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1.
Genes Dev ; 32(9-10): 645-657, 2018 05 01.
Artículo en Inglés | MEDLINE | ID: mdl-29748249

RESUMEN

Cholesterol is a major constituent of myelin membranes, which insulate axons and allow saltatory conduction. Therefore, Schwann cells, the myelinating glia of the peripheral nervous system, need to produce large amounts of cholesterol. Here, we define a crucial role of the transcription factor Maf in myelination and cholesterol biosynthesis and show that Maf acts downstream from Neuregulin1 (Nrg1). Maf expression is induced when Schwann cells begin myelination. Genetic ablation of Maf resulted in hypomyelination that resembled mice with defective Nrg1 signaling. Importantly, loss of Maf or Nrg1 signaling resulted in a down-regulation of the cholesterol synthesis program, and Maf directly binds to enhancers of cholesterol synthesis genes. Furthermore, we identified the molecular mechanisms by which Nrg1 signaling regulates Maf levels. Transcription of Maf depends on calmodulin-dependent kinases downstream from Nrg1, whereas Nrg1-MAPK signaling stabilizes Maf protein. Our results delineate a novel signaling cascade regulating cholesterol synthesis in myelinating Schwann cells.


Asunto(s)
Colesterol/biosíntesis , Vaina de Mielina/metabolismo , Neurregulina-1/metabolismo , Proteínas Proto-Oncogénicas c-maf/metabolismo , Células de Schwann/metabolismo , Transducción de Señal , Animales , Proteínas Quinasas Dependientes de Calcio-Calmodulina/metabolismo , Línea Celular , Colesterol/genética , Regulación de la Expresión Génica , Histona Desacetilasas/metabolismo , Ratones , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Estabilidad Proteica , Proteínas Proto-Oncogénicas c-maf/genética , Ratas , Ratas Wistar
2.
Development ; 146(17)2019 09 06.
Artículo en Inglés | MEDLINE | ID: mdl-31427287

RESUMEN

Feeding and breathing are essential motor functions and rely on the activity of hypoglossal and phrenic motor neurons that innervate the tongue and diaphragm, respectively. Little is known about the genetic programs that control the development of these neuronal subtypes. The transcription factor Tshz1 is strongly and persistently expressed in developing hypoglossal and phrenic motor neurons. We used conditional mutation of Tshz1 in the progenitor zone of motor neurons (Tshz1MNΔ) to show that Tshz1 is essential for survival and function of hypoglossal and phrenic motor neurons. Hypoglossal and phrenic motor neurons are born in correct numbers, but many die between embryonic day 13.5 and 14.5 in Tshz1MNΔ mutant mice. In addition, innervation and electrophysiological properties of phrenic and hypoglossal motor neurons are altered. Severe feeding and breathing problems accompany this developmental deficit. Although motor neuron survival can be rescued by elimination of the pro-apoptotic factor Bax, innervation, feeding and breathing defects persist in Bax-/-; Tshz1MNΔ mutants. We conclude that Tshz1 is an essential transcription factor for the development and physiological function of phrenic and hypoglossal motor neurons.


Asunto(s)
Proteínas de Homeodominio/metabolismo , Nervio Hipogloso/citología , Neuronas Motoras/fisiología , Nervio Frénico/citología , Proteínas Represoras/metabolismo , Animales , Animales Recién Nacidos , Apoptosis/genética , Supervivencia Celular/genética , Diafragma/inervación , Proteínas de Homeodominio/genética , Ratones , Ratones Transgénicos , Mutación , Pletismografía , Proteínas Represoras/genética , Respiración , Lengua/inervación , Proteína X Asociada a bcl-2/genética
3.
Genes Dev ; 28(3): 290-303, 2014 Feb 01.
Artículo en Inglés | MEDLINE | ID: mdl-24493648

RESUMEN

Myelination depends on the synthesis of large amounts of myelin transcripts and proteins and is controlled by Nrg1/ErbB/Shp2 signaling. We developed a novel pulse labeling strategy based on stable isotope labeling with amino acids in cell culture (SILAC) to measure the dynamics of myelin protein production in mice. We found that protein synthesis is dampened in the maturing postnatal peripheral nervous system, and myelination then slows down. Remarkably, sustained activation of MAPK signaling by expression of the Mek1DD allele in mice overcomes the signals that end myelination, resulting in continuous myelin growth. MAPK activation leads to minor changes in transcript levels but massively up-regulates protein production. Pharmacological interference in vivo demonstrates that the effects of activated MAPK signaling on translation are mediated by mTOR-independent mechanisms but in part also by mTOR-dependent mechanisms. Previous work demonstrated that loss of ErbB3/Shp2 signaling impairs Schwann cell development and disrupts the myelination program. We found that activated MAPK signaling strikingly compensates for the absence of ErbB3 or Shp2 during Schwann cell development and myelination.


Asunto(s)
Diferenciación Celular , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Vaina de Mielina/metabolismo , Neurregulina-1/metabolismo , Receptor ErbB-3/metabolismo , Células de Schwann/citología , Alelos , Animales , Regulación de la Expresión Génica/genética , MAP Quinasa Quinasa 1/genética , Diana Mecanicista del Complejo 1 de la Rapamicina , Ratones , Microscopía Electrónica de Transmisión , Complejos Multiproteicos , Mutación , Proteína Tirosina Fosfatasa no Receptora Tipo 11/genética , Proteína Tirosina Fosfatasa no Receptora Tipo 11/metabolismo , Receptor ErbB-3/genética , Células de Schwann/ultraestructura , Transducción de Señal , Serina-Treonina Quinasas TOR
5.
EMBO J ; 32(14): 2015-28, 2013 Jul 17.
Artículo en Inglés | MEDLINE | ID: mdl-23792428

RESUMEN

The protease ß-secretase 1 (Bace1) was identified through its critical role in production of amyloid-ß peptides (Aß), the major component of amyloid plaques in Alzheimer's disease. Bace1 is considered a promising target for the treatment of this pathology, but processes additional substrates, among them Neuregulin-1 (Nrg1). Our biochemical analysis indicates that Bace1 processes the Ig-containing ß1 Nrg1 (IgNrg1ß1) isoform. We find that a graded reduction in IgNrg1 signal strength in vivo results in increasingly severe deficits in formation and maturation of muscle spindles, a proprioceptive organ critical for muscle coordination. Further, we show that Bace1 is required for formation and maturation of the muscle spindle. Finally, pharmacological inhibition and conditional mutagenesis in adult animals demonstrate that Bace1 and Nrg1 are essential to sustain muscle spindles and to maintain motor coordination. Our results assign to Bace1 a role in the control of coordinated movement through its regulation of muscle spindle physiology, and implicate IgNrg1-dependent processing as a molecular mechanism.


Asunto(s)
Secretasas de la Proteína Precursora del Amiloide/fisiología , Ácido Aspártico Endopeptidasas/fisiología , Husos Musculares/crecimiento & desarrollo , Husos Musculares/fisiología , Neurregulina-1/fisiología , Enfermedad de Alzheimer/metabolismo , Secretasas de la Proteína Precursora del Amiloide/deficiencia , Secretasas de la Proteína Precursora del Amiloide/genética , Animales , Ácido Aspártico Endopeptidasas/deficiencia , Ácido Aspártico Endopeptidasas/genética , Humanos , Ratones , Ratones de la Cepa 129 , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Mutantes , Husos Musculares/efectos de los fármacos , Neurregulina-1/deficiencia , Neurregulina-1/genética , Neurogénesis/efectos de los fármacos , Neurogénesis/fisiología , Inhibidores de Proteasas/farmacología , Isoformas de Proteínas/genética , Isoformas de Proteínas/fisiología , Procesamiento Proteico-Postraduccional , Desempeño Psicomotor/fisiología , Pirimidinas/farmacología , Transducción de Señal , Tiazinas/farmacología
6.
Glia ; 61(9): 1542-55, 2013 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-23836548

RESUMEN

The phagocyte NADPH oxidase Nox2 generates superoxide ions implicated in the elimination of microorganisms and the redox control of inflammatory signaling. However, the role of Nox2 in phagocyte functions unrelated to immunity or pathologies is unknown. During development, oriented cell migrations insure the timely recruitment and function of phagocytes in developing tissues. Here, we have addressed the role of Nox2 in the directional migration of microglial cells during development. We show that microglial Nox2 regulates the chemotaxis of purified microglia mediated by the colony stimulating factor-1 receptor (CSF-1R) and the vascular endothelial growth factor receptor-1 (VEGFR1). Stimulation of these receptors triggers activation of Nox2 at the leading edge of polarized cells. In the early postnatal stages of mouse brain development, Nox2 is activated in macrophages / microglial cells in the lateral ventricle or the adjacent subventricular zone (SVZ). Fluorescent microglia injected into the lateral ventricle infiltrate the dorso-caudal SVZ through a mechanism that is blocked by pretreatment of the injected cells with an irreversible Nox inhibitor. Infiltration of endogenous microglia into the caudal SVZ of the cerebral cortex is prevented by (1) Nox2 gene deficiency, (2) treatment with a Nox2 inhibitor (apocynin), and (3) invalidation of the VEGFR1 kinase. We conclude that phagocytes move out of the lateral ventricle soon after birth and infiltrate the cortical SVZ through a mechanism requiring microglial Nox2 and VEGFR1 activation. Nox2 therefore modulates the migration of microglia and their development.


Asunto(s)
Quimiotaxis/fisiología , Proteínas Fluorescentes Verdes/metabolismo , Ventrículos Laterales/citología , Glicoproteínas de Membrana/metabolismo , Microglía/metabolismo , NADPH Oxidasas/metabolismo , Fagocitos/metabolismo , Receptor 1 de Factores de Crecimiento Endotelial Vascular/metabolismo , Acetofenonas/farmacología , Actinas/genética , Animales , Animales Recién Nacidos , Antígenos de Diferenciación/metabolismo , Bromodesoxiuridina , Antígeno CD11b/metabolismo , Movimiento Celular/genética , Células Cultivadas , Corteza Cerebral/anatomía & histología , Quimiotaxis/genética , Pollos , Inhibidores Enzimáticos/farmacología , Factor Estimulante de Colonias de Granulocitos y Macrófagos , Proteínas Fluorescentes Verdes/genética , Glicoproteínas de Membrana/genética , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , NADPH Oxidasa 2 , NADPH Oxidasas/genética , Proteínas Nucleares/metabolismo , Transducción de Señal , Receptor 1 de Factores de Crecimiento Endotelial Vascular/genética
7.
Proc Natl Acad Sci U S A ; 106(39): 16704-9, 2009 Sep 29.
Artículo en Inglés | MEDLINE | ID: mdl-19805360

RESUMEN

The nonreceptor tyrosine phosphatase Shp2 (PTPN11) has been implicated in tyrosine kinase, cytokine, and integrin receptor signaling. We show here that conditional mutation of Shp2 in neural crest cells and in myelinating Schwann cells resulted in deficits in glial development that are remarkably similar to those observed in mice mutant for Neuregulin-1 (Nrg1) or the Nrg1 receptors, ErbB2 and ErbB3. In cultured Shp2 mutant Schwann cells, Nrg1-evoked cellular responses like proliferation and migration were virtually abolished, and Nrg1-dependent intracellular signaling was altered. Pharmacological inhibition of Src family kinases mimicked all cellular and biochemical effects of the Shp2 mutation, implicating Src as a primary Shp2 target during Nrg1 signaling. Together, our genetic and biochemical analyses demonstrate that Shp2 is an essential component in the transduction of Nrg1/ErbB signals.


Asunto(s)
Receptores ErbB/metabolismo , Neurregulina-1/metabolismo , Proteína Tirosina Fosfatasa no Receptora Tipo 11/metabolismo , Células de Schwann/metabolismo , Transducción de Señal , Animales , Técnica del Anticuerpo Fluorescente , Ratones , Cresta Neural/metabolismo , Proteína Tirosina Fosfatasa no Receptora Tipo 11/genética , Células de Schwann/enzimología
8.
Cell Rep ; 34(2): 108623, 2021 01 12.
Artículo en Inglés | MEDLINE | ID: mdl-33440152

RESUMEN

Vesicular glutamate transporters (VGLUTs) fill synaptic vesicles with glutamate. VGLUTs were originally identified as sodium-dependent transporters of inorganic phosphate (Pi), but the physiological relevance of this activity remains unclear. Heterologous expression of all three VGLUTs greatly augments intracellular Pi levels. Using neuronal models, we show that translocation of VGLUTs to the plasma membrane during exocytosis results in highly increased Pi uptake. VGLUT-mediated Pi influx is counteracted by Pi efflux. Synaptosomes prepared from perinatal VGLUT2-/- mice that are virtually free of VGLUTs show drastically reduced cytosolic Pi levels and fail to import Pi. Glutamate partially competes with sodium (Na+)/Pi (NaPi)-uptake mediated by VGLUTs but does not appear to be transported. A nanobody that blocks glutamate transport by binding to the cytoplasmic domain of VGLUT1 abolishes Pi transport when co-expressed with VGLUT1. We conclude that VGLUTs have a dual function that is essential for both vesicular glutamate loading and Pi restoration in neurons.


Asunto(s)
Transporte Biológico/fisiología , Fosfatos/metabolismo , Vesículas Sinápticas/metabolismo , Proteínas de Transporte Vesicular de Glutamato/metabolismo , Animales , Humanos , Ratas , Transfección
9.
J Neurosci ; 28(46): 12039-51, 2008 Nov 12.
Artículo en Inglés | MEDLINE | ID: mdl-19005069

RESUMEN

Reactive oxygen species (ROS) modulate intracellular signaling but are also responsible for neuronal damage in pathological states. Microglia, the resident CNS macrophages, are prominent sources of ROS through expression of the phagocyte oxidase which catalytic subunit Nox2 generates superoxide ion (O2(.-)). Here we show that microglia also express Nox1 and other components of nonphagocyte NADPH oxidases, including p22(phox), NOXO1, NOXA1, and Rac1/2. The subcellular distribution and functions of Nox1 were determined by blocking Nox activity with diphenylene iodonium or apocynin, and by silencing the Nox1 gene in microglia purified from wild-type (WT) or Nox2-KO mice. [Nox1-p22(phox)] dimers localized in intracellular compartments are recruited to phagosome membranes during microglial phagocytosis of zymosan, and Nox1 produces O2(.-) in zymosan-loaded phagosomes. In microglia activated with lipopolysaccharide (LPS), Nox1 produces O2(.-), which enhances cell expression of inducible nitric oxide synthase and secretion of interleukin-1beta. Comparisons of microglia purified from WT, Nox2-KO, or Nox1-KO mice indicate that both Nox1 and Nox2 are required to optimize microglial production of nitric oxide. By injecting LPS in the striatum of WT and Nox1-KO mice, we show that Nox1 also enhances microglial production of cytotoxic nitrite species and promotes loss of presynaptic proteins in striatal neurons. These results demonstrate the functional expression of Nox1 in resident CNS phagocytes, which can promote production of neurotoxic compounds during neuroinflammation. Our study also shows that Nox1- and Nox2-dependent oxidases play distinct roles in microglial activation and that Nox1 is a possible target for the treatment of neuroinflammatory states.


Asunto(s)
Encefalitis/enzimología , Gliosis/enzimología , Microglía/enzimología , NADH NADPH Oxidorreductasas/metabolismo , Estrés Oxidativo/fisiología , Proteínas Adaptadoras Transductoras de Señales , Animales , Cuerpo Estriado/efectos de los fármacos , Cuerpo Estriado/enzimología , Cuerpo Estriado/fisiopatología , Grupo Citocromo b/genética , Grupo Citocromo b/metabolismo , Encefalitis/fisiopatología , Femenino , Gliosis/fisiopatología , Mediadores de Inflamación/farmacología , Lipopolisacáridos/farmacología , Masculino , Glicoproteínas de Membrana/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Microglía/efectos de los fármacos , NADH NADPH Oxidorreductasas/genética , NADPH Oxidasa 1 , NADPH Oxidasa 2 , NADPH Oxidasas/genética , NADPH Oxidasas/metabolismo , Neuropéptidos/genética , Neuropéptidos/metabolismo , Neurotoxinas/farmacología , Nitritos/metabolismo , Estrés Oxidativo/efectos de los fármacos , Proteínas/genética , Proteínas/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Zimosan/metabolismo , Proteínas de Unión al GTP rac/genética , Proteínas de Unión al GTP rac/metabolismo , Proteína de Unión al GTP rac1
10.
Cell Rep ; 23(2): 535-545, 2018 Apr 10.
Artículo en Inglés | MEDLINE | ID: mdl-29642010

RESUMEN

Vesicular glutamate transporters (VGLUTs) fill synaptic vesicles with glutamate and are thus essential for glutamatergic neurotransmission. However, VGLUTs were originally discovered as members of a transporter subfamily specific for inorganic phosphate (Pi). It is still unclear how VGLUTs accommodate glutamate transport coupled to an electrochemical proton gradient ΔµH+ with inversely directed Pi transport coupled to the Na+ gradient and the membrane potential. Using both functional reconstitution and heterologous expression, we show that VGLUT transports glutamate and Pi using a single substrate binding site but different coupling to cation gradients. When facing the cytoplasm, both ions are transported into synaptic vesicles in a ΔµH+-dependent fashion, with glutamate preferred over Pi. When facing the extracellular space, Pi is transported in a Na+-coupled manner, with glutamate competing for binding but at lower affinity. We conclude that VGLUTs have dual functions in both vesicle transmitter loading and Pi homeostasis within glutamatergic neurons.


Asunto(s)
Fosfatos/metabolismo , Proteína 1 de Transporte Vesicular de Glutamato/metabolismo , Animales , Sitios de Unión , Transporte Biológico/efectos de los fármacos , Membrana Celular/metabolismo , Exocitosis/efectos de los fármacos , Ácido Glutámico/metabolismo , Concentración de Iones de Hidrógeno , Cinética , Liposomas/química , Liposomas/metabolismo , Nigericina/farmacología , Células PC12 , Cloruro de Potasio/farmacología , Ratas , Ratas Wistar , Proteínas Recombinantes/biosíntesis , Proteínas Recombinantes/genética , Especificidad por Sustrato , Vesículas Sinápticas/metabolismo , Proteína 1 de Transporte Vesicular de Glutamato/genética
11.
J Neurosci ; 26(12): 3345-56, 2006 Mar 22.
Artículo en Inglés | MEDLINE | ID: mdl-16554485

RESUMEN

Because senile plaques in Alzheimer's disease (AD) contain reactive microglia in addition to potentially neurotoxic aggregates of amyloid-beta (Abeta), we examined the influence of microglia on the viability of rodent neurons in culture exposed to aggregated Abeta 1-40. Microglia enhanced the toxicity of Abeta by releasing glutamate through the cystine-glutamate antiporter system Xc-. This may be relevant to Abeta toxicity in AD, because the system Xc(-)-specific xCT gene is expressed not only in cultured microglia but also in reactive microglia within or surrounding amyloid plaques in transgenic mice expressing mutant human amyloid precursor protein or in wild-type mice injected with Abeta. Inhibition of NMDA receptors or system Xc- prevented the microglia-enhanced neurotoxicity of Abeta but also unmasked a neuroprotective effect of microglia mediated by microglial secretion of apolipoprotein E (apoE) in the culture medium. Immunodepletion of apoE or targeted inactivation of the apoE gene in microglia abrogated neuroprotection by microglial conditioned medium, whereas supplementation by human apoE isoforms restored protection, which was potentiated by the presence of microglia-derived cofactors. These results suggest that inhibition of microglial system Xc- might be of therapeutic value in the treatment of AD. Its inhibition not only prevents glutamate excitotoxicity but also facilitates neuroprotection by apoE.


Asunto(s)
Enfermedad de Alzheimer/metabolismo , Sistema de Transporte de Aminoácidos y+/metabolismo , Péptidos beta-Amiloides/toxicidad , Apolipoproteínas E/metabolismo , Microglía/metabolismo , Degeneración Nerviosa/metabolismo , Neuronas/metabolismo , Fragmentos de Péptidos/toxicidad , Enfermedad de Alzheimer/fisiopatología , Sistema de Transporte de Aminoácidos y+/genética , Animales , Apolipoproteínas E/genética , Comunicación Celular/fisiología , Muerte Celular/efectos de los fármacos , Muerte Celular/fisiología , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/fisiología , Células Cultivadas , Técnicas de Cocultivo , Femenino , Regulación de la Expresión Génica/efectos de los fármacos , Regulación de la Expresión Génica/fisiología , Ácido Glutámico/metabolismo , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Transgénicos , Degeneración Nerviosa/fisiopatología , Neuronas/efectos de los fármacos , Neuronas/patología , Ratas , Ratas Wistar
12.
Curr Opin Neurobiol ; 15(1): 101-7, 2005 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-15721751

RESUMEN

Cell corpses generated during CNS development are eliminated through phagocytosis performed by a variety of cells, including mesenchyme-derived macrophages and microglia, or glial cells originating in the neurogenic ectoderm. Mounting evidence indicates that in different species, phagocytes not only clear cell corpses but also engulf still-living neural cells or axons, and thereby promote cell death or axon pruning. Knowledge of the mechanisms of corpse recognition by engulfing cells provides molecular signals to this new role for phagocytes. These observations support a conserved and instructive role for phagocytosis in the execution of regressive events during neurogenesis.


Asunto(s)
Apoptosis/fisiología , Sistema Nervioso Central/crecimiento & desarrollo , Fagocitosis/fisiología , Animales , Sistema Nervioso Central/citología , Sistema Nervioso Central/fisiología , Humanos , Neuronas/citología , Neuronas/fisiología
13.
Science ; 335(6074): 1373-6, 2012 Mar 16.
Artículo en Inglés | MEDLINE | ID: mdl-22345400

RESUMEN

The sense of touch relies on detection of mechanical stimuli by specialized mechanosensory neurons. The scarcity of molecular data has made it difficult to analyze development of mechanoreceptors and to define the basis of their diversity and function. We show that the transcription factor c-Maf/c-MAF is crucial for mechanosensory function in mice and humans. The development and function of several rapidly adapting mechanoreceptor types are disrupted in c-Maf mutant mice. In particular, Pacinian corpuscles, a type of mechanoreceptor specialized to detect high-frequency vibrations, are severely atrophied. In line with this, sensitivity to high-frequency vibration is reduced in humans carrying a dominant mutation in the c-MAF gene. Thus, our work identifies a key transcription factor specifying development and function of mechanoreceptors and their end organs.


Asunto(s)
Mecanorreceptores/citología , Mecanorreceptores/fisiología , Proteínas Proto-Oncogénicas c-maf/metabolismo , Tacto , Animales , Ganglios Espinales/citología , Ganglios Espinales/embriología , Regulación del Desarrollo de la Expresión Génica , Humanos , Factores de Transcripción Maf de Gran Tamaño/genética , Factores de Transcripción Maf de Gran Tamaño/metabolismo , Ratones , Mutación , Corpúsculos de Pacini/citología , Corpúsculos de Pacini/fisiología , Proteínas Proto-Oncogénicas c-maf/genética , Proteínas Proto-Oncogénicas c-ret/genética , Proteínas Proto-Oncogénicas c-ret/metabolismo , Piel/inervación , Vibración
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