RESUMEN
Precise regulation of Notch signaling is essential for normal vertebrate development. Mind bomb (Mib) is a ubiquitin ligase that is required for activation of Notch by Notch׳s ligand, Delta. Sorting Nexin 5 (SNX5) co-localizes with Mib and Delta complexes and has been shown to directly bind to Mib. We show that microRNA-216a (miR-216a) is expressed in the retina during early development and regulates snx5 to precisely regulate Notch signaling. miR-216a and snx5 have complementary expression patterns. Knocking down miR-216a and/or overexpression of snx5 resulted in increased Notch activation. Conversely, knocking down snx5 and/or miR-216a overexpression caused a decrease in Notch activation. We propose a model in which SNX5, precisely controlled by miR-216a, is a vital partner of Mib in promoting endocytosis of Delta and subsequent activation of Notch signaling.
Asunto(s)
Regulación del Desarrollo de la Expresión Génica/fisiología , MicroARNs/metabolismo , Retina/embriología , Transducción de Señal/fisiología , Nexinas de Clasificación/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Proteínas de Pez Cebra/metabolismo , Pez Cebra/embriología , Análisis de Varianza , Animales , Clonación Molecular , Cartilla de ADN/genética , Perfilación de la Expresión Génica , Regulación del Desarrollo de la Expresión Génica/genética , Técnicas de Silenciamiento del Gen , Procesamiento de Imagen Asistido por Computador , Immunoblotting , Hibridación in Situ , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Proteínas de la Membrana/metabolismo , MicroARNs/genética , Análisis por Micromatrices , Modelos Biológicos , Receptores Notch/metabolismo , Retina/metabolismo , Transducción de Señal/genéticaRESUMEN
microRNAs (miRNAs) are a family of 21-23 nucleotide endogenous non-coding RNAs that post-transcriptionally regulate gene expression in a sequence-specific manner. Typically, miRNAs downregulate target genes by recognizing and recruiting protein complexes to 3'UTRs, followed by translation repression or mRNA degradation. miR-92 is a well-studied oncogene in mammalian systems. Here, using zebrafish as a model system, we uncovered a novel tissue-inductive role for miR-92 during early vertebrate development. Overexpression resulted in reduced endoderm formation during gastrulation with consequent cardia and viscera bifida. By contrast, depletion of miR-92 increased endoderm formation, which led to abnormal Kupffer's vesicle development and left-right patterning defects. Using target prediction algorithms and reporter constructs, we show that gata5 is a target of miR-92. Alteration of gata5 levels reciprocally mirrored the effects of gain and loss of function of miR-92. Moreover, genetic epistasis experiments showed that miR-92-mediated defects could be substantially suppressed by modulating gata5 levels. We propose that miR-92 is a critical regulator of endoderm formation and left-right asymmetry during early zebrafish development and provide the first evidence for a regulatory function for gata5 in the formation of Kupffer's vesicle and left-right patterning.
Asunto(s)
Tipificación del Cuerpo/genética , Endodermo/embriología , MicroARNs/fisiología , Pez Cebra/embriología , Animales , Animales Modificados Genéticamente , Secuencia de Bases , Cardias/embriología , Cardias/metabolismo , Embrión no Mamífero , Endodermo/metabolismo , Factor de Transcripción GATA5/genética , Factor de Transcripción GATA5/metabolismo , Regulación del Desarrollo de la Expresión Génica , MicroARNs/genética , MicroARNs/metabolismo , Distribución Tisular , Vísceras/embriología , Vísceras/metabolismo , Pez Cebra/genética , Pez Cebra/metabolismoRESUMEN
microRNAs (miRNAs) are small ( approximately 22 nt) noncoding RNAs that have been shown to regulate gene expression post-transcriptionally. They function by pairing with the 3' UTR of target mRNAs and repressing translation or by targeting the mRNA for degradation. miRNAs are involved in diverse aspects of development, maintenance, and disease, and are largely evolutionarily conserved in metazoans. Searching the genomes of organisms from viruses to worms to humans has revealed potentially thousands of miRNA genes. Understanding the patterns of genomic organization between species cannot only help to refine tools to identify new miRNAs, but also provide insight into miRNA biogenesis and function.
Asunto(s)
Regulación de la Expresión Génica , Genoma , MicroARNs/metabolismo , ARN Mensajero/metabolismo , Regiones no Traducidas 3' , Animales , Evolución Molecular , Genómica , Humanos , MicroARNs/biosíntesis , Biosíntesis de Proteínas , Estabilidad del ARN , Especificidad de la EspecieRESUMEN
SNAP-25 is a core component of the trimeric SNARE complex mediating vesicle exocytosis during membrane addition for neuronal growth, neuropeptide/growth factor secretion, and neurotransmitter release during synaptic transmission. Here, we report a novel microRNA mechanism of SNAP-25 regulation controlling motor neuron development, neurosecretion, synaptic activity, and movement in zebrafish. Loss of miR-153 causes overexpression of SNAP-25 and consequent hyperactive movement in early zebrafish embryos. Conversely, overexpression of miR-153 causes SNAP-25 down regulation resulting in near complete paralysis, mimicking the effects of treatment with Botulinum neurotoxin. miR-153-dependent changes in synaptic activity at the neuromuscular junction are consistent with the observed movement defects. Underlying the movement defects, perturbation of miR-153 function causes dramatic developmental changes in motor neuron patterning and branching. Together, our results indicate that precise control of SNAP-25 expression by miR-153 is critically important for proper neuronal patterning as well as neurotransmission.