The Arabidopsis THADA homologue modulates TOR activity and cold acclimation.

Low temperature is one of the most important environmental factors that affect global survival of humans and animals and equally importantly the distribution of plants and crop productivity. Survival of metazoan cells under cold stress requires regulation of the sensor-kinase Target Of Rapamycin (TOR). TOR controls growth of eukaryotic cells by adjusting anabolic and catabolic metabolism. Previous studies identified the Thyroid Adenoma Associated (THADA) gene as the major effect locus by positive selection in the evolution of modern human adapted to cold. Here we investigate the role of THADA in TOR signaling and cold acclimation of plants. We applied BLAST searches and homology modeling to identify the AtTHADA (AT3G55160) in Arabidopsis thaliana as the highly probable orthologue protein. Reverse genetics approaches were combined with immunological detection of TOR activity and metabolite profiling to address the role of the TOR and THADA for growth regulation and cold acclimation. Depletion of the AtTHADA gene caused complete or partial loss of full-length mRNA, respectively, and significant retardation of growth under non-stressed conditions. Furthermore, depletion of AtTHADA caused hypersensitivity towards low-temperatures. Atthada displayed a lowered energy charge. This went along with decreased TOR activity, which offers a molecular explanation for the slow growth phenotype of Atthada. Finally, we used TOR RNAi lines to identify the de-regulation of TOR activity as one determinant for sensitivity towards low-temperatures. Taken together our results provide evidence for a conserved function of THADA in cold acclimation of eukaryotes and suggest that cold acclimation in plants requires regulation of TOR.

Implementing carrier screening for cystic fibrosis outside the clinic: ethical analysis in the light of the personalist view.

BACKGROUND: Cystic Fibrosis (CF) is an autosomal recessive genetic disease. Two models for screening CF are normally used: newborn screening and population-based CF carrier screening. In turn, there are three main models of population-based CF carrier screening: prenatal carrier screening, preconception carrier screening, and carrier screening outside clinical settings. AIM: To evaluate, in the light of the personalist view, the use of carrier screenings for CF outside the clinic, i.e. in non-clinical settings, such as school and workplaces. METHODS: Analysis has been carried out according to the "Personalist approach" (also called "Triangular model"), an ethical method for performing ethical analysis within HTA process. It includes factual, anthropological and ethical data in a ''triangular'' normative reflection process. FINDINGS: Implementing carrier screening for cystic fibrosis outside the clinical settings allows acquisition of knowledge for informing reproductive choices, that can be considered as valuable; benefit-risk ratio seems to be not much favorable; autonomous and responsible decisions can be taken only under certain conditions; economic advantage is difficult to determine; therefore, from a personalist view, implementing carrier screenings outside the clinic seems not to be ethically justified. CONCLUSIONS: In accordance with the personalist perspective, public health programs providing carrier screenings outside the clinic should not be implemented.

[Genetic test for cancer and intra-family communication: freedom vs. responsibility]. / Test genetici in oncologia e comunicazione intrafamiliare: autonomia vs responsabilità.

Genetic tests affect not only single patients but also their genetic relatives. In some cases, they in fact allow to acquire information not only about a single patient, but also about those who are genetically linked (genetic relatives). By appealing to the principle of autonomy, the patient can refuse to be informed of the test result, or to inform their relatives on the risk of a pathology. How might the relatives' right to know be reconciled with the will of a patient who refuses to know or to inform? Among the large number of moral dilemmas that this field can raise, the article aims to reply to the above mentioned question and to analyse in depth some aspects of intra-family communication within the field of genetic tests for cancer.

Ethical evaluation of risks related to living donor transplantation programs.

The shortage of available cadaveric organs for transplantation and the growing demand has incresed live donation. To increase the number of transplantations from living donors, programs have been implemented to coordinate donations in direct or indirect form (cross-over, paired, and domino chain). Living donors with complex medical conditions are accepted by several transplantation programs. In this way, the number of transplants from living has exceeded that from cadaver donors in several European countries. No mortality has been reported in the case of lung, pancreas, or intestinal Living donations, but the perioperative complications range from 15% to 30% for pancreas and lung donors. In living kidney donors, the perioperative mortality is 3 per 10,000. Their frequency of end-stage renal disease does not exceed the United States rate for the general population. However, long-term follow-up studies of living donors for kidney transplantations have several limitations. The frequency of complications in live donor liver transplantation is 40%, of these, 48% are possibly life-threatening according to the Clavien classification. Residual disability, liver failure, or death has occurred in 1% of cases. The changes in live donor acceptance criteria raise ethical issues, in particular, the physician's role in evaluating and accepting the risks taken by the living donor. Some workers argue to set aside medical paternalism on behalf of the principle of donor autonomy. In this way the medical rule "primum non nocere" is overcome. Transplantation centers should reason beyond the shortage of organs and think in terms of the care for both donor and recipient.

Drosophila: a model for understanding obesity and diabetic complications.

The molecular mechanisms underlying development of obesity and diabetic complications are not well understood. Drosophila has become a popular model organism for studying a variety of human diseases. We discuss here emerging Drosophila models of obesity and diabetic complications.

Dpp gradient formation in the Drosophila wing imaginal disc.

The secreted signaling protein Dpp acts as a morphogen to pattern the anterior-posterior axis of the Drosophila wing. Dpp activity is required in all cells of the developing wing imaginal disc, but the ligand gradient that supports this activity has not been characterized. Here we make use of a biologically active form of Dpp tagged with GFP to examine the ligand gradient. Dpp-GFP forms an unstable extracellular gradient that spreads rapidly in the wing disc. The activity gradient visualized by MAD phosphorylation differs in shape from the ligand gradient. The pMAD gradient adjusted to compartment size when this was experimentally altered. These observations suggest that the Dpp activity gradient may be shaped at the level of receptor activation.

Chromosome arrangement within a bacterium.

BACKGROUND: The contour length of the circular chromosome of bacteria is greater than a millimeter but must be accommodated within a cell that is only a few micrometers in length. Bacteria do not have nucleosomes and little is known about the arrangement of the chromosome inside a prokaryotic cell. RESULTS: We have investigated the arrangement of chromosomal DNA within the bacterium Bacillus subtilis by using fluorescence microscopy to visualize two sites on the chromosome simultaneously in the same cell. Indirect immunofluorescence with antibodies against the chromosome partition protein Spo0J were used to visualize the replication origin region of the chromosome. Green fluorescent protein fused to the lactose operon repressor Lacl was used to decorate tandem copies of the lactose operon operator lacO. A cassette of tandem operators was separately inserted into the chromosome near the origin (359 degrees), near the replication terminus (181 degrees), or at two points in between (90 degrees and 270 degrees). The results show that the layout of the chromosome is dynamic but is principally arranged with the origin and terminus maximally apart and the quarter points of the chromosome in between. CONCLUSIONS: The use of cytological methods to visualize two chromosomal sites in the same cell has provided a glimpse of the arrangement of a bacterial chromosome. We conclude that, to a first approximation, the folding of the bacterial chromosome is consistent with, and may preserve, the linear order of genes on the DNA.

Use of time-lapse microscopy to visualize rapid movement of the replication origin region of the chromosome during the cell cycle in Bacillus subtilis.

We describe the use of time-lapse fluorescence microscopy to visualize the movement of the DNA replication origin and terminus regions on the Bacillus subtilis chromosome during the course of the cell cycle. The origin and terminus regions were tagged with a cassette of tandem lac operator repeats and visualized through the use of a fusion of the green fluorescent protein to the LacI repressor. We have discovered that origin regions abruptly move apart towards the cell poles during a brief interval of the cell cycle. This movement was also seen in the absence of cell wall growth and in the absence of the product of the parB homologue spo0J. The origin regions moved apart an average distance of 1.4 microm in an 11 min period of abrupt movement, representing an average velocity of 0.17 microm min(-1), and reaching a maximum velocity of greater than 0.27 microm min(-1). The terminus region also exhibited a striking pattern of movement but not as far or a rapid as the origin region. These results provide evidence for a mitotic-like motor that is responsible for segregation of the origin regions of the chromosomes.