Zebrafish Cytosolic Carboxypeptidases 1 and 5 Are Essential for Embryonic Development

The cytosolic carboxypeptidases (CCPs) are a subfamily of metalloenzymes within the larger M14 family of carboxypeptidases that have been implicated in the post-translational modification of tubulin. It has been suggested that at least four of the six mammalian CCPs function as tubulin deglutamylases. However, it is not yet clear whether these enzymes play redundant or unique roles within the cell. To address this question, genes encoding CCPs were identified in the zebrafish genome. Analysis by quantitative polymerase chain reaction indicated that CCP1, CCP2, CCP5, and CCP6 mRNAs were detectable between 2 h and 8 days postfertilization with highest levels 5–8 days postfertilization. CCP1, CCP2, and CCP5 mRNAs were predominantly expressed in tissues such as the brain, olfactory placodes, and pronephric ducts. Morpholino oligonucleotide-mediated knockdown of CCP1 and CCP5 mRNA resulted in a common phenotype including ventral body curvature and hydrocephalus. Confocal microscopy of morphant zebrafish revealed olfactory placodes with defective morphology as well as pronephric ducts with increased polyglutamylation. These data suggest that CCP1 and CCP5 play important roles in developmental processes, particularly the development and functioning of cilia. The robust and similar defects upon knockdown suggest that each CCP may have a function in microtubule modification and ciliary function and that other CCPs are not able to compensate for the loss of one

New approaches to investigating the function of mycelial networks

Fungi play a key role in ecosystem nutrient cycles by scavenging, concentrating, translocating and redistributing nitrogen. To quantify and predict fungal nitrogen redistribution, and assess the importance of the integrity of fungal networks in soil for ecosystem function, we need better understanding of the structures and processes involved. Until recently nitrogen translocation has been experimentally intractable owing to the lack of a suitable radioisotope tracer for nitrogen, and the impossibility of observing nitrogen translocation in real time under realistic conditions. We have developed an imaging method for recording the magnitude and direction of amino acid flow through the whole mycelial network as it captures, assimilates and channels its carbon and nitrogen resources, while growing in realistically heterogeneous soil microcosms. Computer analysis and modeling, based on these digitized video records, can reveal patterns in transport that suggest experimentally testable hypotheses. Experimental approaches that we are developing include genomics and stable isotope NMR to investigate where in the system nitrogen compounds are being acquired and stored, and where they are mobilized for transport or broken down. The results are elucidating the interplay between environment, metabolism, and the development and function of transport networks as mycelium forages in soil. The highly adapted and selected foraging networks of fungi may illuminate fundamental principles applicable to other supply networks

Strategic Scheduling of Infrastructure Repair and Maintenance: Volume 1—Decision Tree for Steel Bridge Painting

Even for the right treatment, improper timing can have consequences: premature application (treatment is applied too early) could mean wasteful spending even if users enjoy the benefits of higher pavement condition; deferred or delayed application (treatment is applied too late) could result in higher user costs due to poor condition and even reduced asset longevity. The objectives of this research were to establish the optimal condition or timing for each of the standard M&R treatment types typically used by INDOT; quantify the consequences of departures from such optimal conditions or timings; and to establish the optimal M&R treatment schedule for each asset family. The study focused on three asset types and their treatments: Painting of Steel Bridges. A painting decision tree was developed, to serve as a framework that would enable INDOT to consider other maintenance treatment types, namely spot repair/painting and overcoating. Bridge Deck Maintenance and Rehabilitation. Life-cycle condition-based deck M&R strategies based on different trigger results were proposed and presented. Pavement Maintenance, Rehabilitation, and Replacement. A framework was established to find the optimal scheduling for multiple treatments and recommend appropriate long-term M&R strategies for flexible and rigid pavements on different functional classes

Why Medical Students Choose Rural Clinical Campuses For Training: A Report From Two Campuses At Opposite Ends Of The Commonwealth

Introduction Although US medical schools have increased their enrollment by about 30%, most of the increase has occurred in urban areas. The affinity model proposes that rural training of a rural student will more likely result in a rural physician, but the exact role of these rural campuses is unclear. Do they solidify and reinforce a pre-existing career plan, do they create social and marital ties that make the transition to rural medicine easier, or could they be replaced with a briefer and more efficient rural rotation? We administered a questionnaire to students attending two different two year rural clinical campuses in the same state in order to explore their opinions regarding the advantages of a rural campus. Methods Two different rural M3-M4 year clinical campuses, affiliated with different medical schools in the same state, administered surveys to 70 medical students across all four years of medical school. Both schools selectively recruit rural students to the rural campuses, and require a campus decision at admission. Both schools require students to attend their first two years at an urban campus, and transfer to the rural campus for clinical education. Questions addressed student opinions on rural campus location, recommendations from others regarding attendance, campus atmosphere and social life, teaching methods and involvement in patient care. Comparisons were analyzed using the non-parametric Mann Whitney U test. Results The top five reasons students chose a rural campus included three aspects of rural training and two features of being rural. There were small differences between the two campuses regarding the importance of more procedures during training and more outdoor activities, the opportunity to study with friends, and strength of local leadership, reflecting differences in the practice setting and the environment of the two campuses. Differences were also noted between upper-level and lower-level students regarding the importance of studying with friends, and the chances of meeting a future spouse. Finally, very rural students (30 miles from urban area) were less concerned with availability of scholarships, and lack of fine dining, but viewed the opportunity to study with friends more favorably. Conclusions This study adds to the published literature by surveying students at multiple rural campuses by year of training. There were many more similarities than differences, but there were differences between the two campuses, and there were also differences as the students progressed in their training, and differences between very rural students and other students attending the campus. Rural campuses provide both clinical and social support for students contemplating rural practice. Results of the survey indicate both are of importance to the students as well, with quality of training the most important factor

Trait‐based approaches reveal fungal adaptations to nutrient‐limiting conditions

The dependency of microbial activity on nutrient availability in soil is only partly understood, but highly relevant for nutrient cycling dynamics. In order to achieve more insight on microbial adaptations to nutrient limiting conditions, precise physiological knowledge is needed. Therefore, we developed an experimental system assessing traits of 16 saprobic fungal isolates in nitrogen (N) limited conditions. We tested the hypotheses that (1) fungal traits are negatively affected by N deficiency to a similar extent and (2) fungal isolates respond in a phylogenetically conserved fashion. Indeed, mycelial density, spore production and fungal activity (respiration and enzymatic activity) responded similarly to limiting conditions by an overall linear decrease. By contrast, mycelial extension and hyphal elongation peaked at lowest N supply (C:N 200), causing maximal biomass production at intermediate N contents. Optimal N supply rates differed among isolates, but only the extent of growth reduction was phylogenetically conserved. In conclusion, growth responses appeared as a switch from explorative growth in low nutrient conditions to exploitative growth in nutrient‐rich patches, as also supported by responses to phosphorus and carbon limitations. This detailed trait‐based pattern will not only improve fungal growth models, but also may facilitate interpretations of microbial responses observed in field studies

Estimation and Prediction of Statewide Vehicle Miles Traveled (VMT) by Highway Category and Vehicle Classification

Vehicle Miles Traveled (VMT) is a critical performance measure that is used extensively in highway transportation management for financial analysis, resource allocation, impact assessments, and reporting to oversight agencies. As highway revenue from fuel taxes continues to plummet and user-based taxes such as VMT fees become increasingly attractive, consistent and reliable VMT estimates have become critical for highway funding evaluation and administration. At the present time, there are several methods for VMT estimation that typically yield estimates that are inconsistent or inaccurate. This study was commissioned by the Indiana Department of Transportation (INDOT) to develop a benchmark method for VMT estimation and provide calibration factors for the VMT estimation methods. The study’s core outcome is a segment-level framework for VMT estimation For the state roads, a comprehensive database was developed which facilitates extensive aggregations of VMT by geographical scope, route, functional class, and vehicle class. For the local roads, a sample of counties of different spatial locations and degrees of urbanization were used, and cluster analysis, geographic information systems (GIS), and spatial interpolation techniques were used to expand the VMT estimates from the local road sample to the population of all counties in the state. The results of this study indicate that there is significant variation in the results from the various VMT estimation methods. The technique developed in this study for reconciling these different VMT estimates was validated using the estimate from the benchmark method (segment-level) as a basis. The implementation platform developed in this study was designed to produce outcomes that address the VMT data needs of the intended end users and stakeholders and could be enhanced in the future as and when data become available

Impact of Tide-Topography Interactions on Basal Melting of Larsen C Ice Shelf, Antarctica

Basal melting of ice shelves around Antarctica contributes to formation of Antarctic Bottom Water and can affect global sea level by altering the offshore flow of grounded ice streams and glaciers. Tides influence ice shelf basal melt rate (w(b)) by contributing to ocean mixing and mean circulation as well as thermohaline exchanges with the ice shelf. We use a three-dimensional ocean model, thermodynamically coupled to a nonevolving ice shelf, to investigate the relationship between topography, tides, and w(b) for Larsen C Ice Shelf (LCIS) in the northwestern Weddell Sea, Antarctica. Using our best estimates of ice shelf thickness and seabed topography, we find that the largest modeled LCIS melt rates occur in the northeast, where our model predicts strong diurnal tidal currents (similar to 0.4 m s(-1)). This distribution is significantly different from models with no tidal forcing, which predict largest melt rates along the deep grounding lines. We compare several model runs to explore melt rate sensitivity to geometry, initial ocean potential temperature (theta(0)), thermodynamic parameterizations of heat and freshwater ice-ocean exchange, and tidal forcing. The resulting range of LCIS-averaged w(b) is similar to 0.11-0.44 m a(-1). The spatial distribution of w(b) is very sensitive to model geometry and thermodynamic parameterization while the overall magnitude of w(b) is influenced by theta(0). These sensitivities in w(b) predictions reinforce a need for high-resolution maps of ice draft and sub-ice-shelf seabed topography together with ocean temperature measurements at the ice shelf front to improve representation of ice shelves in coupled climate system models

Naturally Occurring Carboxypeptidase A6 Mutations: Effect on Enzyme Function and Association with Epilepsy

Carboxypeptidase A6 (CPA6) is a member of the A/B subfamily of M14 metallocarboxypeptidases that is expressed in brain and many other tissues during development. Recently, two mutations in human CPA6 were associated with febrile seizures and/or temporal lobe epilepsy. In this study we screened for additional CPA6 mutations in patients with febrile seizures and focal epilepsy, which encompasses the temporal lobe epilepsy subtype. Mutations found from this analysis as well as CPA6 mutations reported in databases of single nucleotide polymorphisms were further screened by analysis of the modeled proCPA6 protein structure and the functional role of the mutated amino acid. The point mutations predicted to affect activity and/or protein folding were tested by expression of the mutant in HEK293 cells and analysis of the resulting CPA6 protein. Common polymorphisms in CPA6 were also included in this analysis. Several mutations resulted in reduced enzyme activity or CPA6 protein levels in the extracellular matrix. The mutants with reduced extracellular CPA6 protein levels showed normal levels of 50-kDa proCPA6 in the cell, and this could be converted into 37-kDa CPA6 by trypsin, suggesting that protein folding was not greatly affected by the mutations. Interestingly, three of the mutations that reduced extracellular CPA6 protein levels were found in patients with epilepsy. Taken together, these results provide further evidence for the involvement of CPA6 mutations in human epilepsy and reveal additional rare mutations that inactivate CPA6 and could, therefore, also be associated with epileptic phenotypes. © 2012 by The American Society for Biochemistry and Molecular Biology, Inc

The structural and dynamic responses of Stange Ice Shelf to recent environmental change

Stange Ice Shelf is the most south-westerly ice shelf on the Antarctic Peninsula, a region where positive trends in atmospheric and oceanic temperatures have been recently documented. In this paper, we use a range of remotely sensed datasets to evaluate the structural and dynamic responses of Stange Ice Shelf to these environmental changes. Ice shelf extent and surface structures were examined at regular intervals from optical and radar satellite imagery between 1973 and 2011. Surface speeds were estimated in 1989, 2004 and 2010 by tracking surface features in successive satellite images. Surface elevation change was estimated using radar altimetry data acquired between 1992 and 2008 by the European Remote Sensing Satellite (ERS) -1, -2 and Envisat. The mean number of surface melt days was estimated using the intensity of backscatter from Envisat’s Advanced Synthetic Aperture Radar instrument between 2006 and 2012. These results show significant shear fracturing in the southern portion of the ice shelf linked to enhanced flow speed as a consequence of measured thinning. However, we conclude that, despite the observed changes, Stange Ice Shelf is currently stable