Biomedical subjects

Elide Formentin

Publications and source records attributed to Elide Formentin.

7 recordsLinked to original sources

PubMed2006

KDC2, a functional homomeric potassium channel expressed during carrot embryogenesis.

In Daucus carota, the model system for embryogenesis, it has been demonstrated that potassium and K(+) selective channels are involved in embryo development. Here, we report the isolation and cloning of a new carrot Shaker-like potassium channel, potassium D. carota channel 2 (KDC2), whose expression pattern during somatic embryogenesis proceeds along with the establishment of the polar axes and the settlement of the hypocotyl region. In plants, KDC2 transcript is localized at the shoot level, in the epidermis and guard cells, similarly to its Arabidopsis homolog KAT1. Electrophysiological assays indicated KDC2 as the first carrot subunit able to form homomeric functional channels in Xenopus oocytes, with properties similar to those of Arabidopsis KAT1.

Amino Acid Sequence↗

PubMed2005

Molecular cloning and characterization of a phytochelatin synthase gene, PvPCS1, from Pteris vittata L.

Pteris vittata L. is a staggeringly efficient arsenic hyperaccumulator that has been shown to be capable of accumulating up to 23,000 microg arsenic g(-1), and thus represents a species that may fully exploit the adaptive potential of plants to toxic metals. However, the molecular mechanisms of adaptation to toxic metal tolerance and hyperaccumulation remain unknown, and P. vittata genes related to metal detoxification have not yet been identified. Here, we report the isolation of a full-length cDNA sequence encoding a phytochelatin synthase (PCS) from P. vittata. The cDNA, designated PvPCS1, predicts a protein of 512 amino acids with a molecular weight of 56.9 kDa. Homology analysis of the PvPCS1 nucleotide sequence revealed that it has low identity with most known plant PCS genes except AyPCS1, and the homology is largely confined to two highly conserved regions near the 5'-end, where the similarity is as high as 85-95%. The amino acid sequence of PvPCS1 contains two Cys-Cys motifs and 12 single Cys, only 4 of which (Cys-56, Cys-90/91, and Cys-109) in the N-terminal half of the protein are conserved in other known PCS polypeptides. When expressed in Saccharomyces cerevisae, PvPCS1 mediated increased Cd tolerance. Cloning of the PCS gene from an arsenic hyperaccumulator may provide information that will help further our understanding of the genetic basis underlying toxic metal tolerance and hyperaccumulation.

Aminoacyltransferases↗

PubMed2004

DKT1, a novel K+ channel from carrot, forms functional heteromeric channels with KDC1.

We report the isolation and characterisation of DKT1, a new carrot K+ channel alpha-subunit belonging to the Shaker-like family. DKT1 is expressed in many tissues of the adult plant, suggesting that it may play important roles in both nutrition and other important physiological processes. During embryo development, DKT1 is expressed at later phases implying the involvement of K+ in embryo maturation. When co-expressed with KDC1 in Xenopus oocytes, DKT1 is able to form functional, heteromeric channels, suggesting that possible interactions between these two ion channels in plant tissues may modulate K+ uptake.

Amino Acid Sequence↗

PubMed2004

Potassium and carrot embryogenesis: are K+ channels necessary for development?

The expression pattern of the KDC1 gene, coding for an inwardly-rectifying K(+) channel of Daucus carota , is described in several embryo stages and seedling tissues. Relative quantitative RT-PCR experiments indicated that, during (somatic) embryonic development, the KDC1 transcript appears as early as the globular stage and that the transcript level remains constant throughout the successive heart and torpedo stages. Thereafter, the KDC1 transcript is preferentially expressed in plant roots, but is also present in other tissues, and in particular, in the shoot apical meristem. In situ hybridisation experiments showed that in embryos KDC1 mRNA is detectable preferentially in protoderm cells with a stage dependent expression pattern. At later times, the hybridisation signal is particularly evident in root hairs, root epidermis and endodermis, but is also observed in single cell layers corresponding to L1 of the shoot apical meristem and leaf primordia. Promoter studies with the beta -glucuronidase reporter gene confirm preferential expression of KDC1 in embryo protoderm cells and in plant root epidermis and root hairs. Western blot analysis of embryonic proteins and immunolocalisation experiments on somatic embryos sections revealed the presence of KDC1 during embryo development. Consistent with these observations, patch-clamp experiments performed on protoplasts isolated from embryos at the torpedo stage demonstrated the presence of functional inward rectifying K(+) channels. This is the first report on the expression of a plant ion channel during embryo development.

Blotting, Western↗

PubMed2004

Histidines are responsible for zinc potentiation of the current in KDC1 carrot channels.

Unlike all plant inward-rectifying potassium channels, the carrot channel KDC1 has two histidine pairs (H161,H162) in the S3-S4 and (H224,H225) in the S5-S6 linkers. When coinjected with KAT1 in Xenopus oocytes, KDC1 participates in the formation of heteromultimeric KDC1:KAT1 channels and the ionic current is potentiated by extracellular Zn2+. To investigate the potential interactions between KDC1 and zinc, a KDC1-KAT1 dimer was constructed. The dimeric and heteromeric channels displayed similar characteristics and the same sensitivity to zinc and other metals; this result suggests that zinc binding is mediated by residues in a single channel subunit. The KDC1:KAT1 currents were also potentiated by external Pb2+ and Cd2+ and inhibited by Ni2+. To investigate further the role of KDC1-histidines, these amino acids were mutated into alanines. The single mutations H225A, H161A, and H162A did not affect the response of the heteromeric channels to zinc. Conversely, the single mutant H224A and the double mutants (H224A,H225A) and (H161A,H162A) abolished zinc potentiation, but not that induced by Pb2+ or Cd2+. These results suggest that Zn2+ potentiation cannot be ascribed to simple electrostatic interactions between zinc and channel residues and that histidine 224 is crucial for zinc but not for lead potentiation of the current.

Amino Acid Substitution↗

PubMed2002

Cytokinins: new apoptotic inducers in plants.

High concentrations of cytokinins block cell proliferation and induce programmed cell death (PCD) in both carrot ( Daucus carota L.) and Arabidopsis thaliana (L.) Heynh. cell cultures [13 and 27 micro M N(6)-benzylaminopurine (BAP), respectively]. In the present work, cell death was scored by Evan's blue staining and was also demonstrated to be programmed by various parameters, including chromatin condensation, oligonucleosomal DNA degradation (laddering), and release of cytochrome c from mitochondria. In carrot cells, this induction takes approximately 24 h, with proliferating cells being more sensitive than quiescent ones. Two hormones, namely abscisic acid and 2,4-dichlorophenoxyacetic acid (2,4-D), protect cells against the cytokinin-induced death. PCD is not merely a consequence of the inability of the culture to proliferate, since high levels of 2,4-D block carrot cell proliferation without promoting PCD. Increased ethylene production was also observed in BAP-treated cultures, although this increase was not responsible for PCD because inhibitors of ethylene synthesis and action did not block PCD in BAP-treated cultures. Programmed cell death in the form of DNA laddering was also seen in plants treated with cytokinins. This process was accompanied by accelerated senescence in the form of leaf yellowing.

2,4-Dichlorophenoxyacetic Acid↗

PubMed2002

Nitric oxide affects plant mitochondrial functionality in vivo.

In this report, we show that nitric oxide affects mitochondrial functionality in plant cells and reduces total cell respiration due to strong inhibition of the cytochrome pathway. The residual respiration depends on the alternative pathway and novel synthesis of alternative oxidase occurs. These modifications are associated with depolarisation of the mitochondrial membrane potential and release of cytochrome c from mitochondria, suggesting a conserved signalling pathway in plants and animals. This signal cascade is triggered at the mitochondrial level and induces about 20% of cell death. In order to achieve a higher level of cell death, the addition of H(2)O(2) is necessary.

Cell Death↗