Molecular and genetic characterization of novel S-RNases from a natural population of Nicotiana alata

Self-incompatibility in the Solanaceae is mediated by S-RNase alleles expressed in the style, which confer specificity for pollen recognition. Nicotiana alata has been successfully used as an experimental model to elucidate cellular and molecular aspects of S-RNase-based self-incompatibility in Solanaceae. However, S-RNase alleles of this species have not been surveyed from natural populations and consequently the S-haplotype diversity is poorly known. Here the molecular and functional characterization of seven S-RNase candidate sequences, identified from a natural population of N. alata, are reported. Six of these candidates, S 5 , S 27 , S 70 , S 75 , S 107 , and S 210 , showed plant-specific amplification in the natural population and style-specific expression, which increased gradually during bud maturation, consistent with the reported S-RNase expression. In contrast, the S 63 ribonuclease was present in all plants examined and was ubiquitously expressed in different organs and bud developmental stages. Genetic segregation analysis demonstrated that S 27 , S 70 , S 75 , S 107 , and S 210 alleles were fully functional novel S-RNases, while S 5 and S 63 resulted to be non-S-RNases, although with a clearly distinct pattern of expression. These results reveal the importance of performing functional analysis in studies of S-RNase allelic diversity. Comparative phylogenetic analysis of six species of Solanaceae showed that N. alata S-RNases were included in eight transgeneric S-lineages. Phylogenetic pattern obtained from the inclusion of the novel S-RNase alleles confirms that N. alata represents a broad sample of the allelic variation at the S-locus of the Solanaceae.Fil: Roldán, Juan Alfredo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones en Química Biológica de Córdoba. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Centro de Investigaciones en Química Biológica de Córdoba; ArgentinaFil: Quiroga, Rodrigo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones en Química Biológica de Córdoba. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Centro de Investigaciones en Química Biológica de Córdoba; ArgentinaFil: Goldraij, Ariel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones en Química Biológica de Córdoba. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Centro de Investigaciones en Química Biológica de Córdoba; Argentin

Entre máscara, cuerpo y puesta en escena. : Desenmascarando el nacimiento de una obra

The text thoughtfully explores the progress of meanings rhizomatically expanded throughout the rehearsals of a project, “Opaque and Neutral Masks: from the self-cognitive training to the composition of scenic material”. The aim of this work is to look into the ways ideas and emotions that are part of the play in process were twisted and turned. It exposes the dialogue between the apparent expressive rigidity of mask and incitement on a stage that oscillates between what is to be expected and what is alarming, what is banal and what is spectacular, what is human and what is animal, what is permanent and what is ephemeral. More than a review, in this text we inquire into the creative process of a theatrical event that overflows the standards of stage and verbal languages, the idea of direction as something omnipotent, the script as a complete fact that must be closely followed, the work as a product of a creative genius, someone inspired and in solitude, and the mask as a concealing device. Here, the writing process does not have the purpose of establishing a completed knowledge, being witness of a concluded subjectivity. It follows the trail of an experimentation that finds in its threshold a porous and productive limit into which the meanings that seemed unmutable can be poured. En el texto se busca explorar reflexivamente el desarrollo de algunos sentidos que se expandieron de manera rizomática a lo largo de los ensayos del proyecto “Media Opaca y Máscara Neutra: del entrenamiento autocognitivo a la composición del propio material escénico” para indagar las maneras en que se fueron torciendo y retorciendo las ideas y los sentires que configuran el proceso de obra. Se expone el diálogo entre la aparente rigidez expresiva de la máscara y el estímulo de un escenario que oscila entre lo esperable y lo alarmante, lo banal y lo espectacular, lo humano y lo animal, lo permanente y lo efímero. Más que una reseña de obra, en este texto se indaga en el proceso creativo de un suceso teatral que desborda los estándares del lenguaje escénico y verbal, de la idea de dirección como omnipotencia, del guión como hecho consumado que se sigue al pie de la letra, de un ser genio creativo como persona inspirada y solitaria, de la máscara como dispositivo de ocultamiento. Aquí la escritura no actúa como fijación de un saber acabado, testigo de una subjetividad concluida, sino que sigue el tránsito de una experimentación que encuentra en su umbral un límite poroso y productivo por el cual derramar los sentidos que parecían inmutables.

Hospital care for the dying patient with cancer: does an advance care planning invitation influence bereaved relatives' experiences? A two country survey

Objectives: Advance care planning (ACP) is not systematically performed in Argentina or Norway. We used the post-bereavement survey of the ERANet-LAC International Care Of the Dying Evaluation (CODE) project (2017–2020) to examine the proportion of relatives who were offered an ACP conversation, the proportion of those not offered it who would have wanted it and whether the outcomes differed between those offered a conversation and those not. Methods: Relatives after cancer deaths in hospitals answered the CODE questionnaire 6–8 weeks post bereavement, by post (Norway) or interview (Argentina). Two additional questions asked if the relative and patient had been invited to a conversation about wishes for the patient’s remaining lifetime, and, if not invited, whether they would have wanted such a conversation. The data were analysed using mixed-effects ordinal regression models. Results: 276 participants (Argentina 98 and Norway 178) responded (56% spouses, 31% children, 68% women, age 18–80+). Fifty-six per cent had been invited, and they had significantly more positive perceptions about care and support than those not invited. Sixty-eight per cent of the participants not invited would have wanted an invitation, and they had less favourable perceptions about the care, especially concerning emotional and spiritual support. Conclusions: Relatives who had been invited to a conversation about wishes for the patient’s remaining lifetime had more positive perceptions about patient care and support for the relatives in the patient’s final days of life. A majority of the relatives who had not been invited to an ACP conversation would have wanted it.publishedVersio

Genetic evidence that two independent S-loci control RNase-based self-incompatibility in diploid strawberry

The self-incompatibility mechanism that reduces inbreeding in many plants of the Rosaceae is attributed to a multi-allelic S locus which, in the Prunoideae and Maloideae subfamilies, comprises two complementary genes, a stylar-expressed S-RNase and a pollen-expressed SFB. To elucidate incompatibility in the subfamily Rosoideae, stylar-specific RNases and self-(in)compatibility status were analysed in various diploid strawberries, especially Fragaria nubicola and F. viridis, both self-incompatible, and F. vesca, self-compatible, and in various progenies derived from them. Unexpectedly, two unlinked RNase loci, S and T, were found, encoding peptides distinct from Prunoideae and Maloideae S-RNases; the presence of a single active allele at either is sufficient to confer self-incompatibility. By contrast, in diploid Maloideae and Prunoideae a single locus encodes S-RNases that share several conserved regions and two active alleles are required for self-incompatibility. Our evidence implicates the S locus in unilateral inter-specific incompatibility and shows that S and T RNases can, remarkably, confer not only allele-specific rejection of cognate pollen but also unspecific rejection of Sn Tn pollen, where n indicates a null allele, consistent with the the presence of the pollen component, SFB, activating the cognitive function of these RNases. Comparison of relevant linkage groups between Fragaria and Prunus suggests that Prunus S-RNases, unique in having two introns, may have resulted from gene conversion in an ancestor of Prunus. In addition, it is shown that there is a non-S locus that is essential for self-incompatibility in diploid Fragaria

Hospital care for the dying patient with cancer: does an advance care planning invitation influence bereaved relatives’ experiences? A two country survey

Objectives Advance care planning (ACP) is not systematically performed in Argentina or Norway. We used the post-bereavement survey of the ERANet-LAC International Care Of the Dying Evaluation (CODE) project (2017–2020) to examine the proportion of relatives who were offered an ACP conversation, the proportion of those not offered it who would have wanted it and whether the outcomes differed between those offered a conversation and those not. Methods Relatives after cancer deaths in hospitals answered the CODE questionnaire 6–8 weeks post bereavement, by post (Norway) or interview (Argentina). Two additional questions asked if the relative and patient had been invited to a conversation about wishes for the patient’s remaining lifetime, and, if not invited, whether they would have wanted such a conversation. The data were analysed using mixed-effects ordinal regression models. Results 276 participants (Argentina 98 and Norway 178) responded (56% spouses, 31% children, 68% women, age 18–80+). Fifty-six per cent had been invited, and they had significantly more positive perceptions about care and support than those not invited. Sixty-eight per cent of the participants not invited would have wanted an invitation, and they had less favourable perceptions about the care, especially concerning emotional and spiritual support. Conclusions Relatives who had been invited to a conversation about wishes for the patient’s remaining lifetime had more positive perceptions about patient care and support for the relatives in the patient’s final days of life. A majority of the relatives who had not been invited to an ACP conversation would have wanted it

Mutational analysis of the major soybean UreF paralogue involved in urease activation

The soybean genome duplicated ∼14 and 45 million years ago and has many paralogous genes, including those in urease activation (emplacement of Ni and CO2 in the active site). Activation requires the UreD and UreF proteins, each encoded by two paralogues. UreG, a third essential activation protein, is encoded by the single-copy Eu3, and eu3 mutants lack activity of both urease isozymes. eu2 has the same urease-negative phenotype, consistent with Eu2 being a single-copy gene, possibly encoding a Ni carrier. Unexpectedly, two eu2 alleles co-segregated with missense mutations in the chromosome 2 UreF paralogue (Ch02UreF), suggesting lack of expression/function of Ch14UreF. However, Ch02UreF and Ch14UreF transcripts accumulate at the same level. Further, it had been shown that expression of the Ch14UreF ORF complemented a fungal ureF mutant. A third, nonsense (Q2*) allelic mutant, eu2-c, exhibited 5- to 10-fold more residual urease activity than missense eu2-a or eu2-b, though eu2-c should lack all Ch02UreF protein. It is hypothesized that low-level activation by Ch14UreF is ‘spoiled’ by the altered missense Ch02UreF proteins (‘epistatic dominant-negative’). In agreement with active ‘spoiling’ by eu2-b-encoded Ch02UreF (G31D), eu2-b/eu2-c heterozygotes had less than half the urease activity of eu2-c/eu2-c siblings. Ch02UreF (G31D) could spoil activation by Chr14UreF because of higher affinity for the activation complex, or because Ch02UreF (G31D) is more abundant than Ch14UreF. Here, the latter is favoured, consistent with a reported in-frame AUG in the 5' leader of Chr14UreF transcript. Translational inhibition could represent a form of ‘functional divergence’ of duplicated genes

Identification and functional analysis of pistil self-incompatibility factor HT-B of Petunia

Gametophytic self-incompatibility (GSI) in Solanaceae, Rosaceae, and Plantaginaceae is controlled by a multiallelic S-locus. The specificities of pistil and pollen are controlled by separate S-locus genes, S-RNase and SLF/SFB, respectively. Although the S-specificity is determined by the S-locus genes, factors located outside the S-locus are also required for expression of GSI. HT-B is one of the pistil non-S-factors identified in Nicotiana and Solanum, and encodes a small asparagine/aspartate-rich extracellular protein with unknown biochemical function. Here, HT-B was cloned from Petunia and characterized. The structural features and expression pattern of Petunia HT-B were very similar to those of Nicotiana and Solanum. Unlike other solanaceous species, expression of HT-B was also observed in self-compatible Petunia species. RNA interference (RNAi)-mediated suppression of Petunia HT-B resulted in partial breakdown of GSI. Quantitative analysis of the HT-B mRNA accumulation in the transgenics showed that a 100-fold reduction is not sufficient and a >1000-fold reduction is required to achieve partial breakdown of GSI

Plant Ureases and Related Peptides: Understanding Their Entomotoxic Properties

Recently, ureases were included in the arsenal of plant defense proteins, alongside many other proteins with biotechnological potential such as insecticides. Isoforms of Canavalia ensiformis urease (canatoxin—CNTX and jack bean urease—JBURE-I) are toxic to insects of different orders. This toxicity is due in part to the release of a 10 kDa peptide from the native protein, by cathepsin-like enzymes present in the insect digestive tract. The entomotoxic peptide, Jaburetox-2Ec, exhibits potent insecticidal activity against several insects, including many resistant to the native ureases. JBURE-I and Jaburetox-2Ec cause major alterations of post-feeding physiological processes in insects, which contribute to, or can be the cause of, their entomotoxic effect. An overview of the current knowledge on plant urease processing and mechanisms of action in insects is presented in this review

A disulfide bond A-like oxidoreductase is a strong candidate gene for self-incompatibility in apricot (Prunus armeniaca) pollen

[EN] S-RNase based gametophytic self-incompatibility (SI) is a widespread prezygotic reproductive barrier in flowering plants. In the Solanaceae, Plantaginaceae and Rosaceae gametophytic SI is controlled by the pistil-specific S-RNases and the pollen S-locus F-box proteins but non-S-specific factors, namely modifiers, are also required. In apricot, Prunus armeniaca (Rosaceae), we previously mapped two pollen-part mutations that confer self-compatibility in cultivars Canino and Katy at the distal end of chromosome 3 (M-locus) unlinked to the S-locus. Here, we used high-resolution mapping to identify the M-locus with an similar to 134 kb segment containing ParM-1-16 genes. Gene expression analysis identified four genes preferentially expressed in anthers as modifier gene candidates, ParM-6, -7, -9 and -14. Variant calling of WGS Illumina data from Canino, Katy, and 10 self-incompatible cultivars detected a 358 bp miniature inverted-repeat transposable element (MITE) insertion in ParM-7 shared only by self-compatible apricots, supporting ParM-7 as strong candidate gene required for SI. ParM-7 encodes a disulfide bond A-like oxidoreductase protein, which we named ParMDO. The MITE insertion truncates the ParMDO ORF and produces a loss of SI function, suggesting that pollen rejection in Prunus is dependent on redox regulation. Based on phylogentic analyses we also suggest that ParMDO may have originated from a tandem duplication followed by subfunctionalization and pollenspecific expression.This work was supported by two grants from the Ministerio de Economia y Competitividad del Gobierno de Espana (AGL19018-2010 and AGL2015-64625-C2-2-R). The authors want to thank Inmaculada Lopez for her technical contribution and Gary Clark for his assistance with the manuscript. Chris Dardick, Tetyana Zhebentyayeva, and Albert Abbott kindly provided Goldrich and SEO genomic sequences. We also thank Mario Fares, and especially, Bruce McClure for their insights and comments on the manuscript.Muñoz-Sanz, JV.; Zuriaga García, E.; Badenes, ML.; Romero Salvador, C. (2017). A disulfide bond A-like oxidoreductase is a strong candidate gene for self-incompatibility in apricot (Prunus armeniaca) pollen. Journal of Experimental Botany. 68(18):5069-5078. https://doi.org/10.1093/jxb/erx336S50695078681

One chance to get it right: improving clinical handovers for better symptom control at the end of life.

Poor communication contributes to morbidity and mortality, not only in general medical care but also at the end oflife. This leads to issues relating to symptom control and quality of care. As part of an international project focused on bereaved relatives’ perceptions about quality of end-of-life care, we undertook a quality improvement (QI) project in a general hospital in Córdoba city, Argentina. By using two iterative QI cycles, we launched an educational process and introduced a clinical mnemonic tool, I-PASS, during ward handovers. The introduction of the handover tool was intended to improve out-of-hours care. Our clinical outcome measure was ensuring comfort in at least 60% of dying patients, as perceived by family carers, during night shifts in an oncology ward during the project period (March–May 2019). As process-based measures, we selected the proportion of staff completing the I-PASS course (target 60%) and using I-PASS in at least 60% of handovers. Participatory action research was the chosen method. During the study period, 13/16 dying patients were included. We received 23 reports from family carers about the level of patient comfort during the previous night. Sixty-five per cent of healthcare professionals completed the I-PASS training. The percentage of completed handovers increased from 60% in the first Plan-Do-Study-Act (PDSA) cycle to 68% in the second one. The proportion of positive reports about patient comfort increased from 63% (end of the first PDSA cycle) to 87% (last iterative analysis after 3 months). Moreover, positive responses to ‘Did doctors and nurses do enough for the patient to be comfortable during the night?’ increased from 75% to 100% between the first and the second QI cycle. In conclusion, we achieved the successful introduction and staff training for use of the I-PASS tool. This led to improved perceptions by family carers, about comfort for dying patients