La enseñanza bíblica en profundidad de John MacArthur lleva la verdad transformadora de la Palabra de Dios a millones de personas cada día.

La enseñanza bíblica en profundidad de John MacArthur lleva la verdad transformadora de la Palabra de Dios a millones de personas cada día.

La enseñanza bíblica en profundidad de John MacArthur lleva la verdad transformadora de la Palabra de Dios a millones de personas cada día.

La enseñanza bíblica en profundidad de John MacArthur lleva la verdad transformadora de la Palabra de Dios a millones de personas cada día.

La enseñanza bíblica en profundidad de John MacArthur lleva la verdad transformadora de la Palabra de Dios a millones de personas cada día.

La enseñanza bíblica en profundidad de John MacArthur lleva la verdad transformadora de la Palabra de Dios a millones de personas cada día.

La enseñanza bíblica en profundidad de John MacArthur lleva la verdad transformadora de la Palabra de Dios a millones de personas cada día.

La enseñanza bíblica en profundidad de John MacArthur lleva la verdad transformadora de la Palabra de Dios a millones de personas cada día.

La enseñanza bíblica en profundidad de John MacArthur lleva la verdad transformadora de la Palabra de Dios a millones de personas cada día.

La enseñanza bíblica en profundidad de John MacArthur lleva la verdad transformadora de la Palabra de Dios a millones de personas cada día.

La enseñanza bíblica en profundidad de John MacArthur lleva la verdad transformadora de la Palabra de Dios a millones de personas cada día.

La enseñanza bíblica en profundidad de John MacArthur lleva la verdad transformadora de la Palabra de Dios a millones de personas cada día.

La enseñanza bíblica en profundidad de John MacArthur lleva la verdad transformadora de la Palabra de Dios a millones de personas cada día.

La enseñanza bíblica en profundidad de John MacArthur lleva la verdad transformadora de la Palabra de Dios a millones de personas cada día.

La enseñanza bíblica en profundidad de John MacArthur lleva la verdad transformadora de la Palabra de Dios a millones de personas cada día.

La enseñanza bíblica en profundidad de John MacArthur lleva la verdad transformadora de la Palabra de Dios a millones de personas cada día.

La enseñanza bíblica en profundidad de John MacArthur lleva la verdad transformadora de la Palabra de Dios a millones de personas cada día.

La enseñanza bíblica en profundidad de John MacArthur lleva la verdad transformadora de la Palabra de Dios a millones de personas cada día.

La enseñanza bíblica en profundidad de John MacArthur lleva la verdad transformadora de la Palabra de Dios a millones de personas cada día.

La enseñanza bíblica en profundidad de John MacArthur lleva la verdad transformadora de la Palabra de Dios a millones de personas cada día.

La enseñanza bíblica en profundidad de John MacArthur lleva la verdad transformadora de la Palabra de Dios a millones de personas cada día.

The Charities Act 2006 (now the Charities Act 2011) introduced a new form of incorporated charitable entity, the CIO.  There has been a long delay in making CIOs available but their imminent arrival has been heralded by a recent government anno...

Es significativo que uno de los nombres bíblicos de Cristo sea Admirable, Consejero (Isaías 9:6). Él es el Consejero Supremo y Definitivo a quien podemos acudir en busca de consejo; y Su Palabra es la fuente desde donde podemos extraer sabiduría divina. ¿Qué podría ser más admirable que eso? 

De hecho, uno de los aspectos más gloriosos de la perfecta suficiencia de Cristo es el consejo admirable y gran sabiduría que Él suple en tiempos de desesperación, confusión, miedo, ansiedad y tristeza. Él es el Consejero por excelencia. 

Ahora, esto no es para menospreciar la importancia de los cristianos aconsejándose unos a otros. Hay ciertamente una necesidad crucial de consejería bíblica sana dentro del cuerpo de Cristo. No debatiría ni por un momento el importante rol de quienes están dotados espiritualmente para dar aliento, discernimiento, consuelo, consejo, compasión y ayuda a otros. 

De hecho, uno de los problemas que ha llevado a la actual plaga de mala consejería es que las iglesias no han hecho tanto bien como deberían, permitiendo que las personas con esos dones espirituales ministren con excelencia. Las complejidades de esta época moderna hacen mucho más difícil que nunca el tomar el tiempo necesario para escuchar con atención, servir a otros a través de la entrega personal compasiva y proveer la comunión cercana, necesaria para que el cuerpo de la iglesia disfrute de salud y vitalidad.  

 Las iglesias han mirado a la psicología para llenar el vacío, pero no va a funcionar. Los psicólogos profesionales no son sustitutos de gente dotada espiritualmente; y el consejo que la psicología ofrece no puede reemplazar la sabiduría bíblica y el poder divino. Por otra parte, la psicología tiende a volver a la gente dependiente de un terapista, mientras que aquellos que ejercitan dones espirituales verdaderos, llevan a la gente hacia un Salvador todo suficiente y Su Palabra toda suficiente.  

Un Salmo sobre la Suficiencia de la Palabra de Dios 

El rey David fue un ejemplo de alguien quien ocasionalmente buscó consejo de consejeros humanos, pero al final, siempre recurría a Dios en busca de respuestas. Como muchos de los salmos revelan, él era especialmente dependiente únicamente de Dios cuando luchaba con problemas o emociones personales (Salmos 18). Cuando le golpeó la depresión o confusión interna, se volvió hacia Dios y luchó en oración (Salmos 73). Cuando el problema era su propio pecado, él se mostró arrepentido, quebrantado y contrito (Salmos 51). La persona madura espiritualmente siempre se vuelve a Dios por ayuda en tiempos de ansiedad, angustia, confusión o inquietud en el alma ––y se aseguran consejo sabio y liberación. 

Eso es debido a que toda necesidad del alma humana es básicamente espiritual. No existe algo llamado ‘problema psicológico’ no relacionado a causas espirituales o físicas. Dios suple recursos divinos suficientes para suplir todas esas necesidades por completo. David entendió eso. 

Sus escritos reflejaron la profundidad de la experiencia humana, emoción y conocimiento espiritual de alguien quien ha experimentado plenamente los extremos de la vida. Él conoció el regocijo de pasar de pastor a rey. Él escribió acerca de todo, desde el triunfo absoluto, hasta el desaliento amargo. Él luchó con un dolor tan profundo que apenas podía soportar vivir. 

Su propio hijo Absalón intentó matarlo y luego, fue asesinado. Él sufrió una culpabilidad horrible debido a la inmoralidad y asesinato. Sus hijos le trajeron constante aflicción. Él luchaba por entender las dos cosas ––la naturaleza de Dios y su propio corazón. De Dios, él dijo: “Grande es Jehová” (Salmos 145:3), mientras que de sí mismo él dijo: “Lávame más y más de mi maldad, y límpiame de mi pecado” (Salmos 51:2). Él le dijo a Dios lo qué sentía y clamó por alivio ––a pesar que admitió que Dios tenía todo el derecho de castigarlo. 

Al final de algunos de los salmos de David, él miró por una ventana de esperanza, y otras veces, no. Pero David siempre fue a Dios, porque entendió la soberanía de Dios y su propia depravación. Él sabía que sólo su todo-suficiente Salvador tenía las respuestas a sus necesidades y el poder de aplicar las respuestas. Y sabía que esas respuestas eran encontradas en la verdad acerca de Dios revelada en Su Palabra, que es en sí misma perfectamente suficiente. El Dios suficiente se reveló a Sí mismo en Su Palabra suficiente. 

El Salmo 19:7-14 es la declaración más monumental sobre la suficiencia de las Escrituras, que jamás se haya hecho en términos concisos. Escrito por David bajo la inspiración del Espíritu Santo, ofrece un testimonio inquebrantable de Dios mismo acerca de la suficiencia de Su Palabra para cada situación. Refuta las enseñanzas de aquellos que creen que debemos ampliar la Palabra de Dios con verdad obtenida de la psicología moderna. 

En los versículos 7 al 9, David hace seis afirmaciones acerca de las Escrituras. Cada una de las seis afirmaciones resalta una característica de la Palabra de Dios y describe su efecto en la vida de quien la recibe. 

La Escritura es perfecta, convierte el alma. 

En la primera afirmación (v.7), él dice: “La ley de Jehová es perfecta, que convierte el alma”. La Palabra hebrea traducida “ley” es torah, que enfatiza la naturaleza didáctica de la Escritura. Aquí, David la usa para referirse a la suma de lo que Dios ha revelado para nuestra instrucción, sea un credo (lo que creemos), carácter (lo que somos) o conducta (lo que hacemos). 

“Perfecta” es la traducción de una palabra hebrea que significa “entero”, “completo” o “suficiente”. Comunica la idea de algo que es integral, al punto de cubrir todos los aspectos de un problema. La Escritura es integral, corporizando todo lo que es necesario para la vida espiritual de uno. El contraste implícito de David es con el razonamiento imperfecto, insuficiente e incorrecto de los hombres. 

La ley perfecta de Dios, dijo David, afecta a las personas porque “convierte el alma” (v.7). La palabra hebrea traducida “convierte” puede significar “restaura”, “revive” o “refresca”; pero mi sinónimo favorito es “transforma”. La palabra “alma” (en hebreo, nephesh) se refiere a la persona de uno, uno mismo o el corazón. Es traducida de todas esas maneras (y muchas más) en el Antiguo Testamento. La esencia de ella es la persona interior, la persona completa, el verdadero usted. 

Parafraseando las palabras de David, las Escrituras son tan poderosas e integrales que pueden convertir o transformar toda la persona, convirtiendo a alguien exactamente en la persona que Dios quiere que sea. La Palabra de Dios es suficiente para restaurar mediante la salvación incluso a la vida más destrozada, un hecho del que David mismo dio abundante testimonio. 

La Escritura es digna de confianza, imparte sabiduría. 

David desarrolla aún más el alcance de la suficiencia de las Escrituras en el Salmo 19:7, “El testimonio de Jehová es fiel, que hace sabio al sencillo”. “Testimonio” habla de la Escritura como un testigo divino. La Escritura es el testimonio seguro de Dios sobre quién Él es y lo que Él requiere de nosotros. “Fiel” significa que su testimonio es inquebrantable, inamovible, inconfundible, confiable y digno de confianza. Proporciona una base sobre la cual construir nuestras vidas y destinos eternos. 

La Palabra fiel de Dios convierte al sencillo en sabio (v.7). La palabra hebrea traducida “sencillo” viene de una expresión que significa “una puerta abierta”. Evoca la imagen de una persona inocente que no sabe cuándo cerrar su mente a la enseñanza falsa o impura. Él es falto de discernimiento, ignorante, crédulo. Pero la Palabra de Dios lo hace sabio. “Sabio” no habla simplemente de alguien que conoce algunos hechos, sino de uno que es hábil en el arte de la vida piadosa. Él se somete a las Escrituras y sabe cómo aplicarla a sus circunstancias. De esta manera, la Palabra de Dios toma una mente simple y sin discernimiento, y la capacita en todas las cuestiones de la vida. Esto también es en contraste con la sabiduría de los hombres, que en realidad es necedad. 

La Escritura es recta, produce gozo 

David agrega una tercera declaración acerca de las Escrituras. Él escribe: “Los mandamientos de Jehová son rectos, que alegran el corazón”. Los mandamientos son principios divinos y guías para el carácter y la conducta. Ya que Dios nos creó y sabe cómo debemos vivir para ser productivos para Su gloria, Él ha puesto en las Escrituras todos los principios que necesitamos para vivir una vida piadosa. 

Los mandamientos de Dios, dijo David, son “rectos”. En lugar de indicar simplemente qué es correcto y qué es erróneo, la Palabra tiene el sentido de enseñarle a alguien el camino verdadero. Las verdades de la Escritura establecen el camino correcto a través del difícil laberinto de la vida. Esa es una confianza maravillosa. Muchas personas hoy están angustiadas o abatidas porque carecen de dirección y propósito. La mayoría busca respuestas en las fuentes equivocadas. La Palabra de Dios no solamente provee la luz para nuestro camino (Salmo 119:105), sino que establece la ruta enfrente nuestro. 

Debido a que nos conduce a través del camino recto en la vida, la Palabra de Dios trae gran gozo. Si usted está deprimido, ansioso, temeroso o dudoso, aprenda a obedecer el consejo de Dios y comparta el deleite resultante. No recurra a actividades autoindulgentes como la autoestima y la autorrealización. Enfóquese en la verdad divina. Ahí encontrará el gozo verdadero y duradero. Todas las otras fuentes son temporales y fugaces. 

¿No es la Palabra de Dios asombrosa en su suficiencia? Es perfecta, no le falta nada, es confiable y establece el rumbo para una vida productiva. Como tal, nos transforma a la imagen de Cristo, nos otorga sabiduría para cada momento y nos llena con gozo eterno.

¡Qué trágico que es cuando dejamos de lado la fuente de sabiduría divina, prefiriendo la sabiduría del hombre, que es impotente e insuficiente! 

La próxima vez, veremos las siguientes tres declaraciones de la suficiencia de las Escrituras, y saborearemos la dulzura de la Palabra de Dios. 

Únase a la conversación                                                                                                                           

¿Qué Escrituras han tenido un impacto profundo en su vida durante tiempos de lucha con el pecado o el sufrimiento? 

(Adaptado de Nuestra Suficiencia en Cristo) 

Central Asia is improving food safety measures to share with the world some of the more than 8000 plant species, as well as livestock, from the region.

Title: liothyronine sodium, Cytomel, Triostat
Category: Medications
Created: 12/31/1997 12:00:00 AM
Last Editorial Review: 7/31/2019 12:00:00 AM

Title: Endometriosis Risk Can Be Predicted in Young Girls: Study
Category: Health News
Created: 3/10/2020 12:00:00 AM
Last Editorial Review: 3/10/2020 12:00:00 AM

ABSTRACT

The luminous marine Gram-negative bacterium Vibrio (Aliivibrio) fischeri is the natural light organ symbiont of several squid species, including the Hawaiian bobtail squid, Euprymna scolopes, and the Japanese bobtail squid, Euprymna morsei. Work with E. scolopes has shown how the bacteria establish their niche in the light organ of the newly hatched host. Two types of V. fischeri strains have been distinguished based upon their behavior in cocolonization competition assays in juvenile E. scolopes, i.e., (i) niche-sharing or (ii) niche-dominant behavior. This study aimed to determine whether these behaviors are observed with other V. fischeri strains or whether they are specific to those isolated from E. scolopes light organs. Cocolonization competition assays between V. fischeri strains isolated from the congeneric squid E. morsei or from other marine animals revealed the same sharing or dominant behaviors. In addition, whole-genome sequencing of these strains showed that the dominant behavior is polyphyletic and not associated with the presence or absence of a single gene or genes. Comparative genomics of 44 squid light organ isolates from around the globe led to the identification of symbiosis-specific candidates in the genomes of these strains. Colonization assays using genetic derivatives with deletions of these candidates established the importance of two such genes in colonization. This study has allowed us to expand the concept of distinct colonization behaviors to strains isolated from a number of squid and fish hosts.

IMPORTANCE There is an increasing recognition of the importance of strain differences in the ecology of a symbiotic bacterial species and, in particular, how these differences underlie crucial interactions with their host. Nevertheless, little is known about the genetic bases for these differences, how they manifest themselves in specific behaviors, and their distribution among symbionts of different host species. In this study, we sequenced the genomes of Vibrio fischeri isolated from the tissues of squids and fishes and applied comparative genomics approaches to look for patterns between symbiont lineages and host colonization behavior. In addition, we identified the only two genes that were exclusively present in all V. fischeri strains isolated from the light organs of sepiolid squid species. Mutational studies of these genes indicated that they both played a role in colonization of the squid light organ, emphasizing the value of applying a comparative genomics approach in the study of symbioses.

ABSTRACT

The cell cycle is a critical component of cellular proliferation, differentiation, and response to stress, yet its role in the regulation of intracellular symbioses is not well understood. To explore host-symbiont cell cycle coordination in a marine symbiosis, we employed a model for coral-dinoflagellate associations: the tropical sea anemone Aiptasia (Exaiptasia pallida) and its native microalgal photosymbionts (Breviolum minutum and Breviolum psygmophilum). Using fluorescent labeling and spatial point-pattern image analyses to characterize cell population distributions in both partners, we developed protocols that are tailored to the three-dimensional cellular landscape of a symbiotic sea anemone tentacle. Introducing cultured symbiont cells to symbiont-free adult hosts increased overall host cell proliferation rates. The acceleration occurred predominantly in the symbiont-containing gastrodermis near clusters of symbionts but was also observed in symbiont-free epidermal tissue layers, indicating that the presence of symbionts contributes to elevated proliferation rates in the entire host during colonization. Symbiont cell cycle progression differed between cultured algae and those residing within hosts; the endosymbiotic state resulted in increased S-phase but decreased G₂/M-phase symbiont populations. These phenotypes and the deceleration of cell cycle progression varied with symbiont identity and host nutritional status. These results demonstrate that host and symbiont cells have substantial and species-specific effects on the proliferation rates of their mutualistic partners. This is the first empirical evidence to support species-specific regulation of the symbiont cell cycle within a single cnidarian-dinoflagellate association; similar regulatory mechanisms likely govern interpartner coordination in other coral-algal symbioses and shape their ecophysiological responses to a changing climate.

IMPORTANCE Biomass regulation is critical to the overall health of cnidarian-dinoflagellate symbioses. Despite the central role of the cell cycle in the growth and proliferation of cnidarian host cells and dinoflagellate symbionts, there are few studies that have examined the potential for host-symbiont coregulation. This study provides evidence for the acceleration of host cell proliferation when in local proximity to clusters of symbionts within cnidarian tentacles. The findings suggest that symbionts augment the cell cycle of not only their enveloping host cells but also neighboring cells in the epidermis and gastrodermis. This provides a possible mechanism for rapid colonization of cnidarian tissues. In addition, the cell cycles of symbionts differed depending on nutritional regime, symbiotic state, and species identity. The responses of cell cycle profiles to these different factors implicate a role for species-specific regulation of symbiont cell cycles within host cnidarian tissues.

ABSTRACT

OleA, a member of the thiolase superfamily, is known to catalyze the Claisen condensation of long-chain acyl coenzyme A (acyl-CoA) substrates, initiating metabolic pathways in bacteria for the production of membrane lipids and β-lactone natural products. OleA homologs are found in diverse bacterial phyla, but to date, only one homodimeric OleA has been successfully purified to homogeneity and characterized in vitro. A major impediment for the identification of new OleA enzymes has been protein instability and time-consuming in vitro assays. Here, we developed a bioinformatic pipeline to identify OleA homologs and a new rapid assay to screen OleA enzyme activity in vivo and map their taxonomic diversity. The screen is based on the discovery that OleA displayed surprisingly high rates of p-nitrophenyl ester hydrolysis, an activity not shared by other thiolases, including FabH. The high rates allowed activity to be determined in vitro and with heterologously expressed OleA in vivo via the release of the yellow p-nitrophenol product. Seventy-four putative oleA genes identified in the genomes of diverse bacteria were heterologously expressed in Escherichia coli, and 25 showed activity with p-nitrophenyl esters. The OleA proteins tested were encoded in variable genomic contexts from seven different phyla and are predicted to function in distinct membrane lipid and β-lactone natural product metabolic pathways. This study highlights the diversity of unstudied OleA proteins and presents a rapid method for their identification and characterization.

IMPORTANCE Microbially produced β-lactones are found in antibiotic, antitumor, and antiobesity drugs. Long-chain olefinic membrane hydrocarbons have potential utility as fuels and specialty chemicals. The metabolic pathway to both end products share bacterial enzymes denoted as OleA, OleC, and OleD that transform acyl-CoA cellular intermediates into β-lactones. Bacteria producing membrane hydrocarbons via the Ole pathway additionally express a β-lactone decarboxylase, OleB. Both β-lactone and olefin biosynthesis pathways are initiated by OleA enzymes that define the overall structure of the final product. There is currently very limited information on OleA enzymes apart from the single representative from Xanthomonas campestris. In this study, bioinformatic analysis identified hundreds of new, putative OleA proteins, 74 proteins were screened via a rapid whole-cell method, leading to the identification of 25 stably expressed OleA proteins representing seven bacteria phyla.

ABSTRACT

The Candida albicans high-affinity phosphate transporter Pho84 is required for normal Target of Rapamycin (TOR) signaling, oxidative stress resistance, and virulence of this fungal pathogen. It also contributes to C. albicans’ tolerance of two antifungal drug classes, polyenes and echinocandins. Echinocandins inhibit biosynthesis of a major cell wall component, beta-1,3-glucan. Cells lacking Pho84 were hypersensitive to other forms of cell wall stress beyond echinocandin exposure, while their cell wall integrity signaling response was weak. Metabolomics experiments showed that levels of phosphoric intermediates, including nucleotides like ATP and nucleotide sugars, were low in pho84 mutant compared to wild-type cells recovering from phosphate starvation. Nonphosphoric precursors like nucleobases and nucleosides were elevated. Outer cell wall phosphomannan biosynthesis requires a nucleotide sugar, GDP-mannose. The nucleotide sugar UDP-glucose is the substrate of enzymes that synthesize two major structural cell wall polysaccharides, beta-1,3- and beta-1,6-glucan. Another nucleotide sugar, UDP-N-acetylglucosamine, is the substrate of chitin synthases which produce a stabilizing component of the intercellular septum and of lateral cell walls. Lack of Pho84 activity, and phosphate starvation, potentiated pharmacological or genetic perturbation of these enzymes. We posit that low substrate concentrations of beta-d-glucan- and chitin synthases, together with pharmacologic inhibition of their activity, diminish enzymatic reaction rates as well as the yield of their cell wall-stabilizing products. Phosphate import is not conserved between fungal and human cells, and humans do not synthesize beta-d-glucans or chitin. Hence, inhibiting these processes simultaneously could yield potent antifungal effects with low toxicity to humans.

IMPORTANCE Candida species cause hundreds of thousands of invasive infections with high mortality each year. Developing novel antifungal agents is challenging due to the many similarities between fungal and human cells. Maintaining phosphate balance is essential for all organisms but is achieved completely differently by fungi and humans. A protein that imports phosphate into fungal cells, Pho84, is not present in humans and is required for normal cell wall stress resistance and cell wall integrity signaling in C. albicans. Nucleotide sugars, which are phosphate-containing building block molecules for construction of the cell wall, are diminished in cells lacking Pho84. Cell wall-constructing enzymes may be slowed by lack of these building blocks, in addition to being inhibited by drugs. Combined targeting of Pho84 and cell wall-constructing enzymes may provide a strategy for antifungal therapy by which two sequential steps of cell wall maintenance are blocked for greater potency.

A number of partial articulated specimens of Cheiracanthus peachi nov. sp. have been collected from the Mey Flagstone Formation and Rousay Flagstone Formation within the Orcadian Basin of northern Scotland. The new, robust-bodied species is mainly distinguished by the scale ornament of radiating grooves rather than ridges. Compared to other Cheiracanthus species in the Orcadian Basin, C. peachi nov. sp. has quite a short range making it a useful zone fossil. As well as describing the general morphology of the specimens, we have also described and figured SEM images of scales and histological sections of all elements, enabling identification of other, isolated remains. Of particular biological interest is the identification of relatively robust, tooth-like gill rakers. Finally, the species has also been identified from isolated scales in Belarus, where it appears earlier and has a longer stratigraphical range, implying the species evolved in the marine deposits of the east and migrated west into the Orcadian Basin via the river systems.

The chloroplast glutamyl-tRNA (tRNA^Glu) is unique in that it has two entirely different functions. In addition to acting in translation, it serves as the substrate of glutamyl-tRNA reductase (GluTR), the enzyme catalyzing the committed step in the tetrapyrrole biosynthetic pathway. How the tRNA^Glu pool is distributed between the two pathways and whether tRNA^Glu allocation limits tetrapyrrole biosynthesis and/or protein biosynthesis remains poorly understood. We generated a series of transplastomic tobacco (Nicotiana tabacum) plants to alter tRNA^Glu expression levels and introduced a point mutation into the plastid trnE gene, which has been reported to uncouple protein biosynthesis from tetrapyrrole biosynthesis in chloroplasts of the protist Euglena gracilis. We show that, rather than comparable uncoupling of the two pathways, the trnE mutation is lethal in tobacco because it inhibits tRNA processing, thus preventing translation of Glu codons. Ectopic expression of the mutated trnE gene uncovered an unexpected inhibition of glutamyl-tRNA reductase by immature tRNA^Glu. We further demonstrate that whereas overexpression of tRNA^Glu does not affect tetrapyrrole biosynthesis, reduction of GluTR activity through inhibition by tRNA^Glu precursors causes tetrapyrrole synthesis to become limiting in early plant development when active photosystem biogenesis provokes a high demand for de novo chlorophyll biosynthesis. Taken together, our findings provide insight into the roles of tRNA^Glu at the intersection of protein biosynthesis and tetrapyrrole biosynthesis.

The lignin biosynthetic pathway is highly conserved in angiosperms, yet pathway manipulations give rise to a variety of taxon-specific outcomes. Knockout of lignin-associated 4-coumarate:CoA ligases (4CLs) in herbaceous species mainly reduces guaiacyl (G) lignin and enhances cell wall saccharification. Here we show that CRISPR-knockout of 4CL1 in poplar (Populus tremula x alba) preferentially reduced syringyl (S) lignin, with negligible effects on biomass recalcitrance. Concordant with reduced S-lignin was downregulation of ferulate 5-hydroxylases (F5Hs). Lignification was largely sustained by 4CL5, a low-affinity paralog of 4CL1 typically with only minor xylem expression or activity. Levels of caffeate, the preferred substrate of 4CL5, increased in line with significant upregulation of caffeoyl shikimate esterase1. Upregulation of caffeoyl-CoA O-methyltransferase1 and downregulation of F5Hs are consistent with preferential funneling of 4CL5 products toward G-lignin biosynthesis at the expense of S-lignin. Thus, transcriptional and metabolic adaptations to 4CL1-knockout appear to have enabled 4CL5 catalysis at a level sufficient to sustain lignification. Finally, genes involved in sulfur assimilation, the glutathione-ascorbate cycle, and various antioxidant systems were upregulated in the mutants, suggesting cascading responses to perturbed thioesterification in lignin biosynthesis.

Despite the ecological relevance of diatoms, many aspects of their photosynthetic machinery remain poorly understood. Diatoms differ from the green lineage of oxygenic organisms by their photosynthetic pigments and light-harvesting complex (Lhc) proteins, the latter of which are also called fucoxanthin-chlorophyll proteins (FCP). These are composed of three groups of proteins: Lhcf as the main group, Lhcr that are PSI associated, and Lhcx that are involved in photoprotection. The FCP complexes are assembled in trimers and higher oligomers. Several studies have investigated the biochemical properties of purified FCP complexes, but limited knowledge is available about their interaction with the photosystem cores. In this study, isolation of stable supercomplexes from the centric diatom Thalassiosira pseudonana was achieved. To preserve in vivo structure, the separation of thylakoid complexes was performed by native PAGE and sucrose density centrifugation. Different subpopulations of PSI and PSII supercomplexes were isolated and their subunits identified. Analysis of Lhc antenna composition identified Lhc(s) specific for either PSI (Lhcr 1, 3, 4, 7, 10–14, and Lhcf10) or PSII (Lhcf 1–7, 11, and Lhcr2). Lhcx6_1 was reproducibly found in PSII supercomplexes, whereas its association with PSI was unclear. No evidence was found for the interaction between photosystems and higher oligomeric FCPs, comprising Lhcf8 as the main component. Although the subunit composition of the PSII supercomplexes in comparison with that of the trimeric FCP complexes indicated a close mutual association, the higher oligomeric pool is only weakly associated with the photosystems, albeit its abundance in the thylakoid membrane.

Background

Chronic lung disease of prematurity (CLD), also called bronchopulmonary dysplasia, is a major consequence of preterm birth, but the role of the microbiome in its development remains unclear. Therefore, we assessed the progression of the bacterial community in ventilated preterm infants over time in the upper and lower airways, and assessed the gut–lung axis by comparing bacterial communities in the upper and lower airways with stool findings. Finally, we assessed whether the bacterial communities were associated with lung inflammation to suggest dysbiosis.

Staphylococcus aureus is a noted human and animal pathogen. Despite decades of research on this important bacterium, there are still many unanswered questions regarding the pathogenic mechanisms it uses to infect the mammalian host. This can be attributed to it possessing a plethora of virulence factors and complex virulence factor and metabolic regulation. PurR, the purine biosynthesis regulator, was recently also shown to regulate virulence factors in S. aureus, and mutations in purR result in derepression of fibronectin binding proteins (FnBPs) and extracellular toxins, required for a so-called hypervirulent phenotype. Here, we show that hypervirulent strains containing purR mutations can be attenuated with the addition of purine biosynthesis mutations, implicating the necessity for de novo purine biosynthesis in this phenotype and indicating that S. aureus in the mammalian host experiences purine limitation. Using cell culture, we showed that while purR mutants are not altered in epithelial cell binding, compared to that of wild-type (WT) S. aureus, purR mutants have enhanced invasion of these nonprofessional phagocytes, consistent with the requirement of FnBPs for invasion of these cells. This correlates with purR mutants having increased transcription of fnb genes, resulting in higher levels of surface-exposed FnBPs to promote invasion. These data provide important contributions to our understanding of how the pathogenesis of S. aureus is affected by sensing of purine levels during infection of the mammalian host.

Legumes tend to be nodulated by competitive rhizobia that do not maximize nitrogen (N2) fixation, resulting in suboptimal yields. Rhizobial nodulation competitiveness and effectiveness at N2 fixation are independent traits, making their measurement extremely time-consuming with low experimental throughput. To transform the experimental assessment of rhizobial competitiveness and effectiveness, we...

Background

Nephropathic cystinosis, a hereditary lysosomal storage disorder caused by dysfunction of the lysosomal cotransporter cystinosin, leads to cystine accumulation and cellular damage in various organs, particularly in the kidney. Close therapeutic monitoring of cysteamine, the only available disease-modifying treatment, is recommended. White blood cell cystine concentration is the current gold standard for therapeutic monitoring, but the assay is technically demanding and is available only on a limited basis. Because macrophage-mediated inflammation plays an important role in the pathogenesis of cystinosis, biomarkers of macrophage activation could have potential for the therapeutic monitoring of cystinosis.

Multidrug resistance (MDR) in cancer arises from cross-resistance to structurally- and functionally-divergent chemotherapeutic drugs. In particular, MDR is characterized by increased expression and activity of ATP-binding cassette (ABC) superfamily transporters. Sphingolipids are substrates of ABC proteins in cell signaling, membrane biosynthesis, and inflammation, for example, and their products can favor cancer progression. Glucosylceramide (GlcCer) is a ubiquitous glycosphingolipid (GSL) generated by glucosylceramide synthase, a key regulatory enzyme encoded by the UDP-glucose ceramide glucosyltransferase (UGCG) gene. Stressed cells increase de novo biosynthesis of ceramides, which return to sub-toxic levels after UGCG mediates incorporation into GlcCer. Given that cancer cells seem to mobilize UGCG and have increased GSL content for ceramide clearance, which ultimately contributes to chemotherapy failure, here we investigated how inhibition of GSL biosynthesis affects the MDR phenotype of chronic myeloid leukemias. We found that MDR is associated with higher UGCG expression and with a complex GSL profile. UGCG inhibition with the ceramide analog d-threo-1-(3,4,-ethylenedioxy)phenyl-2-palmitoylamino-3-pyrrolidino-1-propanol (EtDO-P4) greatly reduced GSL and monosialotetrahexosylganglioside levels, and co-treatment with standard chemotherapeutics sensitized cells to mitochondrial membrane potential loss and apoptosis. ABC subfamily B member 1 (ABCB1) expression was reduced, and ABCC-mediated efflux activity was modulated by competition with nonglycosylated ceramides. Consistently, inhibition of ABCC-mediated transport reduced the efflux of exogenous C6-ceramide. Overall, UGCG inhibition impaired the malignant glycophenotype of MDR leukemias, which typically overcomes drug resistance through distinct mechanisms. This work sheds light on the involvement of GSL in chemotherapy failure, and its findings suggest that targeted GSL modulation could help manage MDR leukemias.

Endorepellin, the C-terminal fragment of the heparan sulfate proteoglycan perlecan, influences various signaling pathways in endothelial cells by binding to VEGFR2. In this study, we discovered that soluble endorepellin activates the canonical stress signaling pathway consisting of PERK, eIF2α, ATF4, and GADD45α. Specifically, endorepellin evoked transient activation of VEGFR2, which, in turn, phosphorylated PERK at Thr980. Subsequently, PERK phosphorylated eIF2α at Ser51, upregulating its downstream effector proteins ATF4 and GADD45α. RNAi-mediated knockdown of PERK or eIF2α abrogated the endorepellin-mediated up-regulation of GADD45α, the ultimate effector protein of this stress signaling cascade. To functionally validate these findings, we utilized an ex vivo model of angiogenesis. Exposure of the aortic rings embedded in 3D fibrillar collagen to recombinant endorepellin for 2–4 h activated PERK and induced GADD45α vis à vis vehicle-treated counterparts. Similar effects were obtained with the established cellular stress inducer tunicamycin. Notably, chronic exposure of aortic rings to endorepellin for 7–9 days markedly suppressed vessel sprouting, an angiostatic effect that was rescued by blocking PERK kinase activity. Our findings unravel a mechanism by which an extracellular matrix protein evokes stress signaling in endothelial cells, which leads to angiostasis.

Allostery exploits the conformational dynamics of enzymes by triggering a shift in population ensembles toward functionally distinct conformational or dynamic states. Allostery extensively regulates the activities of key enzymes within biosynthetic pathways to meet metabolic demand for their end products. Here, we have examined a critical enzyme, 3-deoxy-d-arabino-heptulosonate 7-phosphate synthase (DAH7PS), at the gateway to aromatic amino acid biosynthesis in Mycobacterium tuberculosis, which shows extremely complex dynamic allostery: three distinct aromatic amino acids jointly communicate occupancy to the active site via subtle changes in dynamics, enabling exquisite fine-tuning of delivery of these essential metabolites. Furthermore, this allosteric mechanism is co-opted by pathway branchpoint enzyme chorismate mutase upon complex formation. In this study, using statistical coupling analysis, site-directed mutagenesis, isothermal calorimetry, small-angle X-ray scattering, and X-ray crystallography analyses, we have pinpointed a critical node within the complex dynamic communication network responsible for this sophisticated allosteric machinery. Through a facile Gly to Pro substitution, we have altered backbone dynamics, completely severing the allosteric signal yet remarkably, generating a nonallosteric enzyme that retains full catalytic activity. We also identified a second residue of prime importance to the inter-enzyme communication with chorismate mutase. Our results reveal that highly complex dynamic allostery is surprisingly vulnerable and provide further insights into the intimate link between catalysis and allostery.

Decreased release of palmitic acid methyl ester (PAME), a vasodilator, from perivascular adipose tissue (PVAT) might contribute to hypertension pathogenesis. However, the PAME biosynthetic pathway remains unclear. In this study, we hypothesized that PAME is biosynthesized from palmitic acid (PA) via human catechol-O-methyltransferase (COMT) catalysis and that decreased PAME biosynthesis plays a role in hypertension pathogenesis. We compared PAME biosynthesis between age-matched normotensive Wistar Kyoto (WKY) rats and hypertensive spontaneously hypertensive rats (SHRs) and investigated the effects of losartan treatment on PAME biosynthesis. Computational molecular modeling indicated that PA binds well at the active site of COMT. Furthermore, in in vitro enzymatic assays in the presence of COMT and S-5'-adenosyl-L-methionine (AdoMet), the stable isotope [¹³C₁₆]-PA was methylated to form [¹³C₁₆]-PAME in incubation medium or the Krebs–Henseleit solution containing 3T3-L1 adipocytes or rat PVAT. The adipocytes and PVATs expressed membrane-bound (MB)-COMT and soluble (S)-COMT proteins. [¹³C₁₆]-PA methylation to form [¹³C₁₆]-PAME in 3T3-L1 adipocytes and rat PVAT was blocked by various COMT inhibitors, such as S-(5'-adenosyl)-L-homocysteine, adenosine-2',3'-dialdehyde, and tolcapone. MB- and S-COMT levels in PVATs of established SHRs were significantly lower than those in PVATs of age-matched normotensive WKY rats, with decreased [¹³C₁₆]-PA methylation to form [¹³C₁₆]-PAME. This decrease was reversed by losartan, an angiotensin II (Ang II) type 1 receptor antagonist. Therefore, PAME biosynthesis in rat PVAT is dependent on AdoMet, catalyzed by COMT, and decreased in SHRs, further supporting the role of PVAT/PAME in hypertension pathogenesis. Moreover, the antihypertensive effect of losartan might be due partly to its increased PAME biosynthesis.

Chitotriosidase (CHIT1) is a highly conserved and regulated chitinase secreted by activated macrophages; it is a member of the 18-glycosylase family (GH18). CHIT1 is the most prominent chitinase in humans, can cleave chitin and participates in the body's immune response and is associated with inflammation, infection, tissue damage and remodelling processes. Recently, CHIT1 has been reported to be involved in the molecular pathogenesis of pulmonary fibrosis, bronchial asthma, COPD and pulmonary infections, shedding new light on the role of these proteins in lung pathophysiology. The potential roles of CHIT1 in lung diseases are reviewed in this article.

Assemblages of upper lower through upper Miocene Discoaster spp. have been quantified from Integrated Ocean Drilling Program (IODP) Site U1338 in the eastern equatorial Pacific Ocean. These assemblages can be grouped into five broad morphological categories: six-rayed with bifurcated ray tips, six-rayed with large central areas, six-rayed with pointed ray tips, five-rayed with bifurcated ray tips and five-rayed with pointed ray tips. Discoaster deflandrei dominates the assemblages prior to 15.8 Ma. The decline in abundance of D. deflandrei close to the early–middle Miocene boundary occurs together with the evolution of the D. variabilis group, including D. signus and D. exilis. Six-rayed discoasters having large central areas become a prominent member of the assemblages for a 400 ka interval in the late middle Miocene. Five- and six-rayed forms having pointed tips become prominent in the early late Miocene and show a strong antiphasing relationship with the D. variabilis group. Discoaster bellus completely dominates the Discoaster assemblages for a 400 ka interval in the middle late Miocene. Abundances of all discoasters, or discoasters at the species level, show only (surprisingly) weak correlations to carbonate contents or oxygen and carbon isotopes of bulk sediment when calculated over the entire sample interval.

Vibralactone, a hybrid compound derived from phenols and a prenyl group, is a strong pancreatic lipase inhibitor with a rare fused bicyclic β-lactone skeleton. Recently, a researcher reported a vibralactone derivative (compound C1) that caused inhibition of pancreatic lipase with a half-maximal inhibitory concentration of 14 nM determined by structure-based optimization, suggesting a potential candidate as a new antiobesity treatment. In the present study, we sought to identify the main gene encoding prenyltransferase in Stereum vibrans, which is responsible for the prenylation of phenol leading to vibralactone synthesis. Two RNA silencing transformants of the identified gene (vib-PT) were obtained through Agrobacterium tumefaciens-mediated transformation. Compared to wild-type strains, the transformants showed a decrease in vib-PT expression ranging from 11.0 to 56.0% at 5, 10, and 15 days in reverse transcription-quantitative PCR analysis, along with a reduction in primary vibralactone production of 37 to 64% at 15 and 21 days, respectively, as determined using ultra-high-performance liquid chromatography-mass spectrometry analysis. A soluble and enzymatically active fusion Vib-PT protein was obtained by expressing vib-PT in Escherichia coli, and the enzyme’s optimal reaction conditions and catalytic efficiency (K_m/k_cat) were determined. In vitro experiments established that Vib-PT catalyzed the C-prenylation at C-3 of 4-hydroxy-benzaldehyde and the O-prenylation at the 4-hydroxy of 4-hydroxy-benzenemethanol in the presence of dimethylallyl diphosphate. Moreover, Vib-PT shows promiscuity toward aromatic compounds and prenyl donors.

IMPORTANCE Vibralactone is a lead compound with a novel skeleton structure that shows strong inhibitory activity against pancreatic lipase. Vibralactone is not encoded by the genome directly but rather is synthesized from phenol, followed by prenylation and other enzyme reactions. Here, we used an RNA silencing approach to identify and characterize a prenyltransferase in a basidiomycete species that is responsible for the synthesis of vibralactone. The identified gene, vib-PT, was expressed in Escherichia coli to obtain a soluble and enzymatically active fusion Vib-PT protein. In vitro characterization of the enzyme demonstrated the catalytic mechanism of prenylation and broad substrate range for different aromatic acceptors and prenyl donors. These characteristics highlight the possibility of Vib-PT to generate prenylated derivatives of aromatics and other compounds as improved bioactive agents or potential prodrugs.

In Lysobacter enzymogenes OH11, RpfB1 and RpfB2 were predicted to encode acyl coenzyme A (CoA) ligases. RpfB1 is located in the Rpf gene cluster. Interestingly, we found an RpfB1 homolog (RpfB2) outside this canonical gene cluster, and nothing is known about its functionality or mechanism. Here, we report that rpfB1 and rpfB2 can functionally replace EcFadD in the Escherichia coli fadD mutant JW1794. RpfB activates long-chain fatty acids (n-C_16:0 and n-C_18:0) for the corresponding fatty acyl-CoA ligase (FCL) activity in vitro, and Glu-361 plays critical roles in the catalytic mechanism of RpfB1 and RpfB2. Deletion of rpfB1 and rpfB2 resulted in significantly increased heat-stable antifungal factor (HSAF) production, and overexpression of rpfB1 or rpfB2 completely suppressed HSAF production. Deletion of rpfB1 and rpfB2 resulted in increased L. enzymogenes diffusible signaling factor 3 (LeDSF3) synthesis in L. enzymogenes. Overall, our results showed that changes in intracellular free fatty acid levels significantly altered HSAF production. Our report shows that intracellular free fatty acids are required for HSAF production and that RpfB affects HSAF production via FCL activity. The global transcriptional regulator Clp directly regulated the expression of rpfB1 and rpfB2. In conclusion, these findings reveal new roles of RpfB in antibiotic biosynthesis in L. enzymogenes.

IMPORTANCE Understanding the biosynthetic and regulatory mechanisms of heat-stable antifungal factor (HSAF) could improve the yield in Lysobacter enzymogenes. Here, we report that RpfB1 and RpfB2 encode acyl coenzyme A (CoA) ligases. Our research shows that RpfB1 and RpfB2 affect free fatty acid metabolism via fatty acyl-CoA ligase (FCL) activity to reduce the substrate for HSAF synthesis and, thereby, block HSAF production in L. enzymogenes. Furthermore, these findings reveal new roles for the fatty acyl-CoA ligases RpfB1 and RpfB2 in antibiotic biosynthesis in L. enzymogenes. Importantly, the novelty of this work is the finding that RpfB2 lies outside the Rpf gene cluster and plays a key role in HSAF production, which has not been reported in other diffusible signaling factor (DSF)/Rpf-producing bacteria.

The response to iron limitation of the Gram-positive soil bacterium Corynebacterium glutamicum was analyzed with respect to secreted metabolites, the transcriptome, and the proteome. During growth in glucose minimal medium, iron limitation caused a shift from lactate to pyruvate as the major secreted organic acid complemented by l-alanine and 2-oxoglutarate. Transcriptome and proteome analyses revealed that a pronounced iron starvation response governed by the transcriptional regulators DtxR and RipA was detectable in the late, but not in the early, exponential-growth phase. A link between iron starvation and thiamine pyrophosphate (TPP) biosynthesis was uncovered by the strong upregulation of thiC. As phosphomethylpyrimidine synthase (ThiC) contains an iron-sulfur cluster, limiting activities of the TPP-dependent pyruvate–2-oxoglutarate dehydrogenase supercomplex probably cause the excretion of pyruvate and 2-oxoglutarate. In line with this explanation, thiamine supplementation could strongly diminish the secretion of these acids. The upregulation of thiC and other genes involved in thiamine biosynthesis and transport is presumably due to TPP riboswitches present at the 5' end of the corresponding operons. The results obtained in this study provide new insights into iron homeostasis in C. glutamicum and demonstrate that the metabolic consequences of iron limitation can be due to the iron dependency of coenzyme biosynthesis.

IMPORTANCE Iron is an essential element for most organisms but causes problems due to poor solubility under oxic conditions and due to toxicity by catalyzing the formation of reactive oxygen species (ROS). Therefore, bacteria have evolved complex regulatory networks for iron homeostasis aiming at a sufficient iron supply while minimizing ROS formation. In our study, the responses of the actinobacterium Corynebacterium glutamicum to iron limitation were analyzed, resulting in a detailed view on the processes involved in iron homeostasis in this model organism. In particular, we provide evidence that iron limitation causes TPP deficiency, presumably due to insufficient activity of the iron-dependent phosphomethylpyrimidine synthase (ThiC). TPP deficiency was deduced from the upregulation of genes controlled by a TPP riboswitch and secretion of metabolites caused by insufficient activity of the TPP-dependent enzymes pyruvate dehydrogenase and 2-oxoglutarate dehydrogenase. To our knowledge, the link between iron starvation and thiamine synthesis has not been elaborated previously.

Antiretroviral therapy (ART) suppresses HIV-1 replication but fails to cure the infection. The presence of an extremely stable viral latent reservoir, primarily in resting memory CD4⁺ T cells, remains a major obstacle to viral eradication. The "shock and kill" strategy targets these latently infected cells and boosts immune recognition and clearance, and thus, it is a promising approach for an HIV-1 functional cure. Although some latency-reversing agents (LRAs) have been reported, no apparent clinical progress has been made, so it is still vital to seek novel and effective LRAs. Here, we report that thiostrepton (TSR), a proteasome inhibitor, reactivates latent HIV-1 effectively in cellular models and in primary CD4⁺ T cells from ART-suppressed individuals ex vivo. TSR does not induce global T cell activation, severe cytotoxicity, or CD8⁺ T cell dysfunction, making it a prospective LRA candidate. We also observed a significant synergistic effect of reactivation when TSR was combined with JQ1, prostratin, or bryostatin-1. Interestingly, six TSR analogues also show reactivation abilities that are similar to or more effective than that of TSR. We further verified that TSR upregulated expression of heat shock proteins (HSPs) in CD4⁺ T cells, which subsequently activated positive transcriptional elongation factor b (p-TEFb) and NF-B signals, leading to viral reactivation. In summary, we identify TSR as a novel LRA which could have important significance for applications to an HIV-1 functional cure in the future.

Advances in synthetic biology have enabled the production of a variety of compounds using bacteria as a vehicle for complex compound biosynthesis. Violacein, a naturally occurring indole pigment with antibiotic properties, can be biosynthetically engineered in Escherichia coli expressing its nonnative synthesis pathway. To explore whether this synthetic biosynthesis platform could be used for drug discovery, here we have screened bacterially derived violacein against the main causative agent of human malaria, Plasmodium falciparum. We show the antiparasitic activity of bacterially derived violacein against the P. falciparum 3D7 laboratory reference strain as well as drug-sensitive and -resistant patient isolates, confirming the potential utility of this drug as an antimalarial agent. We then screen a biosynthetic series of violacein derivatives against P. falciparum growth. The varied activity of each derivative against asexual parasite growth points to the need to further develop violacein as an antimalarial. Towards defining its mode of action, we show that biosynthetic violacein affects the parasite actin cytoskeleton, resulting in an accumulation of actin signal that is independent of actin polymerization. This activity points to a target that modulates actin behavior in the cell either in terms of its regulation or its folding. More broadly, our data show that bacterial synthetic biosynthesis could become a suitable platform for antimalarial drug discovery, with potential applications in future high-throughput drug screening with otherwise chemically intractable natural products.