Retinoic acid regulates avian lung branching through a molecular network

Retinoic acid (RA) is of major importance during vertebrate embryonic development and its levels need to be strictly regulated otherwise congenital malformations will develop. Through the action of specific nuclear receptors, named RAR/RXR, RA regulates the expression of genes that eventually influence proliferation and tissue patterning. RA has been described as crucial for different stages of mammalian lung morphogenesis, and as part of a complex molecular network that contributes to precise organogenesis; nonetheless, nothing is known about its role in avian lung development. The current report characterizes, for the first time, the expression pattern of RA signaling members (stra6, raldh2, raldh3, cyp26a1, rarα, and rarβ) and potential RA downstream targets (sox2, sox9, meis1, meis2, tgfβ2, and id2) by in situ hybridization. In the attempt of unveiling the role of RA in chick lung branching, in vitro lung explants were performed. Supplementation studies revealed that RA stimulates lung branching in a dose-dependent manner. Moreover, the expression levels of cyp26a1, sox2, sox9, rarβ, meis2, hoxb5, tgfβ2, id2, fgf10, fgfr2, and shh were evaluated after RA treatment to disclose a putative molecular network underlying RA effect. In situ hybridization analysis showed that RA is able to alter cyp26a1, sox9, tgfβ2, and id2 spatial distribution; to increase rarβ, meis2, and hoxb5 expression levels; and has a very modest effect on sox2, fgf10, fgfr2, and shh expression levels. Overall, these findings support a role for RA in the proximal–distal patterning and branching morphogenesis of the avian lung and reveal intricate molecular interactions that ultimately orchestrate branching morphogenesis.     

Genetic determinants of neuronal vulnerability to apoptosis

Apoptosis is a common mode of cell death that contributes to neuronal loss associated with neurodegeneration. Single-nucleotide polymorphisms (SNPs) in chromosomal DNA are contributing factors dictating natural susceptibility of humans to disease. Here, the most common SNPs affecting neuronal vulnerability to apoptosis are reviewed in the context of neurological disorders. Polymorphic variants in genes encoding apoptotic proteins, either from the extrinsic (FAS, TNF-α, CASP8) or the intrinsic (BAX, BCL2, CASP3, CASP9) pathways could be highly valuable in the diagnosis of neurodegenerative diseases and stroke. Interestingly, the Arg72Pro SNP in TP53, the gene encoding tumor suppressor p53, was recently revealed a biomarker of poor prognosis in stroke due to its ability to modulate neuronal apoptotic death. Search for new SNPs responsible for genetic variability to apoptosis will ensure the implementation of novel diagnostic and prognostic tools, as well as therapeutic strategies against neurological diseases.     

LI-cadherin cis-dimerizes in the plasma membrane Ca2+ independently and forms highly dynamic trans-contacts

LI-cadherin belongs to the family of 7D-cadherins that is characterized by a low sequence similarity to classical cadherins, seven extracellular cadherin repeats (ECs), and a short cytoplasmic domain. Nevertheless, LI-cadherins mediates Ca²⁺-dependent cell–cell adhesion and induces an epitheloid cellular phenotype in non-polarized CHO cells. Whereas several studies suggest that classical cadherins cis-dimerize in a Ca²⁺-dependent manner and interact in trans by strand-swapping tryptophan 2 of EC1, little is known about the molecular interactions of LI-cadherin, which lacks tryptophan 2. We thus expressed fluorescent LI-cadherin fusion proteins in HEK293 and CHO cells, analyzed their cell–cell adhesive properties and studied their cellular distribution, cis-interaction, and lateral diffusion in the presence and absence of Ca²⁺. LI-cadherin highly concentrates in cell contact areas but rapidly leaves those sites upon Ca²⁺ depletion and redistributes evenly on the cell surface, indicating that it is only kept in the contact areas by trans-interactions. Fluorescence resonance energy transfer analysis of LI-cadherin-CFP and -YFP revealed that LI-cadherin forms cis-dimers that resist Ca²⁺ depletion. As determined by fluorescence redistribution after photobleaching, LI-cadherin freely diffuses in the plasma membrane as a cis-dimer (D?=?0.42?±?0.03?μm²/s). When trapped by trans-binding in cell contact areas, its diffusion coefficient decreases only threefold to D?=?0.12?±?0.01?μm²/s, revealing that, in contrast to classical and desmosomal cadherins, trans-contacts formed by LI-cadherin are highly dynamic.     

<Emphasis Type="Italic">Cis</Emphasis>–<Emphasis Type="Italic">trans</Emphasis> interactions of cell surface receptors: biological roles and structural basis

Cell surface receptors bind ligands expressed on other cells (in trans) in order to communicate with neighboring cells. However, an increasing number of cell surface receptors are found to also interact with ligands expressed on the same cell (in cis). These observations raise questions regarding the biological role of such cis interactions. Specifically, it is important to know whether cis and trans binding have distinct functional effects and, if so, how a single cell discriminates between interactions in cis versus trans. Further, what are the structural features that allow certain cell surface receptors to engage ligand both on the same as well as on an apposed cell membrane? Here, we summarize known examples of receptors that display cis–trans binding and discuss the emerging diversity of biological roles played by these unconventional two-way interactions, along with their structural basis.     

Functional diversity of melanopsins and their global expression in the teleost retina

Melanopsin (OPN4) is an opsin photopigment that, in mammals, confers photosensitivity to retinal ganglion cells and regulates circadian entrainment and pupil constriction. In non-mammalian species, two forms of opn4 exist, and are classified into mammalian-like (m) and non-mammalian-like (x) clades. However, far less is understood of the function of this photopigment family. Here we identify in zebrafish five melanopsins (opn4m-1, opn4m-2, opn4m-3, opn4x-1 and opn4x-2), each encoding a full-length opsin G protein. All five genes are expressed in the adult retina in a largely non-overlapping pattern, as revealed by RNA in situ hybridisation and immunocytochemistry, with at least one melanopsin form present in all neuronal cell types, including cone photoreceptors. This raises the possibility that the teleost retina is globally light sensitive. Electrophysiological and spectrophotometric studies demonstrate that all five zebrafish melanopsins encode a functional photopigment with peak spectral sensitivities that range from 470 to 484 nm, with opn4m-1 and opn4m-3 displaying invertebrate-like bistability, where the retinal chromophore interchanges between cis- and trans-isomers in a light-dependent manner and remains within the opsin binding pocket. In contrast, opn4m-2, opn4x-1 and opn4x-2 are monostable and function more like classical vertebrate-like photopigments, where the chromophore is converted from 11-cis to all-trans retinal upon absorption of a photon, hydrolysed and exits from the binding pocket of the opsin. It is thought that all melanopsins exhibit an invertebrate-like bistability biochemistry. Our novel findings, however, reveal the presence of both invertebrate-like and vertebrate-like forms of melanopsin in the teleost retina, and indicate that photopigment bistability is not a universal property of the melanopsin family. The functional diversity of these teleost melanopsins, together with their widespread expression pattern within the retina, suggests that melanopsins confer global photosensitivity to the teleost retina and might allow for direct “fine-tuning” of retinal circuitry and physiology in the dynamic light environments found in aquatic habitats.     

Sauerstoff-Heteroringe. Die Konfiguration und Synthese desd,l-Homopterocarpins

Summary The racemic form of homopterocarpin (V), a constituent of red sandal-wood, has been synthesized by a method which has previously been proposed by the authors. N. M. R. spectral studies in conjunction with the examination of models have enabled the assignment of thecis configuration (V) to homopterocarpin.

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V. Mitteilung. IV. Mitteilung sieheH. Suginome, Exper.18, 161 (1962).     

Alcohol dehydrogenase 2 is a major hepatic enzyme for human retinol metabolism

The metabolism of all-trans- and 9-cis-retinol/ retinaldehyde has been investigated with focus on the activities of human, mouse and rat alcohol dehydrogenase 2 (ADH2), an intriguing enzyme with apparently different functions in human and rodents. Kinetic constants were determined with an HPLC method and a structural approach was implemented by in silico substrate dockings. For human ADH2, the determined K_m values ranged from 0.05 to 0.3 μM and k_cat values from 2.3 to 17.6 min⁻¹, while the catalytic efficiency for 9-cis-retinol showed the highest value for any substrate. In contrast, poor activities were detected for the rodent enzymes. A mouse ADH2 mutant (ADH2Pro47His) was studied that resembles the human ADH2 setup. This mutation increased the retinoid activity up to 100-fold. The K_m values of human ADH2 are the lowest among all known human retinol dehydrogenases, which clearly support a role in hepatic retinol oxidation at physiological concentrations. Received 12 October 2006; received after revision 6 December 2006; accepted 8 January 2007     

Retinoic acid signaling and neurogenic niche regulation in the developing peripheral nervous system of the cephalochordate amphioxus

The retinoic acid (RA) signaling pathway regulates axial patterning and neurogenesis in the developing central nervous system (CNS) of chordates, but little is known about its roles during peripheral nervous system (PNS) formation and about how these roles might have evolved. This study assesses the requirement of RA signaling for establishing a functional PNS in the cephalochordate amphioxus, the best available stand-in for the ancestral chordate condition. Pharmacological manipulation of RA signaling levels during embryogenesis reduces the ability of amphioxus larvae to respond to sensory stimulation and alters the number and distribution of ectodermal sensory neurons (ESNs) in a stage- and context-dependent manner. Using gene expression assays combined with immunohistochemistry, we show that this is because RA signaling specifically acts on a small population of soxb1c-expressing ESN progenitors, which form a neurogenic niche in the trunk ectoderm, to modulate ESN production during elongation of the larval body. Our findings reveal an important role for RA signaling in regulating neurogenic niche activity in the larval amphioxus PNS. Although only few studies have addressed this issue so far, comparable RA signaling functions have been reported for neurogenic niches in the CNS and in certain neurogenic placode derivatives of vertebrates. Accordingly, the here-described mechanism is likely a conserved feature of chordate embryonic and adult neural development.     

Isomériecis-trans des ménaquinones et oxydations phosphorylantes dans les extraits deMycobacterium phlei

Summary Incubation ofM. phlei washed cells with [¹⁴C³H₃]-l-methionine led to [2-¹⁴C³H₃] dihydromena-quinone-9 with an isotope ratio identical to that of methionine. Chromatography of the doubly labelled quinone indicated, despite a pronounced isotope effect, that bothcis andtrans isomers had the same isotope ratio. This result eliminates any possibility of hydrogen exchange in the 2-methyl group of menaquinones during oxydative phosphorylation, even in thecis isomer. Furthermore, it is confirmed that this compound is certainly formed from natural or synthetic menaquinones during isolation or incubation periods by the effect of daylight irradiation.     

Identification of (3R,4R)-gD 1(6) as an isolation artefact of cannabinoid acids formed by callus cultures ofCannabis sativa L

Summary For the first time we have isolated a major psychoactive cannabinoid, (3R, 4R)- ¹⁽⁶⁾-tetrahydrocannabinol3 from callus cultures ofCannabis sativa L.3 was obtained as an artefact of the actually formed (3R, 4R)- ¹-tetrahydrocannabinol-3- and/or 5-carboxylic acids1 and2 by subjecting the culture material to a decarboxylation step prior to extraction. No attempt was made to isolated acids1 and2. The identity of3 was confirmed by comparison with an authentic sample of (3R, 4R)- ¹-tetrahydrocannabinol. Culture conditions, isolation procedure and identification of the cannabinoid are described.Acknowledgment. The authors are greatly indebted to Dr W. Heller and Dr G. Schroeder-Frey for deriving the callus cultures from seedlings ofC. sativa and to Prof. N. Amrhein for valuable gifts of culture media and growth regulators.     

Tetraspanin TSPAN12 regulates tumor growth and metastasis and inhibits β-catenin degradation

Ablation of tetraspanin protein TSPAN12 from human MDA-MB-231 cells significantly decreased primary tumor xenograft growth, while increasing tumor apoptosis. Furthermore, TSPAN12 removal markedly enhanced tumor-endothelial interactions and increased metastasis to mouse lungs. TSPAN12 removal from human MDA-MB-231 cells also caused diminished association between FZD4 (a key canonical Wnt pathway receptor) and its co-receptor LRP5. The result likely explains substantially enhanced proteosomal degradation of β-catenin, a key effecter of canonical Wnt signaling. Consistent with disrupted canonical Wnt signaling, TSPAN12 ablation altered expression of LRP5, Naked 1 and 2, DVL2, DVL3, Axin 1, and GSKβ3 proteins. TSPAN12 ablation also altered expression of several genes regulated by β-catenin (e.g. CCNA1, CCNE2, WISP1, ID4, SFN, ME1) that may help to explain altered tumor growth and metastasis. In conclusion, these results provide the first evidence for TSPAN12 playing a role in supporting primary tumor growth and suppressing metastasis. TSPAN12 appears to function by stabilizing FZD4–LRP5 association, in support of canonical Wnt-pathway signaling, leading to enhanced β-catenin expression and function.     

Differences in luteinizing hormone release stimulated in rat anterior pituitary cells by leukotriene C4 and by gonadotropin-releasing hormone in vitro

Summary Continuous administration of leukotriene C₄ (LTC₄, 10^–10 M) to superfused rat anterior pituitary cells increased LH release for 40 min only, whereas in a parallel experiment gonadotropin-releasing hormone (GnRH, 10^–9 M) evoked a continuous increase in hormone secretion. In contrast to GnRH, LTC₄ did not desensitize rat anterior pituitary cells. The secretory action resulting from the administration of LTC₄ (10^–10 M) was abolished for 40 min after previous stimulation. The results documented the dual action of LTC₄ on LH exocytosis.     

Neural stem cells in Parkinson’s disease: a role for neurogenesis defects in onset and progression

Parkinson’s disease (PD) is the second most common neurodegenerative disorder, leading to a variety of motor and non-motor symptoms. Interestingly, non-motor symptoms often appear a decade or more before the first signs of motor symptoms. Some of these non-motor symptoms are remarkably similar to those observed in cases of impaired neurogenesis and several PD-related genes have been shown to play a role in embryonic or adult neurogenesis. Indeed, animal models deficient in Nurr1, Pitx3, SNCA and PINK1 display deregulated embryonic neurogenesis and LRRK2 and VPS35 have been implicated in neuronal development-related processes such as Wnt/β-catenin signaling and neurite outgrowth. Moreover, adult neurogenesis is affected in both PD patients and PD animal models and is regulated by dopamine and dopaminergic (DA) receptors, by chronic neuroinflammation, such as that observed in PD, and by differential expression of wild-type or mutant forms of PD-related genes. Indeed, an increasing number of in vivo studies demonstrate a role for SNCA and LRRK2 in adult neurogenesis and in the generation and maintenance of DA neurons. Finally, the roles of PD-related genes, SNCA, LRRK2, VPS35, Parkin, PINK1 and DJ-1 have been studied in NSCs, progenitor cells and induced pluripotent stem cells, demonstrating a role for some of these genes in stem/progenitor cell proliferation and maintenance. Together, these studies strongly suggest a link between deregulated neurogenesis and the onset and progression of PD and present strong evidence that, in addition to a neurodegenerative disorder, PD can also be regarded as a developmental disorder.     

Corticotropin releasing factor increases the adrenocortical responsiveness to adrenocorticotropin

Summary In the course of studying the plasma adrenocorticotropic hormone (ACTH) and corticosterone responses to synthetic corticotropin releasing factor (CRF), we noted some disparity in the responses. A higher dose (20 g compared with 5 g per rat i.a.) produced an equal plasma ACTH but greater plasma corticosterone response in adult male rats. Thus, we examined the possibility that CRF increases adrenocortical responsiveness to ACTH. CRF significantly (p<0.0005) increased the plasma corticosterone response to ACTH in rats pretreated with dexamethasone. Thus, synthetic CRF increases corticosterone secretion in rats not only by stimulating ACTH secretion, but also by increasing the adrenocortical responsiveness to ACTH.Acknowledgments. This research was supported by grant MT-5183 from the Medical Research Council of Canada and by the Department of Medicine, University of Kentucky.     

4,11-Epoxy-cis-eudesmane,soldier cephalic secretion of the Nearctic desert termite,Amitermes minimus Light (Termitidae: Termitinae)

Summary The soldier cephalic secretion of the Nearctic desert termite,Amitermes minimus, consists almost entirely of 4,11-epoxy-cis-eudesmane which was previously identified from soldiers of 2 AfricanAmitermes species. Soldiers ofA. minimus each store circa 61 g of the secretion. Bioassays with the ant,Crematogaster californica, indicate a repellent role for the eudesmane compound in termite defense.Classification as proposed by Sands, W.A., Bull. Br. Mus. nat. Hist., Ent., suppl.18 (1972).Supported by Sigma Xi Grant-in-Aid of Research and California Statewide Critical Applied Research grant.