The PTK7 and ROR2 Protein Receptors Interact in the Vertebrate WNT/Planar Cell Polarity (PCP) Pathway *

International audienceBackground: The planar cell polarity pathway plays important roles in morphogenetic processes. Results: PTK7 and ROR2 form a heterodimeric complex and bind to WNT5A, promoting JNK phosphorylation and regulating expression of paraxial protocadherin. Conclusion: PTK7 and ROR2 promote cell movement in mammalian cells and coordinate cell polarity during morphogenetic movements. Significance: We reveal new mechanisms of action of PTK7 in WNT/PCP signaling. The non-canonical WNT/planar cell polarity (WNT/PCP) pathway plays important roles in morphogenetic processes in vertebrates. Among WNT/PCP components, protein tyrosine kinase 7 (PTK7) is a tyrosine kinase receptor with poorly defined functions lacking catalytic activity. Here we show that PTK7 associates with receptor tyrosine kinase-like orphan receptor 2 (ROR2) to form a heterodimeric complex in mammalian cells. We demonstrate that PTK7 and ROR2 physically and functionally interact with the non-canonical WNT5A ligand, leading to JNK activation and cell movements. In the Xenopus embryo, Ptk7 functionally interacts with Ror2 to regulate protocadherin papc expression and morphogenesis. Furthermore , we show that Ptk7 is required for papc activation induced by Wnt5a. Interestingly, we find that Wnt5a stimulates the release of the tagged Ptk7 intracellular domain, which can translocate into the nucleus and activate papc expression. This study reveals novel molecular mechanisms of action of PTK7 in non-canonical WNT/PCP signaling that may promote cell and tissue movements

Effects of Aroclor 1254 on oxidative stress in developing Xenopus laevis tadpoles.

Over the last decades, amphibians decline has been reported worldwide. Exposure to polychlorinated biphenyls (PCBs) is one of the possible causes in addition to climate changes, UV-radiation or habitat destruction. In the present study, we tested the hypothesis that PCBs could induce oxidative stress in young tadpoles. Developing Xenopus laevis were exposed from 2- to 5-d postfertilization (pf) to 0.1 or 1 mg/l of Aroclor 1254. Lipid peroxidation and antioxidant systems (SOD, CAT, GST, GPx, GR activities and t-GSH level) were investigated in whole organisms. Exposure to both concentrations did not impact on the survival and development whereas the average body weight decreased. Exposure to 1 mg/l of Aroclor 1254 induced a significant (p<0.05) increase of GST activity when compared to controls 0 and DMSO. The other antioxidant enzymes and LPO evaluation remained unchanged. Our results demonstrate that exposure of X. laevis tadpoles to environmental concentrations of Aroclor 1254 interfere with normal growth. They also highlight that very young X. laevis tadpoles express antioxidant systems

Reproductive impacts of tributyltin (TBT) and triphenyltin (TPT) in the hermaphroditic freshwater gastropod Lymnaea stagnalis

Tributyltin (TBT) and triphenyltin (TPT) are emblematic endocrine disruptors, which have been mostly studied in gonochoric prosobranchs. Although both compounds can simultaneously occur in the environment, they have mainly been tested separately for their effects on snail reproduction. Because large discrepancies in experimental conditions occurred in these tests, the present study aimed at comparing the relative toxicity of TBT and TPT under similar laboratory conditions in the 0–600 ng Sn/L range. Tests were performed on the simultaneous hermaphrodite Lymnaea stagnalis, a freshwater snail in which effects of TPT were unknown. Survival, shell length and reproduction were monitored in a 21d semi-static test. Frequency of abnormal eggs was assessed as an additional endpoint. TPT hampered survival while TBT did not. Major effects on shell solidity and reproduction were observed for both compounds, reproductive outputs being more severely hampered by TBT than by TPT. Considering the frequency of abnormal eggs allowed increasing test sensitivity, since snail responses to TBT could be detected at concentrations as low as 19 ng Sn/L. However, the putative mode of action of the two compounds could not be deduced from the structure of the molecules or from the response of apical endpoints. Sensitivity of L. stagnalis to TBT and TPT was compared to the sensitivity of prosobranch molluscs with different habitats and different reproductive strategies

Effets de substances androgéniques et anti-androgéniques sur le gastéropodes dulçaquicole Lymnaea stagnalis

Knowledge on the impacts of endocrine disrupting chemicals (EDCs) on gastropods is scarce and their mechanisms of action are still poorly understood. In this study, effects of 3 androgens (tributyltin, testosterone and fenitrothion), 2 anti-androgens (cyproterone acetate and vinclozolin) and 1 estrogen (chlordecone) on growth and reproduction were investigated in the hermaphrodite gastropod Lymnaea stagnalis

The SCF/KIT pathway implements self-organised epithelial patterning by cell movement

How individual cell behaviours lead to the emergence of global patterns is poorly understood. In the Xenopus embryonic epidermis, multiciliated cells (MCCs) are born in a random pattern within an inner mesenchymal layer, and subsequently intercalate at regular intervals into an outer epithelial layer. Using both experiments and mathematical modelling, we show that this transition from chaotic to ordered distribution relies on mutual repulsion among motile immature MCCs, and affinity towards outer-layer intercellular junctions. Consistently, ARP2/3-mediated actin remodelling is required for MCC pattern emergence. Using multiple functional approaches, we show that the Kit tyrosine kinase receptor, expressed in MCCs, and its ligand Scf, expressed in outer-layer cells, are both required for regular MCC distribution. While Scf behaves as a potent adhesive cue for MCCs, Kit expression is sufficient to confer order to a disordered heterologous cell population. Our work reveals how a single signalling system can implement self-organised large-scale patterning

The Scf/Kit pathway implements self-organized epithelial patterning

International audienceHow individual cell behaviours lead to the emergence of global patterns is poorly understood. In the Xenopus embryonic epidermis, multiciliated cells (MCCs) are born in a random pattern within an inner mesenchymal layer, and subsequently intercalate at regular intervals into an outer epithelial layer. Using both experiments and mathematical modelling, we show that this transition from random to ordered distribution relies on mutual repulsion among motile immature MCCs, and affinity towards outer-layer intercellular junctions. Consistently, Arp2/3-mediated actin remodelling is required for MCC pattern emergence. Using multiple functional approaches, we show that the Kit tyrosine kinase receptor, expressed in MCCs, and its ligand Scf, expressed in outer-layer cells, are both required for regular MCC distribution. Membrane-associated Scf behaves as a potent adhesive cue for MCCs, while its soluble form promotes their mutual repulsion. On the other hand, Kit expression is sufficient to confer order to a disordered heterologous cell population. Our work reveals how a single signalling system can implement self-organised large-scale patterning

Myostatin promotes the terminal differentiation of embryonic muscle progenitors

Myostatin, a TGF-β family member, is an important regulator of adult muscle size. While extensively studied in vitro, the mechanisms by which this molecule mediates its effect in vivo are poorly understood. We addressed this question using chick and mouse embryos. We show that while myostatin overexpression in chick leads to an exhaustion of the muscle progenitor population that ultimately results in muscle hypotrophy, myostatin loss of function in chick and mouse provokes an expansion of this population. Our data demonstrate that myostatin acts in vivo to regulate the balance between proliferation and differentiation of embryonic muscle progenitors by promoting their terminal differentiation through the activation of p21 and MyoD. Previous studies have suggested that myostatin imposes quiescence on muscle progenitors. Our data suggest that myostatin’s effect on muscle progenitors is more complex than previously realized and is likely to be context-dependent. We propose a novel model for myostatin mode of action in vivo, in which myostatin affects the balance between proliferation and differentiation of embryonic muscle progenitors by enhancing their differentiation

Reproductive impact and proteomic analysis of androgenic and anti-androgenic disruptors on the hermaphroditic freshwater gastropod Lymnaea stagnalis

In this study, e ects of two androgens (tributyltin and testosterone), one antiandrogen (cyproterone acetate) and one estrogen (chlordecone) on growth and reproduction were investigated in the hermaphrodite gastropod Lymnaea stagnalis. In this study, exposure to a range of concentrations (ng/l to μg/l) of each chemical was performed during 21 days. e number of clutches and the number of eggs per clutch were monitored. A decrease in clutches laid per snail was observed after exposure to tributyltin (540, 1180 and 2600 ng/l) and chlordecone (10, 22, 50 and 110 μg/l). A signi cant decrease in egg laid per snail was observed after exposure to tributyltin (540, 1180 and 2600 ng/l) and chlordecone (50 and 110 μg/l). An increase of egg abnormalities ratio in exposed snails (atrophied albumen, polyembryonicity,...) was observed in L. stagnalis following exposure to testosterone (2, 22, 50 and 110 ng/l), cyproterone acetate (4,5 and 50 μg/l), tributyltin (110 and 244 ng/l) and chlordecone (4,5 and 10 μg/l). Investigation of alteration in protein expression in exposed snails was performed using proteomic analysis such as 2D-DIGE. Mass spectrometry identi cation was performed on proteins with altered expression. We could establish correlation between reproductive endpoints and changes in proteins involved in egg formation and in egg laying were underlined. Egg yolk ferritin, the main protein of egg yolk, was shown to be reduced signi cantly in relationship with a decrease of egg yolk quality after exposure to tributyltin 540 ng/l and cyproterone acetate 4,5 μg/l. Ovipostatin, a protein proved to reduce egg masses, was signi cantly over expressed in snails exposed to 50 μg/l of chlordecone and were in relationship with a reduction of clutches laid by individuals. Further western blot analysis on those proteins involved in the reproduction are underway. ese analysis will enable us to con rm and re ne with more speci city the 2D-DIGE results for the selected proteins. e results of this study can help to establish new biomarkers of exposure of endocrine disruptors in freshwater environment and can provide new insight on mode of action of endocrine disruptors in L. stagnalis

Lrrcc1 and Ccdc61 are conserved effectors of multiciliated cell function

International audienceCiliated epithelia perform essential functions across animal evolution, ranging from locomotion of marine organisms to mucociliary clearance of airways in mammals. These epithelia are composed of multiciliated cells (MCCs) harbouring myriads of motile cilia, which rest on modified centrioles called basal bodies (BBs), and beat coordinately to generate directed fluid flows. Thus, BB biogenesis and organization is central to MCC function. In basal eukaryotes, the coiled-coil domain proteins Lrrcc1 and Ccdc61 were shown to be required for proper BB construction and function. Here, we used the Xenopus embryonic ciliated epidermis to characterize Lrrcc1 and Ccdc61 in vertebrate MCCs. We found that they both encode BB components, localized proximally at the junction with striated rootlets. Knocking down either gene caused defects in BB docking, spacing, and polarization. Moreover, their depletion impaired the apical cytoskeleton, and altered ciliary beating. Consequently, cilia-powered fluid flow was greatly reduced in morphant tadpoles, which displayed enhanced mortality when exposed to pathogenic bacteria. This work illustrates how integration across organizational scales make elementary BB components essential for the emergence of the physiological function of ciliated epithelia