The bHLH transcription factors TSAR1 and TSAR2 regulate triterpene saponin biosynthesis in Medicago truncatula

Plants respond to stresses by producing a broad spectrum of bioactive specialized metabolites. Hormonal elicitors, such as jasmonates, trigger a complex signaling circuit leading to the concerted activation of specific metabolic pathways. However, for many specialized metabolic pathways, the transcription factors involved remain unknown. Here, we report on two homologous jasmonate-inducible transcription factors of the basic helix-loop-helix family, TRITERPENE SAPONIN BIOSYNTHESIS ACTIVATING REGULATOR1 (TSAR1) and TSAR2, which direct triterpene saponin biosynthesis in Medicago truncatula. TSAR1 and TSAR2 are coregulated with and transactivate the genes encoding 3-HYDROXY-3-METHYLGLUTARYL-COENZYME A REDUCTASE1 (HMGR1) and MAKIBISHI1, the rate-limiting enzyme for triterpene biosynthesis and an E3 ubiquitin ligase that controls HMGR1 levels, respectively. Transactivation is mediated by direct binding of TSARs to the N-box in the promoter of HMGR1. In transient expression assays in tobacco (Nicotiana tabacum) protoplasts, TSAR1 and TSAR2 exhibit different patterns of transactivation of downstream triterpene saponin biosynthetic genes, hinting at distinct functionalities within the regulation of the pathway. Correspondingly, overexpression of TSAR1 or TSAR2 in M. truncatula hairy roots resulted in elevated transcript levels of known triterpene saponin biosynthetic genes and strongly increased the accumulation of triterpene saponins. TSAR2 overexpression specifically boosted hemolytic saponin biosynthesis, whereas TSAR1 overexpression primarily stimulated nonhemolytic soyasaponin biosynthesis. Both TSARs also activated all genes of the precursor mevalonate pathway but did not affect sterol biosynthetic genes, pointing to their specific role as regulators of specialized triterpene metabolism in M. truncatula

Genome-wide profiling of uncapped mRNA

Gene transcripts are under extensive posttranscriptional regulation, including the regulation of their stability. A major route for mRNA degradation produces uncapped mRNAs, which can be generated by decapping enzymes, endonucleases, and small RNAs. Profiling uncapped mRNA molecules is important for the understanding of the transcriptome, whose composition is determined by a balance between mRNA synthesis and degradation. In this chapter, we describe a method to profile these uncapped mRNAs at the genome scale

Mimicry technology : a versatile tool for small RNA suppression

A decade ago the discovery of the target mimicry regulatory process on the activity of a mature microRNA (miRNA) enabled for the first time the customized attenuation of miRNA activity in plants. That powerful technology was named MIMIC and was based on engineering the IPS1 long noncoding transcript to become complementary to the miRNA under study. In order to avoid IPS1 degradation, the predicted miRNA-mediated cleavage site was interrupted by three additional nucleotides giving rise to the so-called MIMIC decoy. Since then, MIMIC technology has been used in several plant species and in basic and translational research. We here provide a detailed guide to produce custom-designed MIMIC decoys to facilitate the study of sRNA functions in plants

Efecto Antibacteriano In vitro del extracto Hidroalcohólico de las hojas de Origanum vulgare L. (Orégano) sobre Streptococcus mutans ATCC 25175

Objetivo: Determinar el efecto antibacteriano in vitro, del extracto hidroalcohólico de las hojas de Origanum vulgare L. (orégano) sobre Streptococcus mutans ATCC 25175. Materiales y métodos: Investigación de tipo experimental, explicativo, prospectivo, transversal. La muestra vegetal estuvo constituida por 2 kilos de hojas secas y la muestra biológica usada consistió en 10 placas Petri. El procedimiento fitoquímico fue la marcha fitoquímica y el método microbiológico usado fue el de difusión en agar o Kirby-Bauer el cual requirió del uso de 10 repeticiones y estuvo constituida por los grupos experimentales al 5 %, 25 %, 50 % y 90 %, el grupo control fue Amoxicilina 25 μg y Dimetilsulfóxido (DMSO). Resultados: Los metabolitos secundarios que se detectaron en la prueba del tamizaje fitoquímico fueron los compuestos fenólicos, antocianinas, terpenos y esteroides, alcaloides y lactonas α, β-insaturadas. Por otro lado, mediante ANOVA y Tukey, se demostró como resultado una diferencia estadísticamente significativa entre los grupos experimentales al 25 %, 50 % y 90 % comparado con el control (DMSO) frente a Streptococcus mutans ATCC 25175. Conclusión: El extracto hidroalcohólico de las hojas de Origanum vulgare L. (orégano) presenta actividad antibacteriana frente a cepas de Streptococcus mutans ATCC 25175

Competencias entre fisioterapeutas y educadores físicos-deportivos respecto al ejercicio físico y terapéutico: una revisión narrativa

El ejercicio físico-terapéutico es una reciente herramienta nacida de la fusión del ejercicio físico y la rehabilitación. Debido a su corta edad, no cuenta con un nivel de estructuración definido, originando conflictos entre fisioterapeutas y educadores físico-de- portivos. El objetivo de esta revisión es analizar la bibliografía disponible para buscar unos criterios consensuados. Se realizó una búsqueda en PubMed, utilizando una estrategia basada en la combinación de términos relacionados con el ejercicio físico, el ejercicio terapéutico y su utilización como tratamiento de diversas patologías. Se analizaron un total de 60 programas, de los cuales en tan sólo 4 se especifica el ejercicio terapéutico con respecto al ejercicio físico. Por lo tanto, concluimos que existe una inestabilidad legal en España en cuanto a competencias profesionales, siendo la carga formativa académica universitaria insuficiente para hacer frente a las necesidades de la población general

Characterization of ripe fruit epidermis-specific transcription factors in strawberry

Transcriptome changes during strawberry fruit ripening have been previously reported using either complete fruits or achenes (actual fruits) and receptacles (fleshy part) separately. In order to perform a more detailed study, we have performed a tissue- and stage-specific transcriptome analysis in receptacles of Fragaria vesca fruits, allowing us to infer Gene Regulatory Networks (GRN) in each tissue and stage. In the study, we have focused on the epidermis at the ripe stage, since it plays an important role in defense, as it is the external cell layer in direct contact with the environment, and, in contrast to receptacles of the commercial species, it is the only part of the fruit that accumulates anthocyanins. MapMan analysis of the GRN in ripe epidermis showed that wax and flavonoid biosynthesis were significantly overrepresented functions. Three out of the several TFs found among the main hubs in this GRN were selected to study their biological role, one of them belonging to the MYB family, and two bHLH genes. Protein interaction assays revealed that the MYB protein physically interacts with the two bHLHs, leading to the subcellular relocalization from the cytoplasm to the nucleus in one of them. DAP-seq analyses showed that the bHLH TFs do not bind DNA by themselves, but that genes involved in cuticle formation and flavonoid biosynthesis are among the MYB targets, which were validated by a transactivation assay using the Luciferase/Renilla system. Consistently, MYB-overexpressing stable lines exhibited an upregulation of genes related to cuticle and wax biosynthesis in ripe fruits, and an accumulation of higher amounts of epicuticular waxes in young leaves compared to the WT. We are currently establishing RNAi and CRISPR lines for these three ripe-epidermis specific TFs to further investigate their biological role and performing analyses to understand the effect on gene expression of the interaction between them.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Characterization of the cytokinin-responsive transcriptome in rice

Abstract Background Cytokinin activates transcriptional cascades important for development and the responses to biotic and abiotic stresses. Most of what is known regarding cytokinin-regulated gene expression comes from studies of the dicotyledonous plant Arabidopsis thaliana. To expand the understanding of the cytokinin-regulated transcriptome, we employed RNA-Seq to analyze gene expression in response to cytokinin in roots and shoots of the monocotyledonous plant rice. Results We identified over 4,600 and approximately 2,400 genes differentially expressed in response to cytokinin in roots and shoots respectively. There were some similarities in the sets of cytokinin-regulated genes identified in rice and Arabidopsis, including an up-regulation of genes that act to reduce cytokinin function. Consistent with this, we found that the preferred DNA-binding motif of a rice type-B response regulator is similar to those from Arabidopsis. Analysis of the genes regulated by cytokinin in rice revealed a large number of transcription factors, receptor-like kinases, and genes involved in protein degradation, as well as genes involved in development and the response to biotic stress. Consistent with the over-representation of genes involved in biotic stress, there is a substantial overlap in the genes regulated by cytokinin and those differentially expressed in response to pathogen infection, suggesting that cytokinin plays an integral role in the transcriptional response to pathogens in rice, including the induction of a large number of WRKY transcription factors. Conclusions These results begin to unravel the complex gene regulation after cytokinin perception in a crop of agricultural importance and provide insight into the processes and responses modulated by cytokinin in monocots

Phytochrome interacting factors 4 and 5 control seedling growth in changing light conditions by directly controlling auxin signaling.

Plant growth is strongly influenced by the presence of neighbors that compete for light resources. In response to vegetational shading shade-intolerant plants such as Arabidopsis display a suite of developmental responses known as the shade-avoidance syndrome (SAS). The phytochrome B (phyB) photoreceptor is the major light sensor to mediate this adaptive response. Control of the SAS occurs in part with phyB, which controls protein abundance of phytochrome-interacting factors 4 and 5 (PIF4 and PIF5) directly. The shade-avoidance response also requires rapid biosynthesis of auxin and its transport to promote elongation growth. The identification of genome-wide PIF5-binding sites during shade avoidance revealed that this bHLH transcription factor regulates the expression of a subset of previously identified SAS genes. Moreover our study suggests that PIF4 and PIF5 regulate elongation growth by controlling directly the expression of genes that code for auxin biosynthesis and auxin signaling components

A PHABULOSA/cytokinin feedback loop controls root growth in arabidopsis

The hormone cytokinin (CK) controls root length in Arabidopsis thaliana by defining where dividing cells, derived from stem cells of the root meristem, start to differentiate [ [1], [2], [3], [4], [5] and [6]]. However, the regulatory inputs directing CK to promote differentiation remain poorly understood. Here, we show that the HD-ZIPIII transcription factor PHABULOSA (PHB) directly activates the CK biosynthesis gene ISOPENTENYL TRANSFERASE 7 (IPT7), thus promoting cell differentiation and regulating root length. We further demonstrate that CK feeds back to repress both PHB and microRNA165, a negative regulator of PHB. These interactions comprise an incoherent regulatory loop in which CK represses both its activator and a repressor of its activator. We propose that this regulatory circuit determines the balance of cell division and differentiation during root development and may provide robustness against CK fluctuations