Nucleocytoplasmic partitioning of the plant photoreceptors phytochrome A,B, C,D, and E is regulated differentially by light and exhibits a diurnal rhythm

The phytochrome family of plant photoreceptors has a central role in the adaptation of plant development to changes in ambient light conditions. The individual phytochrome species regulate different or partly overlapping physiological responses. We generated transgenic Arabidopsis plants expressing phytochrome A to E:green fluorescent protein (GFP) fusion proteins to assess the biological role of intracellular compartmentation of these photoreceptors in light-regulated signaling. We show that all phytochrome:GFP fusion proteins were imported into the nuclei. Translocation of these photoreceptors into the nuclei was regulated differentially by light. Light-induced accumulation of phytochrome species in the nuclei resulted in the formation of speckles. The appearance of these nuclear structures exhibited distinctly different kinetics, wavelengths, and fluence dependence and was regulated by a diurnal rhythm. Furthermore, we demonstrate that the import of mutant phytochrome B:GFP and phytochrome A:GFP fusion proteins, shown to be defective in signaling in vivo, is regulated by light but is not accompanied by the formation of speckles. These results suggest that (1) the differential regulation of the translocation of phytochrome A to E into nuclei plays a role in the specification of functions, and (2) the appearance of speckles is a functional feature of phytochrome-regulated signaling.     

Missense mutation in the PAS2 domain of phytochrome A impairs subnuclear localization and a subset of responses

Phytochrome A signaling shows two photobiologically discrete outputs: so-called very-low-fluence responses (VLFR) and high-irradiance responses (HIR). By modifying previous screening protocols, we isolated two Arabidopsis mutants retaining VLFR and lacking HIR. Phytochrome A negatively or positively regulates phytochrome B signaling, depending on light conditions. These mutants retained the negative but lacked the positive regulation. Both mutants carry the novel phyA-302 allele, in which Glu-777 (a residue conserved in angiosperm phytochromes) changed to Lys in the PAS2 motif of the C-terminal domain. The phyA-302 mutants showed a 50% reduction in phytochrome A levels in darkness, but this difference was compensated for by greater stability under continuous far-red light. phyA-302:green fluorescent protein fusion proteins showed normal translocation from the cytosol to the nucleus under continuous far-red light but failed to produce nuclear spots, suggesting that nuclear speckles could be involved in HIR signaling and phytochrome A degradation. We propose that the PAS2 domain of phytochrome A is necessary to initiate signaling in HIR but not in VLFR, likely via interaction with a specific partner.     

Near-infrared fluorescent nanoparticles as combined MR/optical imaging probes

A number of quantitative three-dimensional tomographic near-infrared fluorescence imaging techniques have recently been developed and combined with MR imaging to yield highly detailed anatomic and molecular information in living organisms (1, 2). Here we describe magnetic nanoparticle based MR contrast agents that have a near-infrared fluorescence (NIRF) that is activated by certain enzymes. The probes are prepared by conjugation of arginyl peptides to cross-linked iron oxide amine (amino-CLIO), either by a disulfide linkage or a thioether linker, followed by the attachment of the indocyanine dye Cy5.5. The NIRF of disulfide-linked conjugate was activated by DTT, while the NIRF of thioether-linked conjugate was activated by trypsin. Fluorescent quenching of the attached fluorochrome occurs in part due to the interaction with iron oxide, as evident by the activation of fluorescence with DTT when nanoparticles that have less than one dye attached per particle. With a SC injection of the probe, axillary and brachial lymph nodes were darkened on MR images and easily delineated by NIRF imaging. The probes may provide the basis for a new class of so-called smart nanoparticles, capable of pinpointing their position through their magnetic properties, while providing information on their environment by optical imaging techniques.     

Untersuchungen über die Verteilung der ABO-Blutgruppen bei verschiedenen Erkrankungen im Säuglingsalter

Summary In the last ten years the ABO blood groups of our infants have been constantly ascertained and their distribution among some of the more prevalent diseases have been investigated. Other infants and healthy new-borns were used for comparison. The following results have been found:The blood group O is slightly more frequent in sick babies than in healthy new-borns. Among patients, who were under the age of 2 months at the time of admission, there were more with blood group O than among the older children. The blood group A tends to increase among the infants with anemia and with acute respiratory diseases, the blood group O tends to increase among those with staphylococcal infections and rachitis. With the latter the blood group B seems also to be found a little more frequently.Statistics show the increase of the blood group A in anemia in the 3. and 4. months of life and in acute respiratory diseases between the 3. and 12. month of life. The increase of the blood group O in staphylococcal infections is sure only among the youngest patients during their 1. month of life, it is obvious among the older patients, and there mainly among those of masculine sex. The increase of the blood groups O and B among infants with rachitis at the age of 3 to 12 months is almost certain. Premature babies show a sure decrease of the blood group O among both sexes, an increase of the blood group A among feminine, and of the blood group B among masculine patients.     

Blood pressure changes following chronic administration to rats of 3beta,16beta-dihydroxy-5-androsten-17-one, 3beta,17beta-dihydroxy-5-androsten-16-one and 21-hydroxy-4-pregnene-3,20-dione-21-acetate.

The urinary excretion of 3beta,16beta-dihydroxy-5-androsten-17-one (16beta-OH-DHEA) is increased in patients with low renin essential hypertension. This steroid and its isomer 3beta,17beta-dihydroxy-5-androsten-16-one (16-oxo-A) have also been reported to have mineralocorticoid activity in adrenalectomized rats. These findings have led to the postulate that excessive secretion of 16beta-OH-DHEA may be responsible for the production of low renin essential hypertension. In this study unilaterally nephrectomized salt loaded rats injected once a week with 30 mg of 11-desoxycorticosterone acetate per/kg of body weight for 2 month periods developed hypertension. Rats given similar amounts of 16beta-OH-DHEA or 16-oxo-A and rats given no steroids did not develop hypertension. We conclude that it is unlikely that 16beta-OH-DHEA and 16-oxo-A are direct causative factors in the production of low renin essential hypertension.     

Nuclear localization of the Arabidopsis blue light receptor cryptochrome 2

The cryptochrome blue light photoreceptor family of Arabidopsis thaliana consists of two members, CRY1 and CRY2 (PHH1). CRY2 contains a putative nuclear localization signal (NLS) within its C-terminal region. We examined whether CRY2 is localized in the nucleus and whether the C-terminal region of CRY2 is involved in nuclear targeting. Total cellular and nuclear protein extracts from Arabidopsis were subjected to immunoblot analysis with CRY2-specific antibodies. Strong CRY2 signals were obtained in the nuclear fraction. Fusion proteins consisting of the green fluorescent protein (GFP) and different fragments of CRY2 were expressed in parsley protoplasts and the localization of the fusion proteins was determined by fluorescence and confocal laser scanning microscopy. GFP-fusions containing the entire CRY2 protein or its C-terminal region were found exclusively in the nucleus. We conclude from these results that CRY2 is localized in the nucleus and that nuclear localization is mediated by the C-terminal region of CRY2.     

Removal of deaminated cytosines and detection of in vivo methylation in ancient DNA

DNA sequences determined from ancient organisms have high error rates, primarily due to uracil bases created by cytosine deamination. We use synthetic oligonucleotides, as well as DNA extracted from mammoth and Neandertal remains, to show that treatment with uracil–DNA–glycosylase and endonuclease VIII removes uracil residues from ancient DNA and repairs most of the resulting abasic sites, leaving undamaged parts of the DNA fragments intact. Neandertal DNA sequences determined with this protocol have greatly increased accuracy. In addition, our results demonstrate that Neandertal DNA retains in vivo patterns of CpG methylation, potentially allowing future studies of gene inactivation and imprinting in ancient organisms.     

Cognitive behavioural therapy versus supportive therapy for persistent positive symptoms in psychotic disorders: The POSITIVE Study, a multicenter, prospective, single-blind, randomised controlled clinical trial

Background

It has been demonstrated that cognitive behavioural therapy (CBT) has a moderate effect on symptom reduction and on general well being of patients suffering from psychosis. However, questions regarding the specific efficacy of CBT, the treatment safety, the cost-effectiveness, and the moderators and mediators of treatment effects are still a major issue. The major objective of this trial is to investigate whether CBT is specifically efficacious in reducing positive symptoms when compared with non-specific supportive therapy (ST) which does not implement CBT-techniques but provides comparable therapeutic attention.

Methods/Design

The POSITIVE study is a multicenter, prospective, single-blind, parallel group, randomised clinical trial, comparing CBT and ST with respect to the efficacy in reducing positive symptoms in psychotic disorders. CBT as well as ST consist of 20 sessions altogether, 165 participants receiving CBT and 165 participants receiving ST. Major methodological aspects of the study are systematic recruitment, explicit inclusion criteria, reliability checks of assessments with control for rater shift, analysis by intention to treat, data management using remote data entry, measures of quality assurance (e.g. on-site monitoring with source data verification, regular query process), advanced statistical analysis, manualized treatment, checks of adherence and competence of therapists. Research relating the psychotherapy process with outcome, neurobiological research addressing basic questions of delusion formation using fMRI and neuropsychological assessment and treatment research investigating adaptations of CBT for adolescents is combined in this network. Problems of transfer into routine clinical care will be identified and addressed by a project focusing on cost efficiency.

Discussion

This clinical trial is part of efforts to intensify psychotherapy research in the field of psychosis in Germany, to contribute to the international discussion on psychotherapy in psychotic disorders, and to help implement psychotherapy in routine care. Furthermore, the study will allow drawing conclusions about the mediators of treatment effects of CBT of psychotic disorders.

Trial Registration

Current Controlled Trials ISRCTN29242879