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1.
Cell ; 187(5): 1127-1144.e21, 2024 Feb 29.
Artículo en Inglés | MEDLINE | ID: mdl-38428393

RESUMEN

Chloroplasts are green plastids in the cytoplasm of eukaryotic algae and plants responsible for photosynthesis. The plastid-encoded RNA polymerase (PEP) plays an essential role during chloroplast biogenesis from proplastids and functions as the predominant RNA polymerase in mature chloroplasts. The PEP-centered transcription apparatus comprises a bacterial-origin PEP core and more than a dozen eukaryotic-origin PEP-associated proteins (PAPs) encoded in the nucleus. Here, we determined the cryo-EM structures of Nicotiana tabacum (tobacco) PEP-PAP apoenzyme and PEP-PAP transcription elongation complexes at near-atomic resolutions. Our data show the PEP core adopts a typical fold as bacterial RNAP. Fifteen PAPs bind at the periphery of the PEP core, facilitate assembling the PEP-PAP supercomplex, protect the complex from oxidation damage, and likely couple gene transcription with RNA processing. Our results report the high-resolution architecture of the chloroplast transcription apparatus and provide the structural basis for the mechanistic and functional study of transcription regulation in chloroplasts.


Asunto(s)
ARN Polimerasas Dirigidas por ADN , Plastidios , Cloroplastos/metabolismo , Microscopía por Crioelectrón , ARN Polimerasas Dirigidas por ADN/genética , Nicotiana/genética , Fotosíntesis , Plastidios/enzimología
2.
Yi Chuan ; 45(5): 425-434, 2023 May 20.
Artículo en Inglés | MEDLINE | ID: mdl-37194589

RESUMEN

Hypothalamic median eminence (ME) is a potential niche for neurons and oligodendrocytes, and trophic factors may regulate hypothalamic function by inducing cellular changes in the ME region. To determine whether diet-induced plasticity exists in hypothalamic stem cells dormant under physiological conditions, we used a combination of a normal diet, a high-fat diet, and a ketogenic diet (a low-carb, high-fat diet) to compare the proliferation of tanycytes (TCs) and oligodendrocyte precursor cells (OPCs) in the ME area of mice under the different diets. The results showed that the ketogenic diet could induce and promote the proliferation of OPCs in the ME area, and blocking the fatty acid oxidation program could inhibit the proliferation of OPCs induced by a ketogenic diet. This study preliminarily revealed the diet-induced effect on OPCs in the ME region and provided enlightenment for further study on the function of OPCs in the ME region.


Asunto(s)
Dieta Cetogénica , Células Precursoras de Oligodendrocitos , Ratones , Animales , Eminencia Media , Proliferación Celular , Ácidos Grasos , Diferenciación Celular
3.
Nucleic Acids Res ; 51(4): 1960-1970, 2023 02 28.
Artículo en Inglés | MEDLINE | ID: mdl-36715319

RESUMEN

Canonical bacterial transcription activators bind to their cognate cis elements at the upstream of transcription start site (TSS) in a form of dimer. Caulobacter crescentus GcrA, a non-canonical transcription activator, can activate transcription from promoters harboring its cis element at the upstream or downstream of TSS in a form of monomer. We determined two cryo-EM structures of C. crescentus GcrA-bound transcription activation complexes, GcrA TACU and GcrA TACD, which comprise GcrA, RNAP, σ70 and promoter DNA with GcrA cis elements at either the upstream or downstream of TSS at 3.6 and 3.8 Å, respectively. In the GcrA-TACU structure, GcrA makes bipartite interactions with both σ70 domain 2 (σ702) and its cis element, while in the GcrA-TACD structure, GcrA retains interaction with σ702 but loses the interaction with its cis element. Our results suggest that GcrA likely forms a functionally specialized GcrA-RNAP-σA holoenzyme, in which GcrA first locates its cis element and then facilitates RNAP to load on core promoter at its proximal region. The sequence-specific interaction of GcrA and DNA is disrupted either at the stage of RPo formation or promoter escape depending on the location of GcrA cis elements relative to TSS.


Asunto(s)
Proteínas Bacterianas , Caulobacter crescentus , Factores de Transcripción , Activación Transcripcional , Proteínas Bacterianas/metabolismo , Caulobacter crescentus/metabolismo , ADN/metabolismo , ARN Polimerasas Dirigidas por ADN/metabolismo , Regulación Bacteriana de la Expresión Génica , Factores de Transcripción/metabolismo , Transcripción Genética
4.
Nat Commun ; 13(1): 4204, 2022 07 20.
Artículo en Inglés | MEDLINE | ID: mdl-35859063

RESUMEN

Pseudomonas aeruginosa (Pae) SutA adapts bacteria to hypoxia and nutrition-limited environment during chronic infection by increasing transcription activity of an RNA polymerase (RNAP) holoenzyme comprising the stress-responsive σ factor σS (RNAP-σS). SutA shows no homology to previously characterized RNAP-binding proteins. The structure and mode of action of SutA remain unclear. Here we determined cryo-EM structures of Pae RNAP-σS holoenzyme, Pae RNAP-σS holoenzyme complexed with SutA, and Pae RNAP-σS transcription initiation complex comprising SutA. The structures show SutA pinches RNAP-ß protrusion and facilitates promoter unwinding by wedging RNAP-ß lobe open. Our results demonstrate that SutA clears an energetic barrier to facilitate promoter unwinding of RNAP-σS holoenzyme.


Asunto(s)
ARN Polimerasas Dirigidas por ADN , Pseudomonas aeruginosa , Proteínas Bacterianas/metabolismo , ADN/metabolismo , ARN Polimerasas Dirigidas por ADN/metabolismo , Holoenzimas/metabolismo , Pseudomonas aeruginosa/metabolismo , Factor sigma/metabolismo , Transcripción Genética
5.
Phytopathology ; 112(7): 1476-1485, 2022 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-35021860

RESUMEN

Sclerotinia sclerotiorum is a notorious phytopathogenic Ascomycota fungus with a host range of >600 plant species worldwide. This homothallic Leotiomycetes species reproduces sexually through a multicellular apothecium that produces and releases ascospores. These ascospores serve as the primary inoculum source for disease initiation in the majority of S. sclerotiorum disease cycles. The regulation of apothecium development for this pathogen and other apothecium-producing fungi remains largely unknown. Here, we report that a C2H2 transcription factor, SsZFH1 (zinc finger homologous protein), is necessary for the proper development and maturation of sclerotia and apothecia in S. sclerotiorum and is required for the normal growth rate of hyphae. Furthermore, ΔSszfh1 strains exhibit decreased H2O2 accumulation in hyphae, increased melanin deposition, and enhanced tolerance to H2O2 in the process of vegetative growth and sclerotia formation. Infection assays on common bean leaves, with thin cuticles, and soybean and tomato leaves, with thick cuticles, suggest that the deletion of Sszfh1 slows the mycelial growth rate, which in turn affects the expansion of leaf lesions. Collectively, our results provide novel insights into a major fungal factor mediating maturation of apothecia with additional effects on hyphae and sclerotia development.


Asunto(s)
Ascomicetos , Factores de Transcripción , Peróxido de Hidrógeno/metabolismo , Enfermedades de las Plantas/microbiología , Esporas Fúngicas , Factores de Transcripción/genética , Factores de Transcripción/metabolismo
6.
Science ; 374(6575): 1579-1586, 2021 Dec 24.
Artículo en Inglés | MEDLINE | ID: mdl-34941388

RESUMEN

DNA methylation affects gene expression and maintains genome integrity. The DNA-dependent RNA polymerase IV (Pol IV), together with the RNA-dependent RNA polymerase RDR2, produces double-stranded small interfering RNA precursors essential for establishing and maintaining DNA methylation in plants. We determined the cryo­electron microscopy structures of the Pol IV­RDR2 holoenzyme and the backtracked transcription elongation complex. These structures reveal that Pol IV and RDR2 form a complex with their active sites connected by an interpolymerase channel, through which the Pol IV­generated transcript is handed over to the RDR2 active site after being backtracked, where it is used as the template for double-stranded RNA (dsRNA) synthesis. Our results describe a 'backtracking-triggered RNA channeling' mechanism underlying dsRNA synthesis and also shed light on the evolutionary trajectory of eukaryotic RNA polymerases.


Asunto(s)
Proteínas de Arabidopsis/química , Arabidopsis/enzimología , Arabidopsis/genética , ARN Polimerasas Dirigidas por ADN/química , ARN Bicatenario/biosíntesis , ARN de Planta/biosíntesis , ARN Polimerasa Dependiente del ARN/química , Secuencias de Aminoácidos , Proteínas de Arabidopsis/metabolismo , Dominio Catalítico , Microscopía por Crioelectrón , Metilación de ADN , ADN de Plantas/metabolismo , ARN Polimerasas Dirigidas por ADN/metabolismo , Holoenzimas/química , Modelos Moleculares , Complejos Multiproteicos/química , Conformación Proteica , Dominios Proteicos , ARN Polimerasa II/química , ARN Interferente Pequeño/biosíntesis , ARN Polimerasa Dependiente del ARN/metabolismo , Elongación de la Transcripción Genética , Factores de Transcripción/metabolismo
7.
Cell Stem Cell ; 28(8): 1483-1499.e8, 2021 08 05.
Artículo en Inglés | MEDLINE | ID: mdl-33887179

RESUMEN

The hypothalamus contains an astounding heterogeneity of neurons that regulate endocrine, autonomic, and behavioral functions. However, its molecular developmental trajectory and origin of neuronal diversity remain unclear. Here, we profile the transcriptome of 43,261 cells derived from Rax+ hypothalamic neuroepithelium to map the developmental landscape of the mouse hypothalamus and trajectory of radial glial cells (RGCs), intermediate progenitor cells (IPCs), nascent neurons, and peptidergic neurons. We show that RGCs adopt a conserved strategy for multipotential differentiation but generate Ascl1+ and Neurog2+ IPCs. Ascl1+ IPCs differ from their telencephalic counterpart by displaying fate bifurcation, and postmitotic nascent neurons resolve into multiple peptidergic neuronal subtypes. Clonal analysis further demonstrates that single RGCs can produce multiple neuronal subtypes. Our study reveals that multiple cell types along the lineage hierarchy contribute to fate diversification of hypothalamic neurons in a stepwise fashion, suggesting a cascade diversification model that deconstructs the origin of neuronal diversity.


Asunto(s)
Hipotálamo , Neuronas , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Diferenciación Celular , Regulación del Desarrollo de la Expresión Génica , Hipotálamo/metabolismo , Ratones , Proteínas del Tejido Nervioso/metabolismo , Neuronas/metabolismo , Células Madre/metabolismo
8.
Nat Commun ; 12(1): 2288, 2021 04 16.
Artículo en Inglés | MEDLINE | ID: mdl-33863883

RESUMEN

Hypothalamic tanycytes in median eminence (ME) are emerging as a crucial cell population that regulates endocrine output, energy balance and the diffusion of blood-born molecules. Tanycytes have recently been considered as potential somatic stem cells in the adult mammalian brain, but their regenerative and tumorigenic capacities are largely unknown. Here we found that Rax+ tanycytes in ME of mice are largely quiescent but quickly enter the cell cycle upon neural injury for self-renewal and regeneration. Mechanistically, Igf1r signaling in tanycytes is required for tissue repair under injury conditions. Furthermore, Braf oncogenic activation is sufficient to transform Rax+ tanycytes into actively dividing tumor cells that eventually develop into a papillary craniopharyngioma-like tumor. Together, these findings uncover the regenerative and tumorigenic potential of tanycytes. Our study offers insights into the properties of tanycytes, which may help to manipulate tanycyte biology for regulating hypothalamic function and investigate the pathogenesis of clinically relevant tumors.


Asunto(s)
Craneofaringioma/patología , Células Ependimogliales/fisiología , Eminencia Media/fisiología , Neoplasias Experimentales/patología , Regeneración , Animales , Carcinogénesis/patología , Autorrenovación de las Células/fisiología , Craneofaringioma/inducido químicamente , Craneofaringioma/genética , Proteínas del Ojo/metabolismo , Femenino , Proteínas de Homeodominio/metabolismo , Eminencia Media/citología , Ratones , Neoplasias Experimentales/inducido químicamente , Neoplasias Experimentales/genética , Proteínas Proto-Oncogénicas B-raf/genética , RNA-Seq , Receptor IGF Tipo 1/metabolismo , Transducción de Señal , Análisis de la Célula Individual , Factores de Transcripción/metabolismo
9.
PLoS Biol ; 16(4): e2005211, 2018 04.
Artículo en Inglés | MEDLINE | ID: mdl-29684005

RESUMEN

The thalamus, a crucial regulator of cortical functions, is composed of many nuclei arranged in a spatially complex pattern. Thalamic neurogenesis occurs over a short period during mammalian embryonic development. These features have hampered the effort to understand how regionalization, cell divisions, and fate specification are coordinated and produce a wide array of nuclei that exhibit distinct patterns of gene expression and functions. Here, we performed in vivo clonal analysis to track the divisions of individual progenitor cells and spatial allocation of their progeny in the developing mouse thalamus. Quantitative analysis of clone compositions revealed evidence for sequential generation of distinct sets of thalamic nuclei based on the location of the founder progenitor cells. Furthermore, we identified intermediate progenitor cells that produced neurons populating more than one thalamic nuclei, indicating a prolonged specification of nuclear fate. Our study reveals an organizational principle that governs the spatial and temporal progression of cell divisions and fate specification and provides a framework for studying cellular heterogeneity and connectivity in the mammalian thalamus.


Asunto(s)
Células Clonales/metabolismo , Regulación del Desarrollo de la Expresión Génica , Células-Madre Neurales/metabolismo , Neurogénesis/genética , Neuronas/metabolismo , Tálamo/metabolismo , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Diferenciación Celular , División Celular , Linaje de la Célula , Rastreo Celular/métodos , Células Clonales/citología , Embrión de Mamíferos , Femenino , Genes Reporteros , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Integrasas/genética , Integrasas/metabolismo , Ratones , Ratones Transgénicos , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Células-Madre Neurales/citología , Neuronas/citología , Embarazo , Tálamo/citología , Tálamo/crecimiento & desarrollo , Proteína con Dedos de Zinc GLI1/genética , Proteína con Dedos de Zinc GLI1/metabolismo
10.
Mol Plant Pathol ; 19(7): 1679-1689, 2018 07.
Artículo en Inglés | MEDLINE | ID: mdl-29227022

RESUMEN

The sclerotium, a multicellular structure composed of the compact aggregation of vegetative hyphae, is critical for the long-term survival and sexual reproduction of the plant-pathogenic fungus Sclerotinia sclerotiorum. The development and carpogenic germination of sclerotia are regulated by integrating signals from both environmental and endogenous processes. Here, we report the regulatory functions of the S. sclerotiorum GATA-type IVb zinc-finger transcription factor SsNsd1 in these processes. SsNsd1 is orthologous to the Aspergillus nidulans NsdD (never in sexual development) and the Neurospora crassa SUB-1 (submerged protoperithecia-1) proteins. Ssnsd1 gene transcript accumulation remains relatively low, but variable, during vegetative mycelial growth and multicellular development. Ssnsd1 deletion mutants (Δnsd1-KOs) produce phialides and phialospores (spermatia) excessively in vegetative hyphae and promiscuously within the interior medulla of sclerotia. In contrast, phialospore development occurs only on the sclerotium surface in the wild-type. Loss of SsNsd1 function affects sclerotium structural integrity and disrupts ascogonia formation during conditioning for carpogenic germination. As a consequence, apothecium development is abolished. The Ssnsd1 deletion mutants are also defective in the transition from hyphae to compound appressorium formation, resulting in a loss of pathogenicity on unwounded hosts. In sum, our results demonstrate that SsNsd1 functions in a regulatory role similar to its ascomycete orthologues in regulating sexual and asexual development. Further, SsNsd1 appears to have evolved as a regulator of pre-penetration infectious development required for the successful infection of its many hosts.


Asunto(s)
Ascomicetos/metabolismo , Ascomicetos/patogenicidad , Proteínas Fúngicas/metabolismo , Factores de Transcripción/metabolismo , Ascomicetos/genética , Proteínas Fúngicas/genética , Regulación Fúngica de la Expresión Génica , Factores de Transcripción/genética
11.
Int J Mol Sci ; 15(5): 8049-62, 2014 May 08.
Artículo en Inglés | MEDLINE | ID: mdl-24815067

RESUMEN

MADS-box proteins, a well-conserved family of transcription factors in eukaryotic organisms, specifically regulate a wide range of cellular functions, including primary metabolism, cell cycle, and cell identity. However, little is known about roles of the MADS-box protein family in the fungal pathogen Sclerotinia sclerotiorum. In this research, the S. sclerotiorum MADS-box gene SsMADS was cloned; it encodes a protein that is highly similar to Mcm1 orthologs from Saccharomyces cerevisiae and other fungi, and includes a highly conserved DNA-binding domain. MADS is a member of the MADS box protein SRF (serum response factor) lineage. SsMADS function was investigated using RNA interference. Silenced strains were obtained using genetic transformation of the RNA interference vectors pS1-SsMADS and pSD-SsMADS. SsMADS expression levels in silenced strains were analyzed using RT-PCR. The results showed that SsMADS mRNA expression in these silenced strains was reduced to different degrees, and growth rate in these silenced strains was significantly decreased. Infecting tomato leaflets with silenced strains indicated that SsMADS was required for leaf pathogenesis in a susceptible host. Our results suggest that the MADS-box transcription factor SsMADS is involved in S. sclerotiorum growth and virulence.


Asunto(s)
Ascomicetos/crecimiento & desarrollo , Ascomicetos/patogenicidad , Proteínas Fúngicas/genética , Enfermedades de las Plantas/microbiología , Solanum lycopersicum/microbiología , Factores de Transcripción/genética , Secuencia de Aminoácidos , Ascomicetos/genética , Proteínas Fúngicas/química , Datos de Secuencia Molecular , Filogenia , Interferencia de ARN , ARN Interferente Pequeño/genética , Alineación de Secuencia , Factores de Transcripción/química , Virulencia
12.
J Environ Sci Health B ; 47(3): 153-60, 2012.
Artículo en Inglés | MEDLINE | ID: mdl-22375586

RESUMEN

By enrichment culturing of the sludge collected from the industrial wastewater treatment pond, we isolated a highly efficient nicosulfuron degrading bacterium Serratia marcescens N80. In liquid medium, Serratia marcescens N80 grows using nicosulfuron as the sole nitrogen source, and the optimal temperature, pH values, and inoculation for degradation are 30-35°C, 6.0-7.0, and 3.0% (v/v), respectively. With the initial concentration of 10 mg L⁻¹, the degradation rate is 93.6% in 96 hours; as the initial concentrations are higher than 10 mg L⁻¹, the biodegradation rates decrease as the nicosulfuron concentrations increase; when the concentration is 400 mg L⁻¹, the degradation rate is only 53.1%. Degradation follows the pesticide degradation kinetic equation at concentrations between 5 mg L⁻¹ and 50 mg L⁻¹. Identification of the metabolites by the liquid chromatography/mass spectrometry (LC/MS) indicates that the degradation of nicosulfuron is achieved by breaking the sulfonylurea bridge. The strain N80 also degraded some other sulfonylurea herbicides, including ethametsulfuron, tribenuron-methyl, metsulfuron-methyl, chlorimuron-ethyl,and rimsulfuron.


Asunto(s)
Herbicidas/metabolismo , Piridinas/metabolismo , Serratia marcescens/metabolismo , Compuestos de Sulfonilurea/metabolismo , Secuencia de Bases , Biodegradación Ambiental , Cromatografía Liquida , ADN Bacteriano , Concentración de Iones de Hidrógeno , Espectrometría de Masas , Datos de Secuencia Molecular , Filogenia , Reacción en Cadena de la Polimerasa , ARN Ribosómico 16S , Análisis de Secuencia de ADN , Serratia marcescens/clasificación , Serratia marcescens/genética , Serratia marcescens/aislamiento & purificación , Aguas del Alcantarillado/microbiología , Temperatura
13.
J Environ Sci Health B ; 45(6): 501-7, 2010 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-20574870

RESUMEN

Enrichment culturing of sludge taken from an industrial wastewater treatment pond led to the identification of a bacterium (Klebsiella jilinsis H. Zhang) that degrades chlorimuron-ethyl with high efficiency. Klebsiella jilinsis strain 2N3 grows with chlorimuron-ethyl as the sole nitrogen source at the optimal temperature range of 30-35 degrees C and pH values between 6.0-7.0. In liquid medium, the degradation activity was further induced by chlorimuron-ethyl. Degradation rates followed the pesticide degradation kinetic equation at concentrations between 20 and 200 mg L(-1). Using initial concentrations of 20 and 100 mg L(-1), the degradation rates of chlorimuron-ethyl were 83.5 % and 92.5 % in 12 hours, respectively. At an initial concentration higher than 200 mg L(-1), the degradation rate decreased slightly as the concentration increased. The 2N3 strain also degraded the sulfonylurea herbicides ethametsulfuron, metsulfuron-methyl, nicosulfuron, rimsulfuron, and tribenuron-methyl. This study provides scientific evidence and support for the application of K. jilinsis in bioremediation to reduce environmental pollution.


Asunto(s)
Klebsiella/aislamiento & purificación , Klebsiella/metabolismo , Plaguicidas/metabolismo , Pirimidinas/metabolismo , Aguas del Alcantarillado/microbiología , Compuestos de Sulfonilurea/metabolismo , Biodegradación Ambiental , ADN Bacteriano/genética , ADN Ribosómico/genética , Cinética , Klebsiella/clasificación , Klebsiella/genética , Datos de Secuencia Molecular , Plaguicidas/química , Filogenia , Pirimidinas/química , ARN Ribosómico 16S/genética , Compuestos de Sulfonilurea/química
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