Helena Moura de Carvalho

Immunocytolocalization of glutamine synthetase in mesophyll and phloem of leaves of Solanum tuberosum L.

SummaryLocalization of glutamine synthetase inSolanum tuberosum leaves was investigated by techniques of Western tissue printing and immunogold electron microscopy. Anti-GS antibodies used in immunolocalization recognize two peptides (45 kDa and 42 kDa) on Western blots. Antibody stained tissue prints on nitrocellulose membranes allowed low resolution localization of GS. Immunostaining was most evident in the adaxial phloem of the leaf midribs and petiole veins. High-resolution localization of glutamine synthetase by immunogold electron microscopy revealed that this enzyme occurs in both the chloroplasts and the cytosol ofS. tuberosum leaf cells. However, GS was specifically associated with the chloroplasts of mesophyll cells and with the cytoplasm of phloem companion cells. The evidence for cell-specific localization of chloroplast and cytosolic GS presented here agrees with the recently reported cell-specific pattern of expression of GUS reporter gene, directed by promoters for chloroplast and cytosolic GS form in tobacco transgenic plants. These data provide additional clues to the interpretation of the functional role of these different isoenzymes and its relationship with their specific localization.

Heteromeric assembly of the cytosolic glutamine synthetase polypeptides of Medicago truncatula: complementation of a glnA Escherichia coli mutant with a plant domain-swapped enzyme

We have cloned and sequenced the cDNAs corresponding to the two cytosolic glutamine synthetase (GS) polypeptides (a and b) of Medicago truncatula. Using these two cDNAs we have prepared a construct encoding the N-terminal domain of b and the C-terminal domain of a in order to produce a domain-swapped polypeptide which should assemble to give an enzyme containing chimeric active sites. Both the native and the domain-swapped enzymes were expressed in Escherichia coli where they were catalytically and physiologically active as they were able to rescue a glnA deletion mutant. The expressed polypeptides were of the correct size and the isoenzymes behaved similarly to their native homologues on ion-exchange chromatography. We have found slight differences in the kinetic properties of the purified enzymes and in the modulation of their activities by several putative cellular effectors. In vitro dissociation of the purified a and b homo-octamers, followed by reassociation, showed that the subunits are able to self-assemble, perhaps randomly, to form heteromeric isoenzymes. Moreover, heteromeric isoenzymes occur in the plant as revealed by studies on the GS isoenzymes of nodules, roots, stems and stipules.

LegumeGRN: a gene regulatory network prediction server for functional and comparative studies.

Building accurate gene regulatory networks (GRNs) from high-throughput gene expression data is a long-standing challenge. However, with the emergence of new algorithms combined with the increase of transcriptomic data availability, it is now reachable. To help biologists to investigate gene regulatory relationships, we developed a web-based computational service to build, analyze and visualize GRNs that govern various biological processes. The web server is preloaded with all available Affymetrix GeneChip-based transcriptomic and annotation data from the three model legume species, i.e., Medicago truncatula, Lotus japonicus and Glycine max. Users can also upload their own transcriptomic and transcription factor datasets from any other species/organisms to analyze their in-house experiments. Users are able to select which experiments, genes and algorithms they will consider to perform their GRN analysis. To achieve this flexibility and improve prediction performance, we have implemented multiple mainstream GRN prediction algorithms including co-expression, Graphical Gaussian Models (GGMs), Context Likelihood of Relatedness (CLR), and parallelized versions of TIGRESS and GENIE3. Besides these existing algorithms, we also proposed a parallel Bayesian network learning algorithm, which can infer causal relationships (i.e., directionality of interaction) and scale up to several thousands of genes. Moreover, this web server also provides tools to allow integrative and comparative analysis between predicted GRNs obtained from different algorithms or experiments, as well as comparisons between legume species. The web site is available at http://legumegrn.noble.org.

AHP6 Inhibits Cytokinin Signaling to Regulate the Orientation of Pericycle Cell Division during Lateral Root Initiation

In Arabidopsis thaliana, lateral roots (LRs) initiate from anticlinal cell divisions of pericycle founder cells. The formation of LR primordia is regulated antagonistically by the phytohormones cytokinin and auxin. It has previously been shown that cytokinin has an inhibitory effect on the patterning events occurring during LR formation. However, the molecular players involved in cytokinin repression are still unknown. In a similar manner to protoxylem formation in Arabidopsis roots, in which AHP6 (ARABIDOPSIS HISTIDINE PHOSPHOTRANSFER PROTEIN 6) acts as a cytokinin inhibitor, we reveal that AHP6 also functions as a cytokinin repressor during early stages of LR development. We show that AHP6 is expressed at different developmental stages during LR formation and is required for the correct orientation of cell divisions at the onset of LR development. Moreover, we demonstrate that AHP6 influences the localization of the auxin efflux carrier PIN1, which is necessary for patterning the LR primordia. In summary, we show that the inhibition of cytokinin signaling through AHP6 is required to establish the correct pattern during LR initiation.

Ser e tornar-se Psicoterapeuta parte I: diálogo entre experiências pessoais e profissionais

The psychotherapist has been considered an element of major importance for the success of therapy, however the study of his/her personal and professional characteristics has been neglected in research. The purpose of this work is to analyze the narratives of 20 psychotherapists experts from different theoretical orientations (behavioral-cognitive, constructivist, humanistic, psychoanalytic and systemic) on their personal and professional development during clinical practice. Data were collected through a semi-structured interview and analyzed in a constructive perspective of grounded theory. In the analysis two main themes emerged. In this article are presented the results of the first theme, relating to experiences of personal and professional context that contributed to the development of psychotherapists. The organizing theme of the interviews emphasized the importance of dialogue and the consistency between the personal and professional self-understanding of the development of the psychotherapist.O psicoterapeuta tem sido considerado um elemento de primordial importância para o sucesso da psicoterapia no entanto, o estudo das suas caracteristicas pessoais e profissionais tem sido negligenciado na investigacao. O objetivo do presente artigo e analisar as narrativas de 20 psicoterapeutas experts (peritos) de diferentes orientacoes teoricas (cognitivo-comportamental, construtivista, humanista, psicanalitica e sistemica) sobre o seu desenvolvimento pessoal e profissional no decurso da pratica clinica. Os dados foram recolhidos atraves de uma entrevista semiestruturada e foram analisados em uma perspetiva construtivista da grounded theory. Na analise emergiram dois grandes temas. Serao apresentados aqui os resultados do primeiro tema analisado, relativo as experiencias relacionais do âmbito pessoal e profissional que contribuiram para o desenvolvimento dos psicoterapeutas. O tema organizador das entrevistas sublinha a importância do dialogo e da coerencia entre o self pessoal e profissional na compreensao do desenvolvimento do psicoterapeuta.

The Arabidopsis HP6 gene is expressed in Medicago truncatula lateral roots and root nodule primordia

Expression patterns of orthologous genes can be similar between distantly related species, suggesting that developmental programs can be conserved between organisms. Here, we show that the promoter of AHP6, a gene which is involved in Arabidopsis lateral root development, also drives the expression of the reporter GUS gene in lateral roots of Medicago truncatula suggesting that similar regulatory elements are involved in lateral root organogenesis in these species. Interestingly, the AHP6 promoter was able to drive GUS expression in root nodules and nodule primordia, structures that are absent in Arabidopsis. We found two AHP6 orthologous genes in the M. truncatula genome and we speculate that these putative cytokinin inhibitors may play a role during lateral root and nodule development in this species.

Being and becoming a Psychotherapist part I: dialogue between personal and professional experiences

The psychotherapist has been considered an element of major importance for the success of therapy, however the study of his/her personal and professional characteristics has been neglected in research. The purpose of this work is to analyze the narratives of 20 psychotherapists experts from different theoretical orientations (behavioral-cognitive, constructivist, humanistic, psychoanalytic and systemic) on their personal and professional development during clinical practice. Data were collected through a semi-structured interview and analyzed in a constructive perspective of grounded theory. In the analysis two main themes emerged. In this article are presented the results of the first theme, relating to experiences of personal and professional context that contributed to the development of psychotherapists. The organizing theme of the interviews emphasized the importance of dialogue and the consistency between the personal and professional self-understanding of the development of the psychotherapist.O psicoterapeuta tem sido considerado um elemento de primordial importância para o sucesso da psicoterapia no entanto, o estudo das suas caracteristicas pessoais e profissionais tem sido negligenciado na investigacao. O objetivo do presente artigo e analisar as narrativas de 20 psicoterapeutas experts (peritos) de diferentes orientacoes teoricas (cognitivo-comportamental, construtivista, humanista, psicanalitica e sistemica) sobre o seu desenvolvimento pessoal e profissional no decurso da pratica clinica. Os dados foram recolhidos atraves de uma entrevista semiestruturada e foram analisados em uma perspetiva construtivista da grounded theory. Na analise emergiram dois grandes temas. Serao apresentados aqui os resultados do primeiro tema analisado, relativo as experiencias relacionais do âmbito pessoal e profissional que contribuiram para o desenvolvimento dos psicoterapeutas. O tema organizador das entrevistas sublinha a importância do dialogo e da coerencia entre o self pessoal e profissional na compreensao do desenvolvimento do psicoterapeuta.

Ser y tornarse Psicoterapeuta parte I: diálogo entre experiencias personales y profesionales

The psychotherapist has been considered an element of major importance for the success of therapy, however the study of his/her personal and professional characteristics has been neglected in research. The purpose of this work is to analyze the narratives of 20 psychotherapists experts from different theoretical orientations (behavioral-cognitive, constructivist, humanistic, psychoanalytic and systemic) on their personal and professional development during clinical practice. Data were collected through a semi-structured interview and analyzed in a constructive perspective of grounded theory. In the analysis two main themes emerged. In this article are presented the results of the first theme, relating to experiences of personal and professional context that contributed to the development of psychotherapists. The organizing theme of the interviews emphasized the importance of dialogue and the consistency between the personal and professional self-understanding of the development of the psychotherapist.O psicoterapeuta tem sido considerado um elemento de primordial importância para o sucesso da psicoterapia no entanto, o estudo das suas caracteristicas pessoais e profissionais tem sido negligenciado na investigacao. O objetivo do presente artigo e analisar as narrativas de 20 psicoterapeutas experts (peritos) de diferentes orientacoes teoricas (cognitivo-comportamental, construtivista, humanista, psicanalitica e sistemica) sobre o seu desenvolvimento pessoal e profissional no decurso da pratica clinica. Os dados foram recolhidos atraves de uma entrevista semiestruturada e foram analisados em uma perspetiva construtivista da grounded theory. Na analise emergiram dois grandes temas. Serao apresentados aqui os resultados do primeiro tema analisado, relativo as experiencias relacionais do âmbito pessoal e profissional que contribuiram para o desenvolvimento dos psicoterapeutas. O tema organizador das entrevistas sublinha a importância do dialogo e da coerencia entre o self pessoal e profissional na compreensao do desenvolvimento do psicoterapeuta.