Autoinhibition of c-Abl

AbstractDespite years of investigation, the molecular mechanism responsible for regulation of the c-Abl tyrosine kinase has remained elusive. We now report inhibition of the catalytic activity of purified c-Abl in vitro, demonstrating that regulation is an intrinsic property of the molecule. We show that the interaction of the N-terminal 80 residues with the rest of the protein mediates autoregulation. This N-terminal “cap” is required to achieve and maintain inhibition, and its loss turns c-Abl into an oncogenic protein and contributes to deregulation of BCR-Abl

BDNF promotes target innervation of Xenopus mandibular trigeminal axons in vivo

<p>Abstract</p> <p>Background</p> <p>Trigeminal nerves consist of ophthalmic, maxillary, and mandibular branches that project to distinct regions of the facial epidermis. In <it>Xenopus </it>embryos, the mandibular branch of the trigeminal nerve extends toward and innervates the cement gland in the anterior facial epithelium. The cement gland has previously been proposed to provide a short-range chemoattractive signal to promote target innervation by mandibular trigeminal axons. Brain derived neurotrophic factor, BDNF is known to stimulate axon outgrowth and branching. The goal of this study is to determine whether BDNF functions as the proposed target recognition signal in the <it>Xenopus </it>cement gland.</p> <p>Results</p> <p>We found that the cement gland is enriched in BDNF mRNA transcripts compared to the other neurotrophins NT3 and NT4 during mandibular trigeminal nerve innervation. BDNF knockdown in <it>Xenopus </it>embryos or specifically in cement glands resulted in the failure of mandibular trigeminal axons to arborise or grow into the cement gland. BDNF expressed ectodermal grafts, when positioned in place of the cement gland, promoted local trigeminal axon arborisation <it>in vivo</it>.</p> <p>Conclusion</p> <p>BDNF is necessary locally to promote end stage target innervation of trigeminal axons <it>in vivo</it>, suggesting that BDNF functions as a short-range signal that stimulates mandibular trigeminal axon arborisation and growth into the cement gland.</p

Abelson Phosphorylation of CLASP2 Modulates its Association With Microtubules and Actin

The Abelson (Abl) non-receptor tyrosine kinase regulates the cytoskeleton during multiple stages of neural development, from neurulation, to the articulation of axons and dendrites, to synapse formation and maintenance. We previously showed that Abl is genetically linked to the microtubule (MT) plus end tracking protein (+TIP) CLASP in Drosophila. Here we show in vertebrate cells that Abl binds to CLASP and phosphorylates it in response to serum or PDGF stimulation. In vitro, Abl phosphorylates CLASP with a Km of 1.89 µM, indicating that CLASP is a bona fide substrate. Abl-phosphorylated tyrosine residues that we detect in CLASP by mass spectrometry lie within previously mapped F-actin and MT plus end interaction domains. Using purified proteins, we find that Abl phosphorylation modulates direct binding between purified CLASP2 with both MTs and actin. Consistent with these observations, Abl-induced phosphorylation of CLASP2 modulates its localization as well as the distribution of F-actin structures in spinal cord growth cones. Our data suggest that the functional relationship between Abl and CLASP2 is conserved and provides a means to control the CLASP2 association with the cytoskeleton. © 2014 The Authors. Cytoskeleton Published by Wiley Periodicals, Inc

Foxm1 regulates neural progenitor fate during spinal cord regeneration

From Wiley via Jisc Publications RouterHistory: received 2020-05-20, rev-recd 2021-06-24, accepted 2021-07-01, pub-electronic 2021-08-24Article version: VoRPublication status: PublishedFunder: Wellcome Trust; Grant(s): 205894/Z/17/ZFunder: Biotechnology and Biological Sciences Research Council Research Training Support; Id: http://dx.doi.org/10.13039/501100000268; Grant(s): BB/M011208/1Funder: UKRI|Medical Research Council (MRC); Id: http://dx.doi.org/10.13039/501100000265; Grant(s): MR/M008908/1Funder: Wellcome Trust (ISSF fund)Abstract: Xenopus tadpoles have the ability to regenerate their tails upon amputation. Although some of the molecular and cellular mechanisms that globally regulate tail regeneration have been characterised, tissue‐specific response to injury remains poorly understood. Using a combination of bulk and single‐cell RNA sequencing on isolated spinal cords before and after amputation, we identify a number of genes specifically expressed in the spinal cord during regeneration. We show that Foxm1, a transcription factor known to promote proliferation, is essential for spinal cord regeneration. Surprisingly, Foxm1 does not control the cell cycle length of neural progenitors but regulates their fate after division. In foxm1−/− tadpoles, we observe a reduction in the number of neurons in the regenerating spinal cord, suggesting that neuronal differentiation is necessary for the regenerative process. Altogether, our data uncover a spinal cord‐specific response to injury and reveal a new role for neuronal differentiation during regeneration

Efa6 protects axons and regulates their growth and branching by inhibiting microtubule polymerisation at the cortex

Cortical collapse factors affect microtubule (MT) dynamics at the plasma membrane. They play important roles in neurons, as suggested by inhibition of axon growth and regeneration through the ARF activator Efa6 in C. elegans, and by neurodevelopmental disorders linked to the mammalian kinesin Kif21A. How cortical collapse factors influence axon growth is little understood. Here we studied them, focussing on the function of Drosophila Efa6 in experimentally and genetically amenable fly neurons. First, we show that Drosophila Efa6 can inhibit MTs directly without interacting molecules via an N-terminal 18 amino acid motif (MT elimination domain/MTED) that binds tubulin and inhibits microtubule growth in vitro and cells. If N-terminal MTED-containing fragments are in the cytoplasm they abolish entire microtubule networks of mouse fibroblasts and whole axons of fly neurons. Full-length Efa6 is membrane-attached, hence primarily blocks MTs in the periphery of fibroblasts, and explorative MTs that have left axonal bundles in neurons. Accordingly, loss of Efa6 causes an increase of explorative MTs: In growth cones they enhance axon growth, in axon shafts they cause excessive branching, as well as atrophy through perturbations of MT bundles. Efa6 over-expression causes the opposite phenotypes. Taken together, our work conceptually links molecular and sub-cellular functions of cortical collapse factors to axon growth regulation and reveals new roles in axon branching and in the prevention of axonal atrophy. Furthermore, the MTED delivers a promising tool that can be used to inhibit MTs in a compartmentalised fashion when fusing it to specifically localising protein domains

Amputation-induced reactive oxygen species are required for successful Xenopus tadpole tail regeneration.

Understanding the molecular mechanisms that promote successful tissue regeneration is critical for continued advancements in regenerative medicine. Vertebrate amphibian tadpoles of the species Xenopus laevis and Xenopus tropicalis have remarkable abilities to regenerate their tails following amputation, through the coordinated activity of numerous growth factor signalling pathways, including the Wnt, Fgf, Bmp, Notch and TGF-β pathways. Little is known, however, about the events that act upstream of these signalling pathways following injury. Here, we show that Xenopus tadpole tail amputation induces a sustained production of reactive oxygen species (ROS) during tail regeneration. Lowering ROS levels, using pharmacological or genetic approaches, reduces the level of cell proliferation and impairs tail regeneration. Genetic rescue experiments restored both ROS production and the initiation of the regenerative response. Sustained increased ROS levels are required for Wnt/β-catenin signalling and the activation of one of its main downstream targets, fgf20 (ref. 7), which, in turn, is essential for proper tail regeneration. These findings demonstrate that injury-induced ROS production is an important regulator of tissue regeneration

MECANISMES MOLECULAIRES DE LA REGULATION DE C-ABL, UNE TYROSINE KINASE A DOMAINE SH2 ET SH3

LA TYROSINE KINASE DE TYPE NON-RECEPTEUR C-ABL EST EXPRIMEE UBIQUITAIREMENT CHEZ TOUS LES METAZOAIRES. EN PLUS DE SON DOMAINE CATALYTIQUE, C-ABL POSSEDE DES DOMAINES D'INTERACTION PROTEINE-PROTEINE QUI SONT ESSENTIELS POUR SA FONCTION. SI LA FONCTION PRECISE DE C-ABL EST INCONNUE, ELLE EST IMPLIQUEE DANS UNE GRANDE VARIETE DE PROCESSUS. L'INACTIVATION DU GENE A UN PHENOTYPE PLEIOTROPIQUE CONDUISANT A UNE MORT EMBRYONNAIRE PRECOCE. L'ACTIVATION CONSTITUTIVE DE C-ABL LA TRANSFORME EN UN ONCOGENE PUISSANT IMPLIQUE DANS DES LEUCEMIES HUMAINES. IL EST DONC VITAL POUR LA CELLULE DE REGULER DE MANIERE EXTREMEMENT PRECISE SON ACTIVITE CATALYTIQUE. AU COURS DE CE TRAVAIL DE THESE, NOUS AVONS ETUDIE LES MECANISMES MOLECULAIRES REGULANT C-ABL. LA TYR 412 EST SITUEE DANS LA BOUCLE D'ACTIVATION DU DOMAINE CATALYTIQUE DE C-ABL. CE RESIDU EST AU CUR DU MECANISME DE REGULATION DE L'ACTIVITE CATALYTIQUE DE C-ABL. SON POSITIONNEMENT A L'INTERIEUR DU DOMAINE CATALYTIQUE EST ESSENTIEL AU MAINTIEN DE LA FORME INACTIVE DE C-ABL. LA PHOSPHORYLATION DE LA TYR 412 EST REQUISE POUR ACTIVER L'ENZYME. LA TYR 412 EST UN SITE DE TRANS-AUTOPHOSPHORYLATION QUI PERMET UNE BOUCLE D'AMPLIFICATION DU SIGNAL. UNE PARTICULARITE DE LA REGULATION DE C-ABL EST LE ROLE D'UN DOMAINE DE 80 ACIDES AMINES SITUE EN N-TERMINAL DE LA MOLECULE ET CONTENANT LE SITE DE MYRISTOYLATION. IL INTERAGIT DE MANIERE INTRAMOLECULAIRE AVEC LE RESTE DE LA MOLECULE. CETTE INTERACTION EST NECESSAIRE POUR LA REGULATION DE C-ABL. LE POINT COMMUN DES FORMES ONCOGENIQUES DE C-ABL EST L'ABSENCE DE CE DOMAINE. LA DELETION DE CE DOMAINE SUFFIT A RENDRE C-ABL ONCOGENIQUE. C'EST PEUT-ETRE LA QUE RESIDE LA DIFFERENCE FONDAMENTALE ENTRE LA FORME SAUVAGE ET LES FORMES ONCOGENIQUES D'ABL. L'ACTIVITE CATALYTIQUE DE C-ABL EST DONC SOUS LE CONTROLE D'UNE COMBINAISON DE FACTEURS -INTERACTIONS INTRAMOLECULAIRES, INTERACTIONS PROTEINE-PROTEINE, EVENEMENT DE PHOSPHORYLATION- QUI PERMETTENT D'ASSURER UNE REPONSE APPROPRIEE A UN STIMULUS DONNE.PARIS-BIUSJ-Thèses (751052125) / SudocCentre Technique Livre Ens. Sup. (774682301) / SudocPARIS-BIUSJ-Physique recherche (751052113) / SudocSudocFranceF

A novel Cripto-related protein reveals an essential role for EGF-CFCs in Nodal signalling in Xenopus embryos

AbstractThe location, timing and intensity of Nodal signalling are all critical for proper patterning of the vertebrate embryo. Genetic evidence from mouse and zebrafish indicates that EGF-CFC family members are essential for Nodal ligands to signal. However, the Xenopus EGF-CFC, FRL1, has been implicated in Wnt signalling and in activation of Erk MAP kinase. Here, we identify two additional Xenopus EGF-CFCs, XCR2 and XCR3. We have focused on the role of XCR1/FRL1 and XCR3, which are both expressed at gastrula stages when Nodal signalling is active. We demonstrate spatial and temporal regulation of XCR1 protein expression, whereas XCR3 appears to be expressed ubiquitously. Using gain and loss of function approaches, we show that XCR1 and XCR3 are required for Nodal-related ligands to signal during early Xenopus development. Moreover, different Nodal-related ligands require different XCRs to signal. When both XCR1 and XCR3 are knocked down, activation of the Nodal intracellular signal transducer, Smad2, is severely inhibited and neither gastrulation nor mesendoderm formation occurs. Together our results indicate that the XCRs are important for modulation of the timing and intensity of Nodal signalling in Xenopus embryos