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1.
Cell ; 187(9): 2129-2142.e17, 2024 Apr 25.
Artículo en Inglés | MEDLINE | ID: mdl-38670071

RESUMEN

Interspecies blastocyst complementation (IBC) provides a unique platform to study development and holds the potential to overcome worldwide organ shortages. Despite recent successes, brain tissue has not been achieved through IBC. Here, we developed an optimized IBC strategy based on C-CRISPR, which facilitated rapid screening of candidate genes and identified that Hesx1 deficiency supported the generation of rat forebrain tissue in mice via IBC. Xenogeneic rat forebrain tissues in adult mice were structurally and functionally intact. Cross-species comparative analyses revealed that rat forebrain tissues developed at the same pace as the mouse host but maintained rat-like transcriptome profiles. The chimeric rate of rat cells gradually decreased as development progressed, suggesting xenogeneic barriers during mid-to-late pre-natal development. Interspecies forebrain complementation opens the door for studying evolutionarily conserved and divergent mechanisms underlying brain development and cognitive function. The C-CRISPR-based IBC strategy holds great potential to broaden the study and application of interspecies organogenesis.


Asunto(s)
Prosencéfalo , Animales , Prosencéfalo/metabolismo , Prosencéfalo/embriología , Ratones , Ratas , Blastocisto/metabolismo , Femenino , Sistemas CRISPR-Cas/genética , Transcriptoma , Organogénesis , Repeticiones Palindrómicas Cortas Agrupadas y Regularmente Espaciadas/genética , Masculino , Ratones Endogámicos C57BL
2.
Cell ; 187(9): 2143-2157.e15, 2024 Apr 25.
Artículo en Inglés | MEDLINE | ID: mdl-38670072

RESUMEN

A central question for regenerative neuroscience is whether synthetic neural circuits, such as those built from two species, can function in an intact brain. Here, we apply blastocyst complementation to selectively build and test interspecies neural circuits. Despite approximately 10-20 million years of evolution, and prominent species differences in brain size, rat pluripotent stem cells injected into mouse blastocysts develop and persist throughout the mouse brain. Unexpectedly, the mouse niche reprograms the birth dates of rat neurons in the cortex and hippocampus, supporting rat-mouse synaptic activity. When mouse olfactory neurons are genetically silenced or killed, rat neurons restore information flow to odor processing circuits. Moreover, they rescue the primal behavior of food seeking, although less well than mouse neurons. By revealing that a mouse can sense the world using neurons from another species, we establish neural blastocyst complementation as a powerful tool to identify conserved mechanisms of brain development, plasticity, and repair.


Asunto(s)
Neuronas , Animales , Ratones , Ratas , Neuronas/metabolismo , Neuronas/citología , Neuronas/fisiología , Blastocisto/metabolismo , Blastocisto/citología , Células Madre Pluripotentes/citología , Células Madre Pluripotentes/metabolismo , Encéfalo/citología , Encéfalo/fisiología , Femenino , Hipocampo/citología , Hipocampo/fisiología , Especificidad de la Especie , Ratones Endogámicos C57BL , Masculino
3.
Cell ; 187(9): 2175-2193.e21, 2024 Apr 25.
Artículo en Inglés | MEDLINE | ID: mdl-38552623

RESUMEN

In addition to long-distance molecular motor-mediated transport, cellular vesicles also need to be moved at short distances with defined directions to meet functional needs in subcellular compartments but with unknown mechanisms. Such short-distance vesicle transport does not involve molecular motors. Here, we demonstrate, using synaptic vesicle (SV) transport as a paradigm, that phase separation of synaptic proteins with vesicles can facilitate regulated, directional vesicle transport between different presynaptic bouton sub-compartments. Specifically, a large coiled-coil scaffold protein Piccolo, in response to Ca2+ and via its C2A domain-mediated Ca2+ sensing, can extract SVs from the synapsin-clustered reserve pool condensate and deposit the extracted SVs onto the surface of the active zone protein condensate. We further show that the Trk-fused gene, TFG, also participates in COPII vesicle trafficking from ER to the ER-Golgi intermediate compartment via phase separation. Thus, phase separation may play a general role in short-distance, directional vesicle transport in cells.


Asunto(s)
Vesículas Cubiertas por Proteínas de Revestimiento , Retículo Endoplásmico , Vesículas Sinápticas , Animales , Vesículas Sinápticas/metabolismo , Vesículas Cubiertas por Proteínas de Revestimiento/metabolismo , Retículo Endoplásmico/metabolismo , Calcio/metabolismo , Aparato de Golgi/metabolismo , Ratas , Transporte Biológico , Terminales Presinápticos/metabolismo , Sinapsinas/metabolismo , Condensados Biomoleculares/metabolismo , Proteínas del Citoesqueleto/metabolismo , Separación de Fases
4.
Cell ; 186(25): 5517-5535.e24, 2023 12 07.
Artículo en Inglés | MEDLINE | ID: mdl-37992713

RESUMEN

Transfer RNA (tRNA) modifications are critical for protein synthesis. Queuosine (Q), a 7-deaza-guanosine derivative, is present in tRNA anticodons. In vertebrate tRNAs for Tyr and Asp, Q is further glycosylated with galactose and mannose to generate galQ and manQ, respectively. However, biogenesis and physiological relevance of Q-glycosylation remain poorly understood. Here, we biochemically identified two RNA glycosylases, QTGAL and QTMAN, and successfully reconstituted Q-glycosylation of tRNAs using nucleotide diphosphate sugars. Ribosome profiling of knockout cells revealed that Q-glycosylation slowed down elongation at cognate codons, UAC and GAC (GAU), respectively. We also found that galactosylation of Q suppresses stop codon readthrough. Moreover, protein aggregates increased in cells lacking Q-glycosylation, indicating that Q-glycosylation contributes to proteostasis. Cryo-EM of human ribosome-tRNA complex revealed the molecular basis of codon recognition regulated by Q-glycosylations. Furthermore, zebrafish qtgal and qtman knockout lines displayed shortened body length, implying that Q-glycosylation is required for post-embryonic growth in vertebrates.


Asunto(s)
ARN de Transferencia , Animales , Humanos , Ratas , Anticodón , Línea Celular , Codón , Glicosilación , Nucleósido Q/química , Nucleósido Q/genética , Nucleósido Q/metabolismo , ARN de Transferencia/química , ARN de Transferencia/metabolismo , Porcinos , Pez Cebra/metabolismo , Conformación de Ácido Nucleico
5.
Cell ; 186(25): 5500-5516.e21, 2023 12 07.
Artículo en Inglés | MEDLINE | ID: mdl-38016470

RESUMEN

Most animals require sleep, and sleep loss induces serious pathophysiological consequences, including death. Previous experimental approaches for investigating sleep impacts in mice have been unable to persistently deprive animals of both rapid eye movement sleep (REMS) and non-rapid eye movement sleep (NREMS). Here, we report a "curling prevention by water" paradigm wherein mice remain awake 96% of the time. After 4 days of exposure, mice exhibit severe inflammation, and approximately 80% die. Sleep deprivation increases levels of prostaglandin D2 (PGD2) in the brain, and we found that elevated PGD2 efflux across the blood-brain-barrier-mediated by ATP-binding cassette subfamily C4 transporter-induces both accumulation of circulating neutrophils and a cytokine-storm-like syndrome. Experimental disruption of the PGD2/DP1 axis dramatically reduced sleep-deprivation-induced inflammation. Thus, our study reveals that sleep-related changes in PGD2 in the central nervous system drive profound pathological consequences in the peripheral immune system.


Asunto(s)
Privación de Sueño , Animales , Ratones , Citocinas/metabolismo , Inflamación , Prostaglandina D2 , Sueño/fisiología , Privación de Sueño/genética , Privación de Sueño/metabolismo , Síndrome , Humanos , Ratas , Línea Celular , Tormentas Ciclónicas , Neutrófilos/metabolismo
6.
Cell ; 184(2): 370-383.e13, 2021 01 21.
Artículo en Inglés | MEDLINE | ID: mdl-33333023

RESUMEN

Proton-coupled monocarboxylate transporters MCT1-4 catalyze the transmembrane movement of metabolically essential monocarboxylates and have been targeted for cancer treatment because of their enhanced expression in various tumors. Here, we report five cryo-EM structures, at resolutions of 3.0-3.3 Å, of human MCT1 bound to lactate or inhibitors in the presence of Basigin-2, a single transmembrane segment (TM)-containing chaperon. MCT1 exhibits similar outward-open conformations when complexed with lactate or the inhibitors BAY-8002 and AZD3965. In the presence of the inhibitor 7ACC2 or with the neutralization of the proton-coupling residue Asp309 by Asn, similar inward-open structures were captured. Complemented by structural-guided biochemical analyses, our studies reveal the substrate binding and transport mechanism of MCTs, elucidate the mode of action of three anti-cancer drug candidates, and identify the determinants for subtype-specific sensitivities to AZD3965 by MCT1 and MCT4. These findings lay out an important framework for structure-guided drug discovery targeting MCTs.


Asunto(s)
Antineoplásicos/farmacología , Transportadores de Ácidos Monocarboxílicos/antagonistas & inhibidores , Transportadores de Ácidos Monocarboxílicos/química , Simportadores/antagonistas & inhibidores , Simportadores/química , Secuencia de Aminoácidos , Animales , Basigina/química , Sitios de Unión , Microscopía por Crioelectrón , Humanos , Ligandos , Modelos Moleculares , Transportadores de Ácidos Monocarboxílicos/ultraestructura , Proteínas Mutantes/química , Proteínas Mutantes/metabolismo , Protones , Pirimidinonas/química , Pirimidinonas/farmacología , Ratas , Homología Estructural de Proteína , Especificidad por Sustrato , Simportadores/ultraestructura , Tiofenos/química , Tiofenos/farmacología
7.
Cell ; 184(20): 5151-5162.e11, 2021 09 30.
Artículo en Inglés | MEDLINE | ID: mdl-34520724

RESUMEN

The heartbeat is initiated by voltage-gated sodium channel NaV1.5, which opens rapidly and triggers the cardiac action potential; however, the structural basis for pore opening remains unknown. Here, we blocked fast inactivation with a mutation and captured the elusive open-state structure. The fast inactivation gate moves away from its receptor, allowing asymmetric opening of pore-lining S6 segments, which bend and rotate at their intracellular ends to dilate the activation gate to ∼10 Å diameter. Molecular dynamics analyses predict physiological rates of Na+ conductance. The open-state pore blocker propafenone binds in a high-affinity pose, and drug-access pathways are revealed through the open activation gate and fenestrations. Comparison with mutagenesis results provides a structural map of arrhythmia mutations that target the activation and fast inactivation gates. These results give atomic-level insights into molecular events that underlie generation of the action potential, open-state drug block, and fast inactivation of cardiac sodium channels, which initiate the heartbeat.


Asunto(s)
Canal de Sodio Activado por Voltaje NAV1.5/química , Canal de Sodio Activado por Voltaje NAV1.5/metabolismo , Animales , Arritmias Cardíacas/genética , Microscopía por Crioelectrón , Células HEK293 , Frecuencia Cardíaca/efectos de los fármacos , Humanos , Activación del Canal Iónico , Modelos Moleculares , Simulación de Dinámica Molecular , Mutación/genética , Miocardio , Canal de Sodio Activado por Voltaje NAV1.5/aislamiento & purificación , Canal de Sodio Activado por Voltaje NAV1.5/ultraestructura , Propafenona/farmacología , Conformación Proteica , Ratas , Sodio/metabolismo , Factores de Tiempo , Agua/química
8.
Cell ; 184(5): 1299-1313.e19, 2021 03 04.
Artículo en Inglés | MEDLINE | ID: mdl-33606976

RESUMEN

It is unclear how binding of antidepressant drugs to their targets gives rise to the clinical antidepressant effect. We discovered that the transmembrane domain of tyrosine kinase receptor 2 (TRKB), the brain-derived neurotrophic factor (BDNF) receptor that promotes neuronal plasticity and antidepressant responses, has a cholesterol-sensing function that mediates synaptic effects of cholesterol. We then found that both typical and fast-acting antidepressants directly bind to TRKB, thereby facilitating synaptic localization of TRKB and its activation by BDNF. Extensive computational approaches including atomistic molecular dynamics simulations revealed a binding site at the transmembrane region of TRKB dimers. Mutation of the TRKB antidepressant-binding motif impaired cellular, behavioral, and plasticity-promoting responses to antidepressants in vitro and in vivo. We suggest that binding to TRKB and allosteric facilitation of BDNF signaling is the common mechanism for antidepressant action, which may explain why typical antidepressants act slowly and how molecular effects of antidepressants are translated into clinical mood recovery.


Asunto(s)
Antidepresivos/farmacología , Receptor trkB/metabolismo , Animales , Antidepresivos/química , Antidepresivos/metabolismo , Sitios de Unión , Factor Neurotrófico Derivado del Encéfalo/metabolismo , Línea Celular , Colesterol/metabolismo , Embrión de Mamíferos , Fluoxetina/química , Fluoxetina/metabolismo , Fluoxetina/farmacología , Hipocampo/metabolismo , Humanos , Ratones , Modelos Animales , Simulación de Dinámica Molecular , Dominios Proteicos , Ratas , Receptor trkB/química , Corteza Visual/metabolismo
9.
Cell ; 184(24): 5886-5901.e22, 2021 11 24.
Artículo en Inglés | MEDLINE | ID: mdl-34822784

RESUMEN

Current therapies for Alzheimer's disease seek to correct for defective cholinergic transmission by preventing the breakdown of acetylcholine through inhibition of acetylcholinesterase, these however have limited clinical efficacy. An alternative approach is to directly activate cholinergic receptors responsible for learning and memory. The M1-muscarinic acetylcholine (M1) receptor is the target of choice but has been hampered by adverse effects. Here we aimed to design the drug properties needed for a well-tolerated M1-agonist with the potential to alleviate cognitive loss by taking a stepwise translational approach from atomic structure, cell/tissue-based assays, evaluation in preclinical species, clinical safety testing, and finally establishing activity in memory centers in humans. Through this approach, we rationally designed the optimal properties, including selectivity and partial agonism, into HTL9936-a potential candidate for the treatment of memory loss in Alzheimer's disease. More broadly, this demonstrates a strategy for targeting difficult GPCR targets from structure to clinic.


Asunto(s)
Enfermedad de Alzheimer/tratamiento farmacológico , Diseño de Fármacos , Receptor Muscarínico M1/agonistas , Anciano , Anciano de 80 o más Años , Envejecimiento/patología , Enfermedad de Alzheimer/complicaciones , Enfermedad de Alzheimer/diagnóstico por imagen , Enfermedad de Alzheimer/patología , Secuencia de Aminoácidos , Animales , Presión Sanguínea/efectos de los fármacos , Células CHO , Inhibidores de la Colinesterasa/farmacología , Cricetulus , Cristalización , Modelos Animales de Enfermedad , Perros , Donepezilo/farmacología , Electroencefalografía , Femenino , Células HEK293 , Frecuencia Cardíaca/efectos de los fármacos , Humanos , Masculino , Ratones Endogámicos C57BL , Modelos Moleculares , Simulación de Dinámica Molecular , Degeneración Nerviosa/complicaciones , Degeneración Nerviosa/patología , Primates , Ratas , Receptor Muscarínico M1/química , Transducción de Señal , Homología Estructural de Proteína
10.
Cell ; 183(7): 2003-2019.e16, 2020 12 23.
Artículo en Inglés | MEDLINE | ID: mdl-33308478

RESUMEN

The ability to record transient cellular events in the DNA or RNA of cells would enable precise, large-scale analysis, selection, and reprogramming of heterogeneous cell populations. Here, we report a molecular technology for stable genetic tagging of cells that exhibit activity-related increases in intracellular calcium concentration (FLiCRE). We used FLiCRE to transcriptionally label activated neural ensembles in the nucleus accumbens of the mouse brain during brief stimulation of aversive inputs. Using single-cell RNA sequencing, we detected FLiCRE transcripts among the endogenous transcriptome, providing simultaneous readout of both cell-type and calcium activation history. We identified a cell type in the nucleus accumbens activated downstream of long-range excitatory projections. Taking advantage of FLiCRE's modular design, we expressed an optogenetic channel selectively in this cell type and showed that direct recruitment of this otherwise genetically inaccessible population elicits behavioral aversion. The specificity and minute resolution of FLiCRE enables molecularly informed characterization, manipulation, and reprogramming of activated cellular ensembles.


Asunto(s)
Conducta Animal , Calcio/metabolismo , Cuerpo Estriado/metabolismo , Animales , Femenino , Células HEK293 , Humanos , Cinética , Masculino , Ratones Endogámicos C57BL , Neuronas/metabolismo , Optogenética , Ratas , Análisis de la Célula Individual , Transcriptoma/genética
11.
Cell ; 180(3): 552-567.e25, 2020 02 06.
Artículo en Inglés | MEDLINE | ID: mdl-32004462

RESUMEN

Cognitive faculties such as imagination, planning, and decision-making entail the ability to represent hypothetical experience. Crucially, animal behavior in natural settings implies that the brain can represent hypothetical future experience not only quickly but also constantly over time, as external events continually unfold. To determine how this is possible, we recorded neural activity in the hippocampus of rats navigating a maze with multiple spatial paths. We found neural activity encoding two possible future scenarios (two upcoming maze paths) in constant alternation at 8 Hz: one scenario per ∼125-ms cycle. Further, we found that the underlying dynamics of cycling (both inter- and intra-cycle dynamics) generalized across qualitatively different representational correlates (location and direction). Notably, cycling occurred across moving behaviors, including during running. These findings identify a general dynamic process capable of quickly and continually representing hypothetical experience, including that of multiple possible futures.


Asunto(s)
Conducta Animal/fisiología , Cognición/fisiología , Toma de Decisiones/fisiología , Hipocampo/fisiología , Potenciales de Acción/fisiología , Animales , Locomoción/fisiología , Masculino , Aprendizaje por Laberinto/fisiología , Red Nerviosa/fisiología , Neuronas/fisiología , Ratas , Ratas Long-Evans , Ritmo Teta/fisiología
12.
Cell ; 180(5): 984-1001.e22, 2020 03 05.
Artículo en Inglés | MEDLINE | ID: mdl-32109414

RESUMEN

Aging causes a functional decline in tissues throughout the body that may be delayed by caloric restriction (CR). However, the cellular profiles and signatures of aging, as well as those ameliorated by CR, remain unclear. Here, we built comprehensive single-cell and single-nucleus transcriptomic atlases across various rat tissues undergoing aging and CR. CR attenuated aging-related changes in cell type composition, gene expression, and core transcriptional regulatory networks. Immune cells were increased during aging, and CR favorably reversed the aging-disturbed immune ecosystem. Computational prediction revealed that the abnormal cell-cell communication patterns observed during aging, including the excessive proinflammatory ligand-receptor interplay, were reversed by CR. Our work provides multi-tissue single-cell transcriptional landscapes associated with aging and CR in a mammal, enhances our understanding of the robustness of CR as a geroprotective intervention, and uncovers how metabolic intervention can act upon the immune system to modify the process of aging.


Asunto(s)
Envejecimiento/genética , Restricción Calórica , Sistema Inmunológico/metabolismo , Transcriptoma/genética , Envejecimiento/metabolismo , Envejecimiento/patología , Animales , Reprogramación Celular/genética , Regulación de la Expresión Génica/genética , Redes Reguladoras de Genes/genética , Humanos , Ratas , Análisis de la Célula Individual
13.
Cell ; 180(2): 221-232, 2020 01 23.
Artículo en Inglés | MEDLINE | ID: mdl-31978342

RESUMEN

Human diseases are increasingly linked with an altered or "dysbiotic" gut microbiota, but whether such changes are causal, consequential, or bystanders to disease is, for the most part, unresolved. Human microbiota-associated (HMA) rodents have become a cornerstone of microbiome science for addressing causal relationships between altered microbiomes and host pathology. In a systematic review, we found that 95% of published studies (36/38) on HMA rodents reported a transfer of pathological phenotypes to recipient animals, and many extrapolated the findings to make causal inferences to human diseases. We posit that this exceedingly high rate of inter-species transferable pathologies is implausible and overstates the role of the gut microbiome in human disease. We advocate for a more rigorous and critical approach for inferring causality to avoid false concepts and prevent unrealistic expectations that may undermine the credibility of microbiome science and delay its translation.


Asunto(s)
Disbiosis/microbiología , Microbioma Gastrointestinal/fisiología , Roedores/microbiología , Animales , Enfermedad/etiología , Trasplante de Microbiota Fecal/métodos , Humanos , Ratones , Microbiota/fisiología , Modelos Animales , Ratas
14.
Cell ; 180(2): 311-322.e15, 2020 01 23.
Artículo en Inglés | MEDLINE | ID: mdl-31883793

RESUMEN

The propagation of electrical impulses along axons is highly accelerated by the myelin sheath and produces saltating or "jumping" action potentials across internodes, from one node of Ranvier to the next. The underlying electrical circuit, as well as the existence and role of submyelin conduction in saltatory conduction remain, however, elusive. Here, we made patch-clamp and high-speed voltage-calibrated optical recordings of potentials across the nodal and internodal axolemma of myelinated neocortical pyramidal axons combined with electron microscopy and experimentally constrained cable modeling. Our results reveal a nanoscale yet conductive periaxonal space, incompletely sealed at the paranodes, which separates the potentials across the low-capacitance myelin sheath and internodal axolemma. The emerging double-cable model reproduces the recorded evolution of voltage waveforms across nodes and internodes, including rapid nodal potentials traveling in advance of attenuated waves in the internodal axolemma, revealing a mechanism for saltation across time and space.


Asunto(s)
Potenciales de Acción/fisiología , Vaina de Mielina/fisiología , Fibras Nerviosas Mielínicas/fisiología , Nódulos de Ranvier/fisiología , Animales , Axones/metabolismo , Axones/fisiología , Masculino , Modelos Neurológicos , Fibras Nerviosas Mielínicas/metabolismo , Técnicas de Placa-Clamp/métodos , Células Piramidales/fisiología , Ratas , Ratas Wistar
15.
Cell ; 182(5): 1125-1139.e18, 2020 09 03.
Artículo en Inglés | MEDLINE | ID: mdl-32822574

RESUMEN

Maternal decidual NK (dNK) cells promote placentation, but how they protect against placental infection while maintaining fetal tolerance is unclear. Here we show that human dNK cells highly express the antimicrobial peptide granulysin (GNLY) and selectively transfer it via nanotubes to extravillous trophoblasts to kill intracellular Listeria monocytogenes (Lm) without killing the trophoblast. Transfer of GNLY, but not other cell death-inducing cytotoxic granule proteins, strongly inhibits Lm in human placental cultures and in mouse and human trophoblast cell lines. Placental and fetal Lm loads are lower and pregnancy success is greatly improved in pregnant Lm-infected GNLY-transgenic mice than in wild-type mice that lack GNLY. This immune defense is not restricted to pregnancy; peripheral NK (pNK) cells also transfer GNLY to kill bacteria in macrophages and dendritic cells without killing the host cell. Nanotube transfer of GNLY allows dNK to protect against infection while leaving the maternal-fetal barrier intact.


Asunto(s)
Antígenos de Diferenciación de Linfocitos T/inmunología , Bacterias/inmunología , Movimiento Celular/inmunología , Células Asesinas Naturales/inmunología , Trofoblastos/inmunología , Animales , Línea Celular , Línea Celular Tumoral , Células Dendríticas/inmunología , Femenino , Células HeLa , Humanos , Macrófagos/inmunología , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Transgénicos , Placenta/inmunología , Placenta/microbiología , Embarazo , Ratas , Células THP-1 , Trofoblastos/microbiología
16.
Cell ; 182(4): 960-975.e15, 2020 08 20.
Artículo en Inglés | MEDLINE | ID: mdl-32763155

RESUMEN

Parental behavior is pervasive throughout the animal kingdom and essential for species survival. However, the relative contribution of the father to offspring care differs markedly across animals, even between related species. The mechanisms that organize and control paternal behavior remain poorly understood. Using Sprague-Dawley rats and C57BL/6 mice, two species at opposite ends of the paternal spectrum, we identified that distinct electrical oscillation patterns in neuroendocrine dopamine neurons link to a chain of low dopamine release, high circulating prolactin, prolactin receptor-dependent activation of medial preoptic area galanin neurons, and paternal care behavior in male mice. In rats, the same parameters exhibit inverse profiles. Optogenetic manipulation of these rhythms in mice dramatically shifted serum prolactin and paternal behavior, whereas injecting prolactin into non-paternal rat sires triggered expression of parental care. These findings identify a frequency-tuned brain-endocrine-brain circuit that can act as a gain control system determining a species' parental strategy.


Asunto(s)
Dopamina/metabolismo , Hipotálamo/fisiología , Neuronas/fisiología , Conducta Paterna/fisiología , Animales , Encéfalo/fisiología , Masculino , Ratones , Ratones Endogámicos C57BL , Neuronas/metabolismo , Optogenética , Técnicas de Placa-Clamp , Prolactina/sangre , Ratas , Ratas Sprague-Dawley , Receptores de Prolactina/deficiencia , Receptores de Prolactina/genética , Receptores de Prolactina/metabolismo
17.
Cell ; 180(1): 122-134.e10, 2020 01 09.
Artículo en Inglés | MEDLINE | ID: mdl-31866066

RESUMEN

Voltage-gated sodium channel Nav1.5 generates cardiac action potentials and initiates the heartbeat. Here, we report structures of NaV1.5 at 3.2-3.5 Å resolution. NaV1.5 is distinguished from other sodium channels by a unique glycosyl moiety and loss of disulfide-bonding capability at the NaVß subunit-interaction sites. The antiarrhythmic drug flecainide specifically targets the central cavity of the pore. The voltage sensors are partially activated, and the fast-inactivation gate is partially closed. Activation of the voltage sensor of Domain III allows binding of the isoleucine-phenylalanine-methionine (IFM) motif to the inactivation-gate receptor. Asp and Ala, in the selectivity motif DEKA, line the walls of the ion-selectivity filter, whereas Glu and Lys are in positions to accept and release Na+ ions via a charge-delocalization network. Arrhythmia mutation sites undergo large translocations during gating, providing a potential mechanism for pathogenic effects. Our results provide detailed insights into Nav1.5 structure, pharmacology, activation, inactivation, ion selectivity, and arrhythmias.


Asunto(s)
Canal de Sodio Activado por Voltaje NAV1.5/genética , Canal de Sodio Activado por Voltaje NAV1.5/metabolismo , Canal de Sodio Activado por Voltaje NAV1.5/ultraestructura , Animales , Línea Celular , Células HEK293 , Corazón/fisiología , Humanos , Activación del Canal Iónico/fisiología , Potenciales de la Membrana/fisiología , Técnicas de Placa-Clamp/métodos , Ratas , Sodio/metabolismo , Canales de Sodio/química , Relación Estructura-Actividad , Canales de Sodio Activados por Voltaje/metabolismo , Canales de Sodio Activados por Voltaje/ultraestructura
18.
Cell ; 180(1): 50-63.e12, 2020 01 09.
Artículo en Inglés | MEDLINE | ID: mdl-31923399

RESUMEN

Mucosal barrier immunity is essential for the maintenance of the commensal microflora and combating invasive bacterial infection. Although immune and epithelial cells are thought to be the canonical orchestrators of this complex equilibrium, here, we show that the enteric nervous system (ENS) plays an essential and non-redundant role in governing the antimicrobial protein (AMP) response. Using confocal microscopy and single-molecule fluorescence in situ mRNA hybridization (smFISH) studies, we observed that intestinal neurons produce the pleiotropic cytokine IL-18. Strikingly, deletion of IL-18 from the enteric neurons alone, but not immune or epithelial cells, rendered mice susceptible to invasive Salmonella typhimurium (S.t.) infection. Mechanistically, unbiased RNA sequencing and single-cell sequencing revealed that enteric neuronal IL-18 is specifically required for homeostatic goblet cell AMP production. Together, we show that neuron-derived IL-18 signaling controls tissue-wide intestinal immunity and has profound consequences on the mucosal barrier and invasive bacterial killing.


Asunto(s)
Inmunidad Mucosa/inmunología , Interleucina-18/inmunología , Mucosa Intestinal/inmunología , Animales , Citocinas/inmunología , Sistema Nervioso Entérico/inmunología , Sistema Nervioso Entérico/metabolismo , Células Epiteliales/inmunología , Femenino , Células Caliciformes/inmunología , Interleucina-18/biosíntesis , Mucosa Intestinal/metabolismo , Intestino Delgado/inmunología , Masculino , Ratones , Ratones Endogámicos C57BL , Neuronas/inmunología , Ratas , Ratas Sprague-Dawley , Infecciones por Salmonella/inmunología , Salmonella typhimurium/inmunología , Transducción de Señal/inmunología
19.
Cell ; 183(2): 335-346.e13, 2020 10 15.
Artículo en Inglés | MEDLINE | ID: mdl-33035452

RESUMEN

Muscle spasticity after nervous system injuries and painful low back spasm affect more than 10% of global population. Current medications are of limited efficacy and cause neurological and cardiovascular side effects because they target upstream regulators of muscle contraction. Direct myosin inhibition could provide optimal muscle relaxation; however, targeting skeletal myosin is particularly challenging because of its similarity to the cardiac isoform. We identified a key residue difference between these myosin isoforms, located in the communication center of the functional regions, which allowed us to design a selective inhibitor, MPH-220. Mutagenic analysis and the atomic structure of MPH-220-bound skeletal muscle myosin confirmed the mechanism of specificity. Targeting skeletal muscle myosin by MPH-220 enabled muscle relaxation, in human and model systems, without cardiovascular side effects and improved spastic gait disorders after brain injury in a disease model. MPH-220 provides a potential nervous-system-independent option to treat spasticity and muscle stiffness.


Asunto(s)
Músculo Esquelético/metabolismo , Miosinas del Músculo Esquelético/efectos de los fármacos , Miosinas del Músculo Esquelético/genética , Adulto , Animales , Miosinas Cardíacas/genética , Miosinas Cardíacas/metabolismo , Línea Celular , Sistemas de Liberación de Medicamentos , Femenino , Humanos , Masculino , Ratones , Contracción Muscular/fisiología , Fibras Musculares Esqueléticas/fisiología , Espasticidad Muscular/genética , Espasticidad Muscular/fisiopatología , Músculo Esquelético/fisiología , Miosinas/efectos de los fármacos , Miosinas/genética , Miosinas/metabolismo , Isoformas de Proteínas , Ratas , Ratas Wistar , Miosinas del Músculo Esquelético/metabolismo
20.
Cell ; 182(5): 1252-1270.e34, 2020 09 03.
Artículo en Inglés | MEDLINE | ID: mdl-32818467

RESUMEN

Aryl hydrocarbon receptor (AHR) activation by tryptophan (Trp) catabolites enhances tumor malignancy and suppresses anti-tumor immunity. The context specificity of AHR target genes has so far impeded systematic investigation of AHR activity and its upstream enzymes across human cancers. A pan-tissue AHR signature, derived by natural language processing, revealed that across 32 tumor entities, interleukin-4-induced-1 (IL4I1) associates more frequently with AHR activity than IDO1 or TDO2, hitherto recognized as the main Trp-catabolic enzymes. IL4I1 activates the AHR through the generation of indole metabolites and kynurenic acid. It associates with reduced survival in glioma patients, promotes cancer cell motility, and suppresses adaptive immunity, thereby enhancing the progression of chronic lymphocytic leukemia (CLL) in mice. Immune checkpoint blockade (ICB) induces IDO1 and IL4I1. As IDO1 inhibitors do not block IL4I1, IL4I1 may explain the failure of clinical studies combining ICB with IDO1 inhibition. Taken together, IL4I1 blockade opens new avenues for cancer therapy.


Asunto(s)
L-Aminoácido Oxidasa/metabolismo , Receptores de Hidrocarburo de Aril/metabolismo , Adulto , Anciano , Animales , Línea Celular , Línea Celular Tumoral , Progresión de la Enfermedad , Femenino , Glioma/inmunología , Glioma/metabolismo , Glioma/terapia , Células HEK293 , Humanos , Inhibidores de Puntos de Control Inmunológico/farmacología , Indolamina-Pirrol 2,3,-Dioxigenasa/metabolismo , Leucemia Linfocítica Crónica de Células B/inmunología , Leucemia Linfocítica Crónica de Células B/metabolismo , Leucemia Linfocítica Crónica de Células B/terapia , Masculino , Ratones , Ratones Endogámicos C57BL , Persona de Mediana Edad , Ratas
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