RESUMEN
Mitogen-activated protein (MAP) kinases mediate cellular responses to a wide variety of stimuli. Activation of a MAP kinase occurs after phosphorylation by an upstream dual-specificity protein kinase, known as a MAP kinase kinase or MEK. The Arabidopsis thaliana genome encodes 10 MEKs but few of these have been shown directly to activate any of the 20 Arabidopsis MAP kinases. We show here that functional complementation of the cell lysis phenotype of a mutant yeast strain depends on the co-expression of the Arabidopsis MEK AtMKK6 and the MAP kinase AtMPK13. The kinase activity of AtMPK13 is stimulated in the presence of AtMKK6 in yeast cells. RT-PCR analysis showed the co-expression of these two genes in diverse plant tissues. These data show that AtMKK6 can functionally activate the MAP kinase AtMPK13.
Asunto(s)
Arabidopsis/enzimología , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Arabidopsis/genética , Activación Enzimática , Expresión Génica , Prueba de Complementación Genética , Genoma de Planta , Quinasas de Proteína Quinasa Activadas por Mitógenos/genética , Proteínas Quinasas Activadas por Mitógenos/genética , Mutación , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Saccharomyces cerevisiae/genética , Transducción de Señal , Técnicas del Sistema de Dos HíbridosRESUMEN
Higher plant microspores, when subjected to various stress treatments in vitro, are able to reprogram their regular gametophytic development towards the sporophytic pathway to form haploid embryos and plants. Suppression subtractive hybridization (SSH) and metabolic profiling were used to characterize this developmental switch. Following differential reverse Northern hybridizations 90 distinct up-regulated sequences were identified in stressed, embryogenic microspores (accessible at www.univie.ac.at/ntsm). Sequence analyses allowed the classification of these genes into functional clusters such as metabolism, chromosome remodeling, signaling, transcription and translation, while the putative functions of half of the sequences remained unknown. A comparison of metabolic profiles of non-stressed and stressed microspores using gas chromatography/mass spectrometry (GC/MS) identified 70 compounds, partly displaying significant changes in metabolite levels, e.g., highly elevated levels of isocitrate and isomaltose in stressed microspores compared to non-stressed microspores. The formation of embryogenic microspores is discussed on the basis of the identified transcriptional and metabolic profiles.
Asunto(s)
Nicotiana/genética , Nicotiana/metabolismo , Esporas/genética , Esporas/metabolismo , Northern Blotting , Cromatografía de Gases y Espectrometría de Masas , Regulación del Desarrollo de la Expresión Génica , Regulación de la Expresión Génica de las Plantas , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Nicotiana/embriologíaRESUMEN
The Igf2r imprinted cluster is an epigenetic silencing model in which expression of a ncRNA silences multiple genes in cis. Here, we map a 250 kb region in mouse embryonic fibroblast cells to show that histone modifications associated with expressed and silent genes are mutually exclusive and localized to discrete regions. Expressed genes were modified at promoter regions by H3K4me3 + H3K4me2 + H3K9Ac and on putative regulatory elements flanking active promoters by H3K4me2 + H3K9Ac. Silent genes showed two types of nonoverlapping profile. One type spread over large domains of tissue-specific silent genes and contained H3K27me3 alone. A second type formed localized foci on silent imprinted gene promoters and a nonexpressed pseudogene and contained H3K9me3 + H4K20me3 +/- HP1. Thus, mammalian chromosome arms contain active chromatin interspersed with repressive chromatin resembling the type of heterochromatin previously considered a feature of centromeres, telomeres, and the inactive X chromosome.
Asunto(s)
Cromatina/genética , Cromatina/metabolismo , Proteínas Cromosómicas no Histona/metabolismo , Regulación de la Expresión Génica , Genoma , Impresión Genómica , Histonas/metabolismo , Animales , Inmunoprecipitación de Cromatina , Proteínas Cromosómicas no Histona/genética , Cromosomas , Embrión de Mamíferos , Fibroblastos/metabolismo , Silenciador del Gen , Metilación , Ratones , Mapeo Físico de CromosomaRESUMEN
Mitotic progression is timely regulated by the accumulation and degradation of A- and B-type cyclins. In plants, there are three classes of A-, and two classes of B-type cyclins, but their specific roles are not known. We have generated transgenic tobacco plants in which the ectopic expression of a plant cyclin B2 gene is under the control of a tetracycline-inducible promoter. We show that the induction of cyclin B2 expression in cultured cells during G2 phase accelerates the entry into mitosis and allows cells to override the replication checkpoint induced by hydroxyurea in the simultaneous presence of caffeine or okadaic acid, drugs that are known to alleviate checkpoint control. These results indicate that in plants, a B2-type cyclin is a rate-limiting regulator for the entry into mitosis and a cyclin B2-CDK complex might be a target for checkpoint control pathways. The cyclin B2 localization and the timing of its degradation during mitosis corroborate these conclusions: cyclin B2 protein is confined to the nucleus and during mitosis it is only present during a short time window until mid prophase, but it is effectively degraded from this timepoint onwards. Although cyclin B2 is not present in cells arrested by the spindle checkpoint in metaphase, cyclin B1 is accumulating in these cells. Ectopic expression of cyclin B2 in developing plants interferes with differentiation events and specifically blocks root regeneration, indicating the importance of control mechanisms at the G2- to M-phase transition during plant developmental processes.