RESUMO
Plant development is profoundly regulated by ambient light cues through the red/far-red photoreceptors, the phytochromes. Early phytochrome signaling events include the translocation of phytochromes from the cytoplasm to subnuclear domains called photobodies and the degradation of antagonistically acting phytochrome-interacting factors (PIFs). We recently identified a key phytochrome signaling component, HEMERA (HMR), that is essential for both phytochrome B (phyB) localization to photobodies and PIF degradation. However, the signaling mechanism linking phytochromes and HMR is unknown. Here we show that phytochromes directly interact with HMR to promote HMR protein accumulation in the light. HMR binds more strongly to the active form of phytochromes. This interaction is mediated by the photosensory domains of phytochromes and two phytochrome-interacting regions in HMR. Missense mutations in either HMR or phyB that alter the phytochrome/HMR interaction can also change HMR levels and photomorphogenetic responses. HMR accumulation in a constitutively active phyB mutant (YHB) is required for YHB-dependent PIF3 degradation in the dark. Our genetic and biochemical studies strongly support a novel phytochrome signaling mechanism in which photoactivated phytochromes directly interact with HMR and promote HMR accumulation, which in turn mediates the formation of photobodies and the degradation of PIFs to establish photomorphogenesis.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Luz , Fitocromo/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Mutação , Ligação Proteica , Estrutura Terciária de Proteína , Transdução de SinaisRESUMO
Methods to assess the diversity of the diazotroph assemblage in the rhizosphere of the salt marsh cordgrass, Spartina alterniflora were examined. The effectiveness of nifH PCR-denaturing gradient gel electrophoresis (DGGE) was compared to that of nifH clone library analysis. Seventeen DGGE gel bands were sequenced and yielded 58 nonidentical nifH sequences from a total of 67 sequences determined. A clone library constructed using the GC-clamp nifH primers that were employed in the PCR-DGGE (designated the GC-Library) yielded 83 nonidentical sequences from a total of 257 nifH sequences. A second library constructed using an alternate set of nifH primers (N-Library) yielded 83 nonidentical sequences from a total of 138 nifH sequences. Rarefaction curves for the libraries did not reach saturation, although the GC-Library curve was substantially dampened and appeared to be closer to saturation than the N-Library curve. Phylogenetic analyses showed that DGGE gel band sequencing recovered nifH sequences that were frequently sampled in the GC-Library, as well as sequences that were infrequently sampled, and provided a species composition assessment that was robust, efficient, and relatively inexpensive to obtain. Further, the DGGE method permits a large number of samples to be examined for differences in banding patterns, after which bands of interest can be sampled for sequence determination.
Assuntos
Bactérias/classificação , Bactérias/genética , Biodiversidade , DNA Bacteriano/genética , Poaceae/microbiologia , Bactérias/isolamento & purificação , Proteínas de Bactérias/genética , Análise por Conglomerados , Primers do DNA/genética , DNA Bacteriano/química , Eletroforese em Gel de Campo Pulsado , Dados de Sequência Molecular , Oxirredutases/genética , Filogenia , Raízes de Plantas/microbiologia , Análise de Sequência de DNA , Homologia de Sequência , Microbiologia do SoloRESUMO
Esa1 is the only essential histone acetyltransferase (HAT) in budding yeast. It is the catalytic subunit of at least two multiprotein complexes, NuA4 and Piccolo NuA4 (picNuA4), and its essential function is believed to be its catalytic HAT activity. To examine the role of Esa1 in DNA damage repair, we isolated viable esa1 mutants with a range of hypersensitivities to the toposide camptothecin. Here we show that the sensitivity of these mutants to a variety of stresses is inversely proportional to their level of histone H4 acetylation, demonstrating the importance of Esa1 catalytic activity for resistance to genotoxic stress. Surprisingly, individual mutations in two residues directly involved in catalysis were not lethal even though the mutant enzymes appear catalytically inactive both in vivo and in vitro. However, the double-point mutant is lethal, demonstrating that the essential function of Esa1 relies on residues within the catalytic pocket but not catalysis. We propose that the essential function of Esa1 may be to bind acetyl-CoA or lysine substrates and positively regulate the activities of NuA4 and Piccolo NuA4.
