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1.
Nat Immunol ; 16(7): 766-774, 2015 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-25985233

RESUMEN

Childhood acute lymphoblastic leukemia (ALL) can often be traced to a pre-leukemic clone carrying a prenatal genetic lesion. Postnatally acquired mutations then drive clonal evolution toward overt leukemia. The enzymes RAG1-RAG2 and AID, which diversify immunoglobulin-encoding genes, are strictly segregated in developing cells during B lymphopoiesis and peripheral mature B cells, respectively. Here we identified small pre-BII cells as a natural subset with increased genetic vulnerability owing to concurrent activation of these enzymes. Consistent with epidemiological findings on childhood ALL etiology, susceptibility to genetic lesions during B lymphopoiesis at the transition from the large pre-BII cell stage to the small pre-BII cell stage was exacerbated by abnormal cytokine signaling and repetitive inflammatory stimuli. We demonstrated that AID and RAG1-RAG2 drove leukemic clonal evolution with repeated exposure to inflammatory stimuli, paralleling chronic infections in childhood.


Asunto(s)
Linfocitos B/inmunología , Evolución Clonal/inmunología , Leucemia-Linfoma Linfoblástico de Células Precursoras/inmunología , Células Precursoras de Linfocitos B/inmunología , Adolescente , Animales , Diversidad de Anticuerpos/genética , Diversidad de Anticuerpos/inmunología , Linfocitos B/metabolismo , Niño , Preescolar , Evolución Clonal/genética , Citidina Desaminasa/genética , Citidina Desaminasa/inmunología , Citidina Desaminasa/metabolismo , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/inmunología , Proteínas de Unión al ADN/metabolismo , Femenino , Citometría de Flujo , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/inmunología , Proteínas de Homeodominio/metabolismo , Humanos , Immunoblotting , Lactante , Masculino , Ratones Endogámicos NOD , Ratones Noqueados , Ratones SCID , Ratones Transgénicos , Microscopía Fluorescente , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/metabolismo , Células Precursoras de Linfocitos B/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Células Tumorales Cultivadas
2.
Biochim Biophys Acta ; 1853(9): 2104-14, 2015 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-25764978

RESUMEN

Bax inhibitor-1 (BI-1) is an evolutionarily conserved pH-dependent Ca²âº leak channel in the endoplasmic reticulum and the founding member of a family of six highly hydrophobic mammalian proteins named transmembrane BAX inhibitor motif containing (TMBIM) 1-6 with BI-1 being TMBIM6. Here we compared the structure, subcellular localization, tissue expression and the effect on the cellular Ca²âº homeostasis of all family members side by side. We found that all TMBIM proteins possess the di-aspartyl pH sensor responsible for pH sensing identified in TMBIM6 and its bacterial homologue BsYetJ. TMBIM1-3 and TMBIM4-6 represent two phylogenetically distinct groups that are localized in the Golgi apparatus (TMBIM1-3), endoplasmic reticulum (TMBIM4-6) or mitochondria (TMBIM5) but share a common structure of at least seven transmembrane domains with the last domain being semi-hydrophobic. TMBIM1 is mainly expressed in muscle, TMBIM2 and 3 in the nervous system, TMBIM4 and 5 are ubiquitously expressed and TMBIM6 in skeletal muscle, kidney, liver and spleen. All TMBIM proteins reduce the Ca²âº content of the endoplasmic reticulum, and all but TMBIM5 also reduce the cytosolic resting Ca²âº concentration. These results suggest that the TMBIM family has comparable functions in the maintenance of intracellular Ca²âº homeostasis in a wide variety of tissues. This article is part of a Special Issue entitled: 13th European Symposium on Calcium.


Asunto(s)
Calcio/metabolismo , Retículo Endoplásmico/metabolismo , Regulación de la Expresión Génica/fisiología , Aparato de Golgi/metabolismo , Homeostasis/fisiología , Proteínas de la Membrana/biosíntesis , Secuencias de Aminoácidos , Línea Celular , Retículo Endoplásmico/genética , Aparato de Golgi/genética , Humanos , Proteínas de la Membrana/genética , Especificidad de Órganos/fisiología
3.
J Biol Chem ; 288(52): 37204-15, 2013 Dec 27.
Artículo en Inglés | MEDLINE | ID: mdl-24240096

RESUMEN

Apoptosis and autophagy are fundamental homeostatic processes in eukaryotic organisms fulfilling essential roles in development and adaptation. Recently, the anti-apoptotic factor Bcl-2 has been reported to also inhibit autophagy, thus establishing a potential link between these pathways, but the mechanistic details are only beginning to emerge. Here we show that Bcl-2 directly binds to the phagophore-associated protein GABARAP. NMR experiments revealed that the interaction critically depends on a three-residue segment (EWD) of Bcl-2 adjacent to the BH4 region, which is anchored to one of the two hydrophobic pockets on the GABARAP molecule. This is at variance with the majority of GABARAP interaction partners identified previously, which occupy both hydrophobic pockets simultaneously. Bcl-2 affinity could also be detected for GEC1, but not for other mammalian Atg8 homologs. Finally, we provide evidence that overexpression of Bcl-2 inhibits lipidation of GABARAP, a key step in autophagosome formation, possibly via competition with the lipid conjugation machinery. These results support the regulatory role of Bcl-2 in autophagy and define GABARAP as a novel interaction partner involved in this intricate connection.


Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/metabolismo , Autofagia/fisiología , Proteínas del Citoesqueleto/metabolismo , Lipoilación/fisiología , Proteínas de la Membrana/metabolismo , Proteínas Asociadas a Microtúbulos/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Proteínas Adaptadoras Transductoras de Señales/química , Proteínas Adaptadoras Transductoras de Señales/genética , Secuencias de Aminoácidos , Animales , Apoptosis/fisiología , Proteínas Reguladoras de la Apoptosis , Línea Celular Transformada , Proteínas del Citoesqueleto/química , Proteínas del Citoesqueleto/genética , Humanos , Interacciones Hidrofóbicas e Hidrofílicas , Proteínas de la Membrana/química , Proteínas de la Membrana/genética , Ratones , Proteínas Asociadas a Microtúbulos/química , Proteínas Asociadas a Microtúbulos/genética , Resonancia Magnética Nuclear Biomolecular , Unión Proteica , Estructura Cuaternaria de Proteína , Estructura Terciaria de Proteína , Proteínas Proto-Oncogénicas c-bcl-2/química , Proteínas Proto-Oncogénicas c-bcl-2/genética , Ratas
4.
Hum Mol Genet ; 21(1): 150-62, 2012 Jan 01.
Artículo en Inglés | MEDLINE | ID: mdl-21965300

RESUMEN

Mutations in GDAP1 lead to recessively or dominantly inherited peripheral neuropathies (Charcot-Marie-Tooth disease, CMT), indicating that GDAP1 is essential for the viability of cells in the peripheral nervous system. GDAP1 contains domains characteristic of glutathione-S-transferases (GSTs), is located in the outer mitochondrial membrane and induces fragmentation of mitochondria. We found GDAP1 upregulated in neuronal HT22 cells selected for resistance against oxidative stress. GDAP1 over-expression protected against oxidative stress caused by depletion of the intracellular antioxidant glutathione (GHS) and against effectors of GHS depletion that affect the mitochondrial membrane integrity like truncated BH3-interacting domain death agonist and 12/15-lipoxygenase. Gdap1 knockdown, in contrast, increased the susceptibility of motor neuron-like NSC34 cells against GHS depletion. Over-expression of wild-type GDAP1, but not of GDAP1 with recessively inherited mutations that cause disease and reduce fission activity, increased the total cellular GHS content and the mitochondrial membrane potential up to a level where it apparently limits mitochondrial respiration, leading to reduced mitochondrial Ca(2+) uptake and superoxide production. Fibroblasts from autosomal-recessive CMT4A patients had reduced GDAP1 levels, reduced GHS concentration and a reduced mitochondrial membrane potential. Thus, our results suggest that the potential GST GDAP1 is implicated in the control of the cellular GHS content and mitochondrial activity, suggesting an involvement of oxidative stress in the pathogenesis of CMT4A.


Asunto(s)
Enfermedad de Charcot-Marie-Tooth/metabolismo , Glutatión/metabolismo , Potencial de la Membrana Mitocondrial , Proteínas del Tejido Nervioso/metabolismo , Línea Celular , Enfermedad de Charcot-Marie-Tooth/genética , Humanos , Mitocondrias/metabolismo , Proteínas del Tejido Nervioso/genética , Estrés Oxidativo
5.
J Biol Chem ; 287(50): 42042-52, 2012 Dec 07.
Artículo en Inglés | MEDLINE | ID: mdl-23076152

RESUMEN

Calcium ions are involved in a plethora of cellular functions including cell death and mitochondrial energy metabolism. Store-operated Ca(2+) entry over the plasma membrane is activated by depletion of intracellular Ca(2+) stores and is mediated by the sensor STIM1 and the channel ORAI1. We compared cell death susceptibility to oxidative stress in STIM1 knock-out and ORAI1 knockdown mouse embryonic fibroblasts and in knock-out cells with reconstituted wild type and dominant active STIM1. We show that STIM1 and ORAI1 deficiency renders cells more susceptible to oxidative stress, which can be rescued by STIM1 and ORAI1 overexpression. STIM1 knock-out mitochondria are tubular, have a higher Ca(2+) concentration, and are metabolically more active, resulting in constitutive oxidative stress causing increased nuclear translocation of the antioxidant transcription factor NRF2 triggered by increased phosphorylation of the translation initiation factor eIF2α and the protein kinase-like endoplasmic reticulum kinase PERK. This leads to increased transcription of antioxidant genes and a high basal glutathione in STIM1 knock-out cells, which is, however, more rapidly expended upon additional stress, resulting in increased release and nuclear translocation of apoptosis-inducing factor with subsequent cell death. Our data suggest that store-operated Ca(2+) entry and STIM1 are involved in the regulation of mitochondrial shape and bioenergetics and play a role in oxidative stress.


Asunto(s)
Embrión de Mamíferos/metabolismo , Metabolismo Energético/fisiología , Fibroblastos/metabolismo , Glicoproteínas de Membrana/metabolismo , Mitocondrias/metabolismo , Estrés Oxidativo/fisiología , Transporte Activo de Núcleo Celular/fisiología , Animales , Apoptosis/fisiología , Calcio/metabolismo , Canales de Calcio/genética , Canales de Calcio/metabolismo , Células Cultivadas , Embrión de Mamíferos/citología , Factor 2 Eucariótico de Iniciación/genética , Factor 2 Eucariótico de Iniciación/metabolismo , Fibroblastos/citología , Glicoproteínas de Membrana/genética , Ratones , Ratones Noqueados , Mitocondrias/genética , Proteína ORAI1 , Fosforilación/fisiología , Molécula de Interacción Estromal 1 , eIF-2 Quinasa/genética , eIF-2 Quinasa/metabolismo
6.
J Biol Chem ; 287(4): 2544-57, 2012 Jan 20.
Artículo en Inglés | MEDLINE | ID: mdl-22128171

RESUMEN

Bax inhibitor-1 (BI-1) is a multitransmembrane domain-spanning endoplasmic reticulum (ER)-located protein that is evolutionarily conserved and protects against apoptosis and ER stress. Furthermore, BI-1 is proposed to modulate ER Ca(2+) homeostasis by acting as a Ca(2+)-leak channel. Based on experimental determination of the BI-1 topology, we propose that its C terminus forms a Ca(2+) pore responsible for its Ca(2+)-leak properties. We utilized a set of C-terminal peptides to screen for Ca(2+) leak activity in unidirectional (45)Ca(2+)-flux experiments and identified an α-helical 20-amino acid peptide causing Ca(2+) leak from the ER. The Ca(2+) leak was independent of endogenous ER Ca(2+)-release channels or other Ca(2+)-leak mechanisms, namely translocons and presenilins. The Ca(2+)-permeating property of the peptide was confirmed in lipid-bilayer experiments. Using mutant peptides, we identified critical residues responsible for the Ca(2+)-leak properties of this BI-1 peptide, including a series of critical negatively charged aspartate residues. Using peptides corresponding to the equivalent BI-1 domain from various organisms, we found that the Ca(2+)-leak properties were conserved among animal, but not plant and yeast orthologs. By mutating one of the critical aspartate residues in the proposed Ca(2+)-channel pore in full-length BI-1, we found that Asp-213 was essential for BI-1-dependent ER Ca(2+) leak. Thus, we elucidated residues critically important for BI-1-mediated Ca(2+) leak and its potential channel pore. Remarkably, one of these residues was not conserved among plant and yeast BI-1 orthologs, indicating that the ER Ca(2+)-leak properties of BI-1 are an added function during evolution.


Asunto(s)
Canales de Calcio/metabolismo , Calcio/metabolismo , Retículo Endoplásmico/metabolismo , Membranas Intracelulares/metabolismo , Proteínas de la Membrana/metabolismo , Animales , Calcio/química , Canales de Calcio/química , Canales de Calcio/genética , Retículo Endoplásmico/química , Retículo Endoplásmico/genética , Evolución Molecular , Células HeLa , Humanos , Membranas Intracelulares/química , Membrana Dobles de Lípidos/química , Membrana Dobles de Lípidos/metabolismo , Proteínas de la Membrana/química , Proteínas de la Membrana/genética , Ratones , Mapeo Peptídico , Péptidos/química , Péptidos/genética , Péptidos/metabolismo , Plantas/química , Plantas/genética , Plantas/metabolismo , Estructura Secundaria de Proteína , Estructura Terciaria de Proteína , Levaduras/química , Levaduras/genética , Levaduras/metabolismo
7.
J Exp Med ; 204(5): 1157-66, 2007 May 14.
Artículo en Inglés | MEDLINE | ID: mdl-17485517

RESUMEN

The Philadelphia chromosome (Ph) encoding the oncogenic BCR-ABL1 kinase defines a subset of acute lymphoblastic leukemia (ALL) with a particularly unfavorable prognosis. ALL cells are derived from B cell precursors in most cases and typically carry rearranged immunoglobulin heavy chain (IGH) variable (V) region genes devoid of somatic mutations. Somatic hypermutation is restricted to mature germinal center B cells and depends on activation-induced cytidine deaminase (AID). Studying AID expression in 108 cases of ALL, we detected AID mRNA in 24 of 28 Ph(+) ALLs as compared with 6 of 80 Ph(-) ALLs. Forced expression of BCR-ABL1 in Ph(-) ALL cells and inhibition of the BCR-ABL1 kinase showed that aberrant expression of AID depends on BCR-ABL1 kinase activity. Consistent with aberrant AID expression in Ph(+) ALL, IGH V region genes and BCL6 were mutated in many Ph(+) but unmutated in most Ph(-) cases. In addition, AID introduced DNA single-strand breaks within the tumor suppressor gene CDKN2B in Ph(+) ALL cells, which was sensitive to BCR-ABL1 kinase inhibition and silencing of AID expression by RNA interference. These findings identify AID as a BCR-ABL1-induced mutator in Ph(+) ALL cells, which may be relevant with respect to the particularly unfavorable prognosis of this leukemia subset.


Asunto(s)
Linfocitos B/inmunología , Citidina Desaminasa/metabolismo , Regulación Neoplásica de la Expresión Génica/inmunología , Cromosoma Filadelfia , Leucemia-Linfoma Linfoblástico de Células Precursoras/inmunología , Proteínas Tirosina Quinasas/genética , Linfocitos B/metabolismo , Secuencia de Bases , Western Blotting , Análisis Mutacional de ADN , Proteínas de Unión al ADN/genética , Citometría de Flujo , Proteínas de Fusión bcr-abl , Genes myc/genética , Humanos , Región Variable de Inmunoglobulina/genética , Datos de Secuencia Molecular , Mutación/genética , Análisis de Secuencia por Matrices de Oligonucleótidos , Oligonucleótidos , Leucemia-Linfoma Linfoblástico de Células Precursoras/metabolismo , Proteínas Tirosina Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-bcl-6 , Interferencia de ARN , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Alineación de Secuencia
8.
J Neuroinflammation ; 9: 163, 2012 Jul 07.
Artículo en Inglés | MEDLINE | ID: mdl-22769044

RESUMEN

BACKGROUND: Neuronal degeneration in multiple sclerosis has been linked to oxidative stress. Dimethyl fumarate is a promising novel oral therapeutic option shown to reduce disease activity and progression in patients with relapsing-remitting multiple sclerosis. These effects are presumed to originate from a combination of immunomodulatory and neuroprotective mechanisms. We aimed to clarify whether neuroprotective concentrations of dimethyl fumarate have immunomodulatory effects. FINDINGS: We determined time- and concentration-dependent effects of dimethyl fumarate and its metabolite monomethyl fumarate on viability in a model of endogenous neuronal oxidative stress and clarified the mechanism of action by quantitating cellular glutathione content and recycling, nuclear translocation of transcription factors, and the expression of antioxidant genes. We compared this with changes in the cytokine profiles released by stimulated splenocytes measured by ELISPOT technology and analyzed the interactions between neuronal and immune cells and neuronal function and viability in cell death assays and multi-electrode arrays. Our observations show that dimethyl fumarate causes short-lived oxidative stress, which leads to increased levels and nuclear localization of the transcription factor nuclear factor erythroid 2-related factor 2 and a subsequent increase in glutathione synthesis and recycling in neuronal cells. Concentrations that were cytoprotective in neuronal cells had no negative effects on viability of splenocytes but suppressed the production of proinflammatory cytokines in cultures from C57BL/6 and SJL mice and had no effects on neuronal activity in multi-electrode arrays. CONCLUSIONS: These results suggest that immunomodulatory concentrations of dimethyl fumarate can reduce oxidative stress without altering neuronal network activity.


Asunto(s)
Fumaratos/farmacología , Inmunomodulación/inmunología , Fármacos Neuroprotectores/farmacología , Animales , Muerte Celular/efectos de los fármacos , Muerte Celular/inmunología , Células Cultivadas , Dimetilfumarato , Femenino , Inmunomodulación/efectos de los fármacos , Ratones , Ratones Endogámicos C57BL , Estrés Oxidativo/efectos de los fármacos , Estrés Oxidativo/inmunología , Bazo/citología , Bazo/efectos de los fármacos , Bazo/inmunología , Resultado del Tratamiento
9.
J Neurochem ; 111(2): 332-43, 2009 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-19694903

RESUMEN

In amyotrophic lateral sclerosis, down-regulation of the astrocyte-specific glutamate excitatory amino acid transporter 2 is hypothesized to increase extracellular glutamate, thereby leading to excitotoxic motor neuron death. The antibiotic ceftriaxone was recently reported to induce excitatory amino acid transporter 2 and to prolong the survival of mutant superoxide dismutase 1 transgenic mice. Here we show that ceftriaxone also protects fibroblasts and the hippocampal cell line HT22, which are not sensitive to excitotoxicity, against oxidative glutamate toxicity, where extracellular glutamate blocks cystine import via the glutamate/cystine-antiporter system x(c)(-). Lack of intracellular cystine leads to glutathione depletion and cell death because of oxidative stress. Ceftriaxone increased system x(c)(-) and glutathione levels independently of its effect on excitatory amino acid transporters by induction of the transcription factor Nrf2 (nuclear factor erythroid 2-related factor 2), a known inducer of system x(c)(-), and the specific x(c)(-) subunit xCT. No significant effect was apparent in fibroblasts deficient in Nrf2 or xCT. Similar ceftriaxone-stimulated changes in Nrf2, system x(c)(-), and glutathione were observed in rat cortical and spinal astrocytes. In addition, ceftriaxone induced xCT mRNA expression in stem cell-derived human motor neurons. We conclude that ceftriaxone-mediated neuroprotection might relate more strongly to activation of the antioxidant defense system including Nrf2 and system x(c)(-) than to excitatory amino acid transporter induction.


Asunto(s)
Sistema de Transporte de Aminoácidos y+/metabolismo , Ceftriaxona/farmacología , Neuronas Motoras/efectos de los fármacos , Factor 2 Relacionado con NF-E2/metabolismo , Fármacos Neuroprotectores/farmacología , Sistemas de Transporte de Aminoácidos Acídicos , Animales , Antibacterianos/farmacología , Astrocitos/citología , Astrocitos/efectos de los fármacos , Astrocitos/metabolismo , Línea Celular , Fibroblastos/citología , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Ácido Glutámico/toxicidad , Glutatión/metabolismo , Hipocampo/citología , Humanos , Técnicas In Vitro , Ratones , Ratones Noqueados , Neuronas Motoras/citología , Neuronas Motoras/metabolismo , Estrés Oxidativo/efectos de los fármacos , Ratas , Médula Espinal/citología , Células Madre/citología , Superóxido Dismutasa/genética , Superóxido Dismutasa/metabolismo , Superóxido Dismutasa-1
10.
Neurochem Int ; 62(5): 610-9, 2013 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-23357478

RESUMEN

Cell death in response to oxidative stress plays a role in a variety of neurodegenerative diseases and can be studied in detail in the neuronal cell line HT22, where extracellular glutamate causes glutathione depletion by inhibition of the glutamate/cystine antiporter system xc(-), elevation of reactive oxygen species and eventually programmed cell death caused by cytotoxic calcium influx. Using this paradigm, we screened 54 putative extracellular peptide or small molecule ligands for effects on cell death and identified extracellular cyclic guanosine monophosphate (cGMP) as a protective substance. Extracellular cGMP was protective, whereas the cell-permeable cGMP analog 8-pCPT-cGMP or the inhibition of cGMP degradation by phosphodiesterases was toxic. Interestingly, metabolites GMP and guanosine were even more protective than cGMP and the inhibition of the conversion of GMP to guanosine attenuated its effect, suggesting that GMP offers protection through its conversion to guanosine. Guanosine increased system xc(-) activity and cellular glutathione levels in the presence of glutamate, which can be explained by transcriptional upregulation of xCT, the functional subunit of system xc(-). However, guanosine also provided protection when added late in the cell death cascade and significantly reduced the number of calcium peaking cells, which was most likely not mediated by transcriptional mechanisms. We observed no changes in the classical protective pathways such as phosphorylation of Akt, ERK1/2 or induction of Nrf2 or ATF4. We conclude that extracellular guanosine protects against endogenous oxidative stress by two probably independent mechanisms involving system xc(-) induction and inhibition of cytotoxic calcium influx.


Asunto(s)
GMP Cíclico/fisiología , Ácido Glutámico/toxicidad , Guanosina/fisiología , Estrés Oxidativo , Animales , Calcio/metabolismo , Línea Celular , Espacio Extracelular/metabolismo , Ratones , Fosforilación , Proteínas Quinasas/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa
11.
Cell Calcium ; 50(3): 251-60, 2011 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-21663964

RESUMEN

Bax inhibitor-1 (BI-1) was initially identified for its ability to inhibit BAX-induced apoptosis in yeast cells and is the founding member of a family of highly hydrophobic proteins localized in diverse cellular membranes. It is evolutionarily conserved and orthologues from plants can substitute for mammalian BI-1 in regard to its anti-apoptotic function suggesting a high degree of functional conservation. BI-1 interacts with BCL-2 and BCL-XL and, similar to these two anti-apoptotic proteins, the effect of BI-1 on cell death involves changes in the amount of Ca(2+) releasable from intracellular stores. However, BI-1 is also a negative regulator of the endoplasmic reticulum stress sensor IRE1 α, it interacts with G-actin and increases actin polymerization, enhances cancer metastasis by altering glucose metabolism and activating the sodium-hydrogen exchanger, and reduces the production of reactive oxygen species through direct interaction with NADPH-P450 reductase. In this contribution, we summarize what is known about the expression, intracellular localization and structure of BI-1 and specifically illuminate its effects on the intracellular Ca(2+) homeostasis and how this might relate to its other functions. We also present a thorough phylogenetic analysis of BI-1 proteins from major phyla together with paralogues from all BI-1 family members.


Asunto(s)
Proteínas Reguladoras de la Apoptosis/metabolismo , Calcio/metabolismo , Proteínas de la Membrana/metabolismo , Actinas/metabolismo , Animales , Proteínas Reguladoras de la Apoptosis/genética , Muerte Celular , Endorribonucleasas/metabolismo , Homeostasis , Humanos , Mamíferos , Proteínas de la Membrana/genética , NADPH-Ferrihemoproteína Reductasa/metabolismo , Neoplasias/metabolismo , Filogenia , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Proteína bcl-X/metabolismo
12.
Exp Neurol ; 232(2): 185-94, 2011 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-21906591

RESUMEN

Store-operated Ca(2+) entry (SOCE) over the plasma membrane is activated by depletion of intracellular Ca(2+) stores and has only recently been shown to play a role in CNS processes like synaptic plasticity. However, the direct effect of SOCE on the excitability of neuronal networks in vitro and in vivo has never been determined. We confirmed the presence of SOCE and the expression of the calcium sensors STIM1 and STIM2, which convey information about the calcium load of the stores to channel proteins at the plasma membrane, in neurons and astrocytes. Inhibition of SOCE by pharmacological agents 2-APB and ML-9 reduced the steady-state neuronal Ca(2+) concentration, reduced network activity, and increased synchrony of primary neuronal cultures grown on multi-electrode arrays, which prompted us to elucidate the relative expression of STIM proteins in conditions of pathologic excitability. Both proteins were increased in brains of chronic epileptic rodents and strongly expressed in hippocampal specimens from medial temporal lobe epilepsy patients. Pharmacologic inhibition of SOCE in chronic epileptic hippocampal slices suppressed interictal spikes and rhythmized epileptic burst activity. Our results indicate that SOCE modulates the activity of neuronal networks in vitro and in vivo and delineates SOCE as a potential drug target.


Asunto(s)
Proteínas de Unión al Calcio/metabolismo , Calcio/metabolismo , Epilepsia del Lóbulo Temporal/metabolismo , Glicoproteínas de Membrana/metabolismo , Proteínas de la Membrana/metabolismo , Neuronas/metabolismo , Animales , Astrocitos/citología , Astrocitos/metabolismo , Moléculas de Adhesión Celular/metabolismo , Membrana Celular/metabolismo , Enfermedad Crónica , Corteza Entorrinal/citología , Corteza Entorrinal/fisiopatología , Epilepsia del Lóbulo Temporal/fisiopatología , Hipocampo/citología , Hipocampo/fisiopatología , Humanos , Proteínas de Neoplasias/metabolismo , Red Nerviosa/metabolismo , Red Nerviosa/fisiopatología , Neuronas/citología , Técnicas de Cultivo de Órganos , Cultivo Primario de Células , Ratas , Molécula de Interacción Estromal 1 , Molécula de Interacción Estromal 2
13.
Cancer Cell ; 16(3): 232-45, 2009 Sep 08.
Artículo en Inglés | MEDLINE | ID: mdl-19732723

RESUMEN

Chronic myeloid leukemia (CML) is induced by BCR-ABL1 and can be effectively treated for many years with Imatinib until leukemia cells acquire drug resistance through BCR-ABL1 mutations and progress into fatal B lymphoid blast crisis (LBC). Despite its clinical significance, the mechanism of progression into LBC is unknown. Here, we show that LBC but not CML cells express the B cell-specific mutator enzyme AID. We demonstrate that AID expression in CML cells promotes overall genetic instability by hypermutation of tumor suppressor and DNA repair genes. Importantly, our data uncover a causative role of AID activity in the acquisition of BCR-ABL1 mutations leading to Imatinib resistance, thus providing a rationale for the rapid development of drug resistance and blast crisis progression.


Asunto(s)
Linfocitos B/efectos de los fármacos , Crisis Blástica/tratamiento farmacológico , Citidina Desaminasa/metabolismo , Resistencia a Antineoplásicos/genética , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Mutación , Piperazinas/uso terapéutico , Pirimidinas/uso terapéutico , Animales , Linfocitos B/patología , Benzamidas , Línea Celular Tumoral , Proteínas de Fusión bcr-abl/genética , Proteínas Fluorescentes Verdes/metabolismo , Humanos , Mesilato de Imatinib , Leucemia Mielógena Crónica BCR-ABL Positiva/genética , Leucemia Mielógena Crónica BCR-ABL Positiva/patología , Luciferasas de Renilla/metabolismo , Ratones , Ratones Endogámicos BALB C , Ratones Noqueados , Ratones SCID , Ratones Transgénicos , Ensayos Antitumor por Modelo de Xenoinjerto
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