophil activation through JNK-microRNA-16 pathway. Importantly, the Wip1 expression level was negatively correlated with inflammatory cytokine productions of neutrophils in sepsis patients. Wip1-deficient mice displayed increased bactericidal activities to Staphylococcus aureus and were hypersensitive to LPS-induced acute lung damage with increased neutrophil infiltration and inflammation. Mechanism studies showed that the enhanced inflammatory activity of neutrophils caused by Wip1 deficiency was mediated by p38 MAPK-STAT1 and NF-κB pathways. The increased migration ability of Wip1KO neutrophils was mediated by the decreased CXCR2 internalization and desensitization, which was directly regulated by p38 MAPK activity. Thus, our findings identify a previously unrecognized function of Wip1 as an intrinsic negative regulator for neutrophil proinflammatory cytokine production and migration through multiple signal pathways.

Blood-brain barrier (BBB) integrity is critical for proper function of the central nervous system (CNS). Here, we show that the endothelial Unc5B receptor controls BBB integrity by maintaining Wnt/β-catenin signaling. Inducible endothelial-specific deletion of Unc5B in adult mice leads to BBB leak from brain capillaries that convert to a barrier-incompetent state with reduced Claudin-5 and increased PLVAP expression. Loss of Unc5B decreases BBB Wnt/β-catenin signaling, and β-catenin overexpression rescues Unc5B mutant BBB defects. Mechanistically, the Unc5B ligand Netrin-1 enhances Unc5B interaction with the Wnt co-receptor LRP6, induces its phosphorylation and activates Wnt/β-catenin downstream signaling. Intravenous delivery of antibodies blocking Netrin-1 binding to Unc5B causes a transient BBB breakdown and disruption of Wnt signaling, followed by neurovascular barrier resealing. These data identify Netrin-1-Unc5B signaling as a ligand-receptor pathway that regulates BBB integrity, with implications for CNS diseases.