Brown adipose tissue specific lack of uncoupling protein 3 is associated with impaired cold tolerance and reduced transcript levels of metabolic genes

Uncoupling protein 3 (Ucp3) is located within the mitochondrial inner membrane of brown adipose tissue and skeletal muscle. It is thought to be implicated in lipid metabolism and defense against reactive oxygen species. We previously reported on a mutation in our breeding colony of Djungarian hamsters (Phodopus sungorus) that leads to brown adipose tissue specific lack of Ucp3 expression. In this study we compared wildtype with mutant hamsters on a broad genetic background. Hamsters lacking Ucp3 in brown adipose tissue displayed a reduced cold tolerance due to impaired nonshivering thermogenesis. This phenotype is associated with a global decrease in expression of metabolic genes but not of uncoupling protein 1. These data implicate that Ucp3 is necessary to sustain high metabolic rates in brown adipose tissue.     

Dose-dependent and sequence-sensitive effects of amyloid-beta peptide on neurosteroidogenesis in human neuroblastoma cells

Interactions between neurosteroidogenesis and proteins involved in age-related diseases are unknown. High concentrations of amyloid-beta (A beta) peptides induce plaques in Alzheimer's disease but several studies demonstrated that physiological or non-toxic doses are neuroprotective. We compared the effects of non-toxic and toxic concentrations of A beta 1-42 and A beta 25-35 on neurosteroidogenesis in human neuroblastoma SH-SY5Y cells. Viability assays revealed that nanomolar doses of A beta are devoid of cytotoxicity while 12 microM induced cell death. Pulse-chase, high-performance liquid chromatography and flow-scintillation analyses showed that non-toxic A beta 1-42 concentrations, acting selectively, decreased [3H]progesterone but increased [3H]estradiol production from the precursor [3H]pregnenolone. Non-toxic A beta 25-35 doses reduced [3H]progesterone formation but had no effect on [3H]estradiol biosynthesis. At 12 microM, both A beta 1-42 and A beta 25-35 inhibited [3H]progesterone formation but only A beta 1-42 reduced [3H]estradiol production. The results demonstrate a selective and amino-acid sequence-dependent action of A beta on neurosteroidogenesis. The fact that non-toxic A beta 1-42 doses stimulated neuroprotective-neurosteroid estradiol synthesis, which is inhibited by high A beta 1-42 doses, may explain A beta 1-42 ability to exert either protective or deleterious effects on nerve cells.     

Comparative genomics of the pIPO2/pSB102 family of environmental plasmids: sequence, evolution, and ecology of pTer331 isolated from Collimonas fungivorans Ter331

Plasmid pTer331 from the bacterium Collimonas fungivorans Ter331 is a new member of the pIPO2/pSB102 family of environmental plasmids. The 40 457-bp sequence of pTer331 codes for 44 putative ORFs, most of which represent genes involved in replication, partitioning and transfer of the plasmid. We confirmed that pTer331 is stably maintained in its native host. Deletion analysis identified a mini-replicon capable of replicating autonomously in Escherichia coli and Pseudomonas putida. Furthermore, plasmid pTer331 was able to mobilize and retromobilize IncQ plasmid pSM1890 at typical rates of 10(-4) and 10(-8), respectively. Analysis of the 91% DNA sequence identity between pTer331 and pIPO2 revealed functional conservation of coding sequences, the deletion of DNA fragments flanked by short direct repeats (DR), and sequence preservation of long DRs. In addition, we experimentally established that pTer331 has no obvious contribution in several of the phenotypes that are characteristic of its host C. fungivorans Ter331, including the ability to efficiently colonize plant roots. Based on our findings, we hypothesize that cryptic plasmids such as pTer331 and pIPO2 might not confer an individual advantage to bacteria, but, due to their broad-host-range and ability to retromobilize, benefit bacterial populations by accelerating the intracommunal dissemination of the mobile gene pool.     

Chloroplast nitrite uptake is enhanced in Arabidopsis PII mutants

In higher plants, the PII protein is a nuclear-encoded plastid protein that regulates the activity of a key enzyme of arginine biosynthesis. We have previously observed that Arabidopsis PII mutants are more sensitive to nitrite toxicity. Using intact chloroplasts isolated from Arabidopsis leaves and (15)N-labelled nitrite we show that a light-dependent nitrite uptake into chloroplasts is increased in PII knock-out mutants when compared to the wild-type. This leads to a higher incorporation of (15)N into ammonium and amino acids in the mutant chloroplasts. However, the uptake differences do not depend on GS/GOGAT activities. Our observations suggest that PII is involved in the regulation of nitrite uptake into higher plant chloroplasts.     

Distributed delays stabilize neural feedback systems

We consider the effect of distributed delays in neural feedback systems. The avian optic tectum is reciprocally connected with the isthmic nuclei. Extracellular stimulation combined with intracellular recordings reveal a range of signal delays from 3 to 9 ms between isthmotectal elements. This observation together with prior mathematical analysis concerning the influence of a delay distribution on system dynamics raises the question whether a broad delay distribution can impact the dynamics of neural feedback loops. For a system of reciprocally connected model neurons, we found that distributed delays enhance system stability in the following sense. With increased distribution of delays, the system converges faster to a fixed point and converges slower toward a limit cycle. Further, the introduction of distributed delays leads to an increased range of the average delay value for which the system's equilibrium point is stable. The system dynamics are determined almost exclusively by the mean and the variance of the delay distribution and show only little dependence on the particular shape of the distribution.     

Chemical Hypoxia Facilitates Alternative Splicing of EAAT2 in Presymptomatic APP23 Transgenic Mice

Hypoxia is one of the major common components of vascular risk factors for pathogenesis of Alzheimer’s disease. This study investigated the possible relationship between hypoxia and alternative splicing of the excitatory amino acid transporter 2 (EAAT2) in a transgenic model for Alzheimer’s disease. We used an APP23 mouse model prior to amyloid deposition and subjected it to chemical hypoxia treatment as induced by 3-nitropropionic acid. One hour after administration of 3-nitropropionic acid changes in the expression of the 5′-splice forms mEAAT2/5UT3, mEAAT2/5UT4, and mEAAT2/5UT5 were found in the frontal cortex, hippocampus and cerebellum of the APP23 model. In untreated APP23 animals the expression of EAAT2 splice variants was unchanged. Our results demonstrate that hypoxia facilitates alternative splicing of EAAT2 in the APP23 model. This may be a molecular mechanism linking vascular factors to early pathophysiology of Alzheimer’s disease.     

Indirect readout in drug-DNA recognition: role of sequence-dependent DNA conformation

DNA-binding drugs have numerous applications in the engineered gene regulation. However, the drug-DNA recognition mechanism is poorly understood. Drugs can recognize specific DNA sequences not only through direct contacts but also indirectly through sequence-dependent conformation, in a similar manner to the indirect readout mechanism in protein-DNA recognition. We used a knowledge-based technique that takes advantage of known DNA structures to evaluate the conformational energies. We built a dataset of non-redundant free B-DNA crystal structures to calculate the distributions of adjacent base-step and base-pair conformations, and estimated the effective harmonic potentials of mean force (PMF). These PMFs were used to calculate the conformational energy of drug-DNA complexes, and the Z-score as a measure of the binding specificity. Comparing the Z-scores for drug-DNA complexes with those for free DNA structures with the same sequence, we observed that in several cases the Z-scores became more negative upon drug binding. Furthermore, the specificity is position-dependent within the drug-bound region of DNA. These results suggest that DNA conformation plays an important role in the drug-DNA recognition. The presented method provides a tool for the analysis of drug-DNA recognition and can facilitate the development of drugs for targeting a specific DNA sequence.     

Cooperative activity of DNA methyltransferases for maintenance of symmetrical and non-symmetrical cytosine methylation in Arabidopsis thaliana

Maintenance of cytosine methylation in plants is controlled by three DNA methyltransferases. MET1 maintains CG methylation, and DRM1/2 and CMT3 act redundantly to enforce non-CG methylation. RPS, a repetitive hypermethylated DNA fragment from Petunia hybrida, attracts DNA methylation when transferred into Petunia or other species. In Arabidopsis thaliana, which does not contain any RPS homologues, RPS transgenes are efficiently methylated in all sequence contexts. To test which DNA methylation pathways regulate RPS methylation, we examined maintenance of RPS methylation in various mutant backgrounds. Surprisingly, CG methylation was lost in a drm1/2/cmt3 mutant, and non-CG methylation was almost completely eliminated in a met1 mutant. An unusual cooperative activity of all three DNA methyltransferases is therefore required for maintenance of both CG and non-CG methylation in RPS. Other unusual features of RPS methylation are the independence of its non-CG methylation from the RNA-directed DNA methylation (RdDM) pathway and the exceptional maintenance of methylation at a CC(m)TGG site in some epigenetic mutants. This is indicative of activity of a methylation system in plants that may have evolved from the DCM methylation system that controls CC(m)WGG methylation in bacteria. Our data suggest that strict separation of CG and non-CG methylation pathways does not apply to all target regions, and that caution is required in generalizing methylation data obtained for individual genomic regions.     

Identification of metabolic and biomass QTL in Arabidopsis thaliana in a parallel analysis of RIL and IL populations

Plant growth and development are tightly linked to primary metabolism and are subject to natural variation. In order to obtain an insight into the genetic factors controlling biomass and primary metabolism and to determine their relationships, two Arabidopsis thaliana populations [429 recombinant inbred lines (RIL) and 97 introgression lines (IL), derived from accessions Col-0 and C24] were analyzed with respect to biomass and metabolic composition using a mass spectrometry-based metabolic profiling approach. Six and 157 quantitative trait loci (QTL) were identified for biomass and metabolic content, respectively. Two biomass QTL coincide with significantly more metabolic QTL (mQTL) than statistically expected, supporting the notion that the metabolic profile and biomass accumulation of a plant are linked. On the same basis, three out the six biomass QTL can be simulated purely on the basis of metabolic composition. QTL based on analysis of the introgression lines were in substantial agreement with the RIL-based results: five of six biomass QTL and 55% of the mQTL found in the RIL population were also found in the IL population at a significance level of P  ≤ 0.05, with >80% agreement on the allele effects. Some of the differences could be attributed to epistatic interactions. Depending on the search conditions, metabolic pathway-derived candidate genes were found for 24–67% of all tested mQTL in the database AraCyc 3.5. This dataset thus provides a comprehensive basis for the detection of functionally relevant variation in known genes with metabolic function and for identification of genes with hitherto unknown roles in the control of metabolism.     

As good as dead? Sublethal predation facilitates lethal predation on an intertidal clam

Although ecologists have speculated that sublethal predation can impact prey dynamics, consequences of these predator effects have seldom been experimentally tested. In soft‐sediment marine communities, fishes crop extended feeding siphons of buried clams, potentially causing clams to reduce their burial depth, thereby enhancing their susceptibility to excavating lethal predators. We simulated cropping of the confamilial clams, Protothaca staminea and Venerupis philippinarum, by removing the top 40% of siphons, which caused each species to burrow 33–50% shallower than conspecifics with intact siphons. To examine subsequent consequences of reduced burial depth, we exposed cropped and intact clams to natural levels of predation in the field. Because of a naturally longer siphon, Protothaca, even after cropping, remained at relatively safe burial depths. In contrast, siphon cropping nearly doubled the mortality rate of Venerupis. Thus, while sublethal predation facilitates lethal predation, this linkage depends on specific life history characteristics, even among ecologically similar species.