Not so crystal clear: the structure of the human telomere G-quadruplex in solution differs from that present in a crystal

The structure of human telomere DNA is of intense interest because of its role in the biology of both cancer and aging. The sequence [5′-AGGG(TTAGGG)(3)] has been used as a model for telomere DNA in both NMR and X-ray crystallographic studies, the results of which show dramatically different structures. In Na(+) solution, NMR revealed an antiparallel G-quadruplex structure that featured both diagonal and lateral TTA loops. Crystallographic studies in the presence of K(+) revealed a flattened, propeller-shaped structure featuring a parallel-stranded G-quadruplex with symmetrical external TTA loops. We report the results of biophysical experiments in solution and computational studies that are inconsistent with the reported crystal structure, indicating that a different structure exists in K(+) solutions. Sedimentation coefficients were determined experimentally in both Na(+) and K(+) solutions and were compared with values calculated using bead models for the reported NMR and crystal structures. Although the solution NMR structure accurately predicted the observed S-value in Na(+) solution, the crystal structure predicted an S-value that differed dramatically from that experimentally observed in K(+) solution. The environments of loop adenines were probed by quantitative fluorescence studies using strategic and systematic single-substitutions of 2-aminopurine for adenine bases. Both fluorescence intensity and quenching experiments in K(+) yielded results at odds with quantitative predictions from the reported crystal structure. Circular dichroism and fluorescence quenching studies in the presence of the crowding agent polyethylene glycol showed dramatic changes in the quadruplex structure in K(+) solutions, but not in Na(+) solutions, suggesting that the crystal environment may have selected for a particular conformational form. Molecular dynamics simulations were performed to yield model structures for the K(+) quadruplex form that are consistent with our biophysical results and with previously reported chemical modification studies. These models suggest that the biologically relevant structure of the human telomere quadruplex in K(+) solution is not the one determined in the published crystalline state

Visual Experiences during Paralysis

Rationale: Paralyzed human volunteers (n = 6) participated in several studies the primary one of which required full neuromuscular paralysis while awake. After the primary experiment, while still paralyzed and awake, subjects undertook studies of humor and of attempted eye-movement. The attempted eye-movements tested a central, intentional component to one’s internal visual model and are the subject of this report. Methods: Subjects reclined in a supportive chair and were ventilated after paralysis (cisatracurium, 20 mg intravenously). In illumination, subjects were requested to focus alternately on the faces of investigators standing on the left and the right within peripheral vision. In darkness, subjects were instructed to look away from a point source of light. Subjects were to report their experiences after reversal of paralysis. Results: During attempted eye-movement in illumination, one subject had an illusion of environmental movement but four subjects perceived faces as clearly as if they were in central vision. In darkness, four subjects reported movement of the target light in the direction of attempted eye-movements and three could control the movement of the light at will. Conclusion: The hypothesis that internal visual models receive intended ocular-movement-information directly from oculomotor centers is strengthened by this evidence

Visual Experiences during Paralysis

Rationale: Paralyzed human volunteers (n = 6) participated in several studies the primary one of which required full neuromuscular paralysis while awake. After the primary experiment, while still paralyzed and awake, subjects undertook studies of humor and of attempted eye-movement. The attempted eye-movements tested a central, intentional component to one’s internal visual model and are the subject of this report. Methods: Subjects reclined in a supportive chair and were ventilated after paralysis (cisatracurium, 20 mg intravenously). In illumination, subjects were requested to focus alternately on the faces of investigators standing on the left and the right within peripheral vision. In darkness, subjects were instructed to look away from a point source of light. Subjects were to report their experiences after reversal of paralysis. Results: During attempted eye-movement in illumination, one subject had an illusion of environmental movement but four subjects perceived faces as clearly as if they were in central vision. In darkness, four subjects reported movement of the target light in the direction of attempted eye-movements and three could control the movement of the light at will. Conclusion: The hypothesis that internal visual models receive intended ocular-movement-information directly from oculomotor centers is strengthened by this evidence

Coordination of opposing sex-specific and core muscle groups regulates male tail posture during Caenorhabditis elegans male mating behavior

Background To survive and reproduce, animals must be able to modify their motor behavior in response to changes in the environment. We studied a complex behavior of Caenorhabditis elegans, male mating behavior, which provided a model for understanding motor behaviors at the genetic, molecular as well as circuit level. C. elegans male mating behavior consists of a series of six sub-steps: response to contact, backing, turning, vulva location, spicule insertion, and sperm transfer. The male tail contains most of the sensory structures required for mating, in addition to the copulatory structures, and thus to carry out the steps of mating behavior, the male must keep his tail in contact with the hermaphrodite. However, because the hermaphrodite does not play an active role in mating and continues moving, the male must modify his tail posture to maintain contact. We provide a better understanding of the molecular and neuro-muscular pathways that regulate male tail posture during mating. Results Genetic and laser ablation analysis, in conjunction with behavioral assays were used to determine neurotransmitters, receptors, neurons and muscles required for the regulation of male tail posture. We showed that proper male tail posture is maintained by the coordinated activity of opposing muscle groups that curl the tail ventrally and dorsally. Specifically, acetylcholine regulates both ventral and dorsal curling of the male tail, partially through anthelmintic levamisole-sensitive, nicotinic receptor subunits. Male-specific muscles are required for acetylcholine-driven ventral curling of the male tail but dorsal curling requires the dorsal body wall muscles shared by males and hermaphrodites. Gamma-aminobutyric acid activity is required for both dorsal and ventral acetylcholine-induced curling of the male tail and an inhibitory gamma-aminobutyric acid receptor, UNC-49, prevents over-curling of the male tail during mating, suggesting that cross-inhibition of muscle groups helps maintain proper tail posture. Conclusion Our results demonstrated that coordination of opposing sex-specific and core muscle groups, through the activity of multiple neurotransmitters, is required for regulation of male tail posture during mating. We have provided a simple model for regulation of male tail posture that provides a foundation for studies of how genes, molecular pathways, and neural circuits contribute to sensory regulation of this motor behavior

The Proximal Drivers of Large Fires: A Pyrogeographic Study

Variations in global patterns of burning and fire regimes are relatively well measured, however, the degree of influence of the complex suite of biophysical and human drivers of fire remains controversial and incompletely understood. Such an understanding is required in order to support current fire management and to predict the future trajectory of global fire patterns in response to changes in these determinants. In this study we explore and compare the effects of four fundamental controls on fire, namely the production of biomass, its drying, the influence of weather on the spread of fire and sources of ignition. Our study area is southern Australia, where fire is currently limited by either fuel production or fuel dryness. As in most fire-prone environments, the majority of annual burned area is due to a raelatively small number of large fires. We train and test an Artificial Neural Networks ability to predict spatial patterns in the probability of large fires (>1,250 ha) in forests and grasslands as a function of proxies of the four major controls on fire activity. Fuel load is represented by predicted forested biomass and remotely sensed grass biomass, drying is represented by fraction of the time monthly potential evapotranspiration exceeds precipitation, weather is represented by the frequency of severe fire weather conditions and ignitions are represented by the average annual density of reported ignitions. The response of fire to these drivers is often non-linear. Our results suggest that fuel management will have limited capacity to alter future fire occurrence unless it yields landscape-scale changes in fuel amount, and that shifts between, rather than within, vegetation community types may be more important. We also find that increased frequency of severe fire weather could increase the likelihood of large fires in forests but decrease it in grasslands. These results have the potential to support long-term strategic planning and risk assessment by fire management agencies.OP’s salary was provided by the NSW Rural Fire Service. MB was partly financially supported by the Bushfires and Natural Hazards Cooperative Research Centre

Prospective Molecular Profiling of Canine Cancers Provides a Clinically Relevant Comparative Model for Evaluating Personalized Medicine (PMed) Trials.

Background Molecularly-guided trials (i.e. PMed) now seek to aid clinical decision-making by matching cancer targets with therapeutic options. Progress has been hampered by the lack of cancer models that account for individual-to-individual heterogeneity within and across cancer types. Naturally occurring cancers in pet animals are heterogeneous and thus provide an opportunity to answer questions about these PMed strategies and optimize translation to human patients. In order to realize this opportunity, it is now necessary to demonstrate the feasibility of conducting molecularly-guided analysis of tumors from dogs with naturally occurring cancer in a clinically relevant setting. Methodology A proof-of-concept study was conducted by the Comparative Oncology Trials Consortium (COTC) to determine if tumor collection, prospective molecular profiling, and PMed report generation within 1 week was feasible in dogs. Thirty-one dogs with cancers of varying histologies were enrolled. Twenty-four of 31 samples (77%) successfully met all predefined QA/QC criteria and were analyzed via Affymetrix gene expression profiling. A subsequent bioinformatics workflow transformed genomic data into a personalized drug report. Average turnaround from biopsy to report generation was 116 hours (4.8 days). Unsupervised clustering of canine tumor expression data clustered by cancer type, but supervised clustering of tumors based on the personalized drug report clustered by drug class rather than cancer type. Conclusions Collection and turnaround of high quality canine tumor samples, centralized pathology, analyte generation, array hybridization, and bioinformatic analyses matching gene expression to therapeutic options is achievable in a practical clinical window (\u3c1 \u3eweek). Clustering data show robust signatures by cancer type but also showed patient-to-patient heterogeneity in drug predictions. This lends further support to the inclusion of a heterogeneous population of dogs with cancer into the preclinical modeling of personalized medicine. Future comparative oncology studies optimizing the delivery of PMed strategies may aid cancer drug development

THE RADIAL AND ROTATIONAL VELOCITIES OF PSO J318.5338-22.8603, A NEWLY CONFIRMED PLANETARY-MASS MEMBER OF THE β PICTORIS MOVING GROUP

PSO J318.5338$-$22.8603 is an extremely-red planetary-mass object that has been identified as a candidate member of the $\beta$ Pictoris moving group based on its spatial position and tangential velocity. We present a high resolution $K$-band spectrum of PSO J318.5338$-$22.8603. Using a forward-modeling Markov Chain Monte Carlo approach, we report the first measurement of the radial velocity and $v$ sin($i$) of PSO J318.5$-$22, $-$6.0$^{+0.8}_{-1.1}$ km s$^{-1}$ and 17.5$^{+2.3}_{-2.8}$ km s$^{-1}$, respectively. We calculate the space velocity and position of PSO J318.5$-$22 and confirm that it is a member of the $\beta$ Pictoris moving group. Adopting an age of 23$\pm$3 Myr for PSO J318.5$-$22, we determine a mass of $8.3\pm0.5$ $M_{\rm{Jup}}$ and effective temperature of $1127^{+24}_{-26}$ K using evolutionary models. PSO J318.5338$-$22.8603 is intermediate in mass and temperature to the directly-imaged planets $\beta$ Pictoris b and 51 Eridani b, making it an important benchmark object in the sequence of planetary-mass members of the $\beta$ Pictoris moving group. Combining our $v$ sin($i$) measurement with recent photometric variability data, we constrain the inclination of PSO J318.5$-$22 to $>29^{\circ}$ and its rotational period to 5-10.2 hours. The equatorial velocity of PSO J318.5$-$22 indicates that its rotation is consistent with an extrapolation of the velocity-mass relationship for solar system planets.Comment: 8 pages, 5 figures, 2 tables, ApJ accepte