Dramatic expansion of the black widow toxin arsenal uncovered by multi-tissue transcriptomics and venom proteomics.

BackgroundAnimal venoms attract enormous interest given their potential for pharmacological discovery and understanding the evolution of natural chemistries. Next-generation transcriptomics and proteomics provide unparalleled, but underexploited, capabilities for venom characterization. We combined multi-tissue RNA-Seq with mass spectrometry and bioinformatic analyses to determine venom gland specific transcripts and venom proteins from the Western black widow spider (Latrodectus hesperus) and investigated their evolution.ResultsWe estimated expression of 97,217 L. hesperus transcripts in venom glands relative to silk and cephalothorax tissues. We identified 695 venom gland specific transcripts (VSTs), many of which BLAST and GO term analyses indicate may function as toxins or their delivery agents. ~38% of VSTs had BLAST hits, including latrotoxins, inhibitor cystine knot toxins, CRISPs, hyaluronidases, chitinase, and proteases, and 59% of VSTs had predicted protein domains. Latrotoxins are venom toxins that cause massive neurotransmitter release from vertebrate or invertebrate neurons. We discovered ≥ 20 divergent latrotoxin paralogs expressed in L. hesperus venom glands, significantly increasing this biomedically important family. Mass spectrometry of L. hesperus venom identified 49 proteins from VSTs, 24 of which BLAST to toxins. Phylogenetic analyses showed venom gland specific gene family expansions and shifts in tissue expression.ConclusionsQuantitative expression analyses comparing multiple tissues are necessary to identify venom gland specific transcripts. We present a black widow venom specific exome that uncovers a trove of diverse toxins and associated proteins, suggesting a dynamic evolutionary history. This justifies a reevaluation of the functional activities of black widow venom in light of its emerging complexity

Control of Silica Dust in Slate Milling Operations

The Mine Safety and Health Administration (MSHA) conducted a study to assess the present state of technology used for controlling employees\u27 respirable silica-bearing dust exposures during dimension slate milling operations. For this study, twelve slate milling operations were visited and various measurements were taken and observations made. These measurements and observations included exhaust and room airflow, types of enclosures, water usage, and building volumes for each of the milling processes. These mills usually employed between five and twenty people. From the study, general guidelines were developed from the best practices observed and can be used at the various sawing, splitting, trimming, and drilling operations throughout the industry

Sustainable Water Management in the Southwestern United States: Reality or Rhetoric?

Background: While freshwater sustainability is generally defined as the provisioning of water for both people and the environment, in practice it is largely focused only on supplying water to furnish human population growth. Symptomatic of this is the state of Arizona, where rapid growth outside of the metropolitan Phoenix-Tucson corridor relies on the same groundwater that supplies year-round flow in rivers. Using Arizona as a case study, we present the first study in the southwestern United States that evaluates the potential impact of future population growth and water demand on streamflow depletion across multiple watersheds. Methodology/Principal Findings: We modeled population growth and water demand through 2050 and used four scenarios to explore the potential effects of alternative growth and water management strategies on river flows. Under the base population projection, we found that rivers in seven of the 18 study watersheds could be dewatered due to municipal demand. Implementing alternative growth and water management strategies, however, could prevent four of these rivers from being dewatered. Conclusions/Significance: The window of opportunity to implement water management strategies is narrowing. Because impacts from groundwater extraction are cumulative and cannot be immediately reversed, proactive water management strategies should be implemented where groundwater will be used to support new municipal demand. Our approach provides a low-cost method to identify where alternative water and growth management strategies may have the most impact, and demonstrates that such strategies can maintain a continued water supply for both people and the environment

Effectiveness of Cabs for Dust and Silica Control on Mobile Mining Equipment

The Mine Safety and Health Administration (MSHA) has conducted a study to evaluate the effectiveness of cabs for controlling silica dust exposure during operation of mobile mining equipment. This study focused on bulldozers, front -end loaders and haul trucks, was conducted at surface coal mining operations and underground metal and nonmetal mining operations. Each piece of equipment tested was equipped with a cab. The vehicles sampled were from a range of manufacturers having different types of filter media and air intake configurations. The purpose of this study was to determine the reduction of dust and silica exposure that could be achieved through the use of a well-maintained cab. For each piece of equipment, dust and silica concentrations inside and outside the cab were determined and compared. In some cases, filtration efficiencies could be calculated. A properly designed environmental cab is sealed, has an intake air filtration system, and a heating and cooling system. Cabs should have good seals around the doors and windows. Factors such as cab pressurization filtration systems, filter media, and maintenance practices were also examined. In some cases, dust and silica reduction of 90 to 95% were observed

Ovarian Transcriptomic Analyses in the Urban Human Health Pest, the Western Black Widow Spider

Due to their abundance and ability to invade diverse environments, many arthropods have become pests of economic and health concern, especially in urban areas. Transcriptomic analyses of arthropod ovaries have provided insight into life history variation and fecundity, yet there are few studies in spiders despite their diversity within arthropods. Here, we generated a de novo ovarian transcriptome from 10 individuals of the western black widow spider (Latrodectus hesperus), a human health pest of high abundance in urban areas, to conduct comparative ovarian transcriptomic analyses. Biological processes enriched for metabolism—specifically purine, and thiamine metabolic pathways linked to oocyte development—were significantly abundant in L. hesperus. Functional and pathway annotations revealed overlap among diverse arachnid ovarian transcriptomes for highly-conserved genes and those linked to fecundity, such as oocyte maturation in vitellogenin and vitelline membrane outer layer proteins, hormones, and hormone receptors required for ovary development, and regulation of fertility-related genes. Comparative studies across arachnids are greatly needed to understand the evolutionary similarities of the spider ovary, and here, the identification of ovarian proteins in L. hesperus provides potential for understanding how increased fecundity is linked to the success of this urban pest

Effects of climate variability and accelerated forest thinning on watershed-scale runoff in southwestern USA ponderosa pine forests

The recent mortality of up to 20% of forests and woodlands in the southwestern United States, along with declining stream flows and projected future water shortages, heightens the need to understand how management practices can enhance forest resilience and functioning under unprecedented scales of drought and wildfire. To address this challenge, a combination of mechanical thinning and fire treatments are planned for 238,000 hectares (588,000 acres) of ponderosa pine (Pinus ponderosa) forests across central Arizona, USA. Mechanical thinning can increase runoff at fine scales, as well as reduce fire risk and tree water stress during drought, but the effects of this practice have not been studied at scales commensurate with recent forest disturbances or under a highly variable climate. Modifying a historical runoff model, we constructed scenarios to estimate increases in runoff from thinning ponderosa pine at the landscape and watershed scales based on driving variables: pace, extent and intensity of forest treatments and variability in winter precipitation. We found that runoff on thinned forests was about 20% greater than unthinned forests, regardless of whether treatments occurred in a drought or pluvial period. The magnitude of this increase is similar to observed declines in snowpack for the region, suggesting that accelerated thinning may lessen runoff losses due to warming effects. Gains in runoff were temporary (six years after treatment) and modest when compared to mean annual runoff from the study watersheds (0–3%). Nonetheless gains observed during drought periods could play a role in augmenting river flows on a seasonal basis, improving conditions for water-dependent natural resources, as well as benefit water supplies for downstream communities. Results of this study and others suggest that accelerated forest thinning at large scales could improve the water balance and resilience of forests and sustain the ecosystem services they provide

Multi-tissue transcriptomics of the black widow spider reveals expansions, co-options, and functional processes of the silk gland gene toolkit

Background: Spiders (Order Araneae) are essential predators in every terrestrial ecosystem largely because they have evolved potent arsenals of silk and venom. Spider silks are high performance materials made almost entirely of proteins, and thus represent an ideal system for investigating genome level evolution of novel protein functions. However, genomic level resources remain limited for spiders. Results: We de novo assembled a transcriptome for the Western black widow (Latrodectus hesperus) from deeply sequenced cDNAs of three tissue types. Our multi-tissue assembly contained ~100,000 unique transcripts, of which > 27,000 were annotated by homology. Comparing transcript abundance among the different tissues, we identified 647 silk gland-specific transcripts, including the few known silk fiber components (e.g. six spider fibroins, spidroins). Silk gland specific transcripts are enriched compared to the entire transcriptome in several functions, including protein degradation, inhibition of protein degradation, and oxidation-reduction. Phylogenetic analyses of 37 gene families containing silk gland specific transcripts demonstrated novel gene expansions within silk glands, and multiple co-options of silk specific expression from paralogs expressed in other tissues. Conclusions: We propose a transcriptional program for the silk glands that involves regulating gland specific synthesis of silk fiber and glue components followed by protecting and processing these components into functional fibers and glues. Our black widow silk gland gene repertoire provides extensive expansion of resources for biomimetic applications of silk in industry and medicine. Furthermore, our multi-tissue transcriptome facilitates evolutionary analysis of arachnid genomes and adaptive protein systems. Electronic supplementary material The online version of this article (doi:10.1186/1471-2164-15-365) contains supplementary material, which is available to authorized users

Alternative Transcription at Venom Genes and Its Role as a Complementary Mechanism for the Generation of Venom Complexity in the Common House Spider

The complex composition of venom, a proteinaceous secretion used by diverse animal groups for predation or defense, is typically viewed as being driven by gene duplication in conjunction with positive selection, leading to large families of diversified toxins with selective venom gland expression. Yet, the production of alternative transcripts at venom genes is often overlooked as another potentially important process that could contribute proteins to venom, and requires comprehensive datasets integrating genome and transcriptome sequences together with proteomic characterization of venom to be fully documented. In the common house spider, Parasteatoda tepidariorum, we used RNA sequencing of four tissue types in conjunction with the sequenced genome to provide a comprehensive transcriptome annotation. We also used mass spectrometry to identify a minimum of 99 distinct proteins in P. tepidariorum venom, including at least 33 latrotoxins, pore-forming neurotoxins shared with the confamilial black widow. We found that venom proteins are much more likely to come from multiple transcript genes, whose transcripts produced distinct protein sequences. The presence of multiple distinct proteins in venom from transcripts at individual genes was confirmed for eight loci by mass spectrometry, and is possible at 21 others. Alternative transcripts from the same gene, whether encoding or not encoding a protein found in venom, showed a range of expression patterns, but were not necessarily restricted to the venom gland. However, approximately half of venom protein encoding transcripts were found among the 1,318 transcripts with strongly venom gland biased expression. Our findings revealed an important role for alternative transcription in generating venom protein complexity and expanded the traditional model of venom evolution

Monocoque structure for the SKITTER three-legged walker

The SKITTER 2 design is a monocoque version of the proposed lunar three-legged walker. By the definition of monocoque, the body and legs are a shell with no internal ribbing or supports added for absorbing stresses. The purpose of the monocoque is to encase the elements used for power transmission, power supply, and control of the motion. The material for the structure is a vinyl ester resin, Derakane 8084. This material is easily formable and locally obtainable. The body consists of a hexagonally shaped cylinder with truncated hexagonal pyramids on the top and botton. The legs are eight inch diameter cylinders. The legs are comprised of a tibia section and a femur section. The SKITTER 2 is powered by six actuators which provide linear forces that are transformed into rotary torques by a series of chains and sprockets. The joints connect the femur to the body and the tibia to the femur. Surrounding the joints are flexible rubber hoses that fully encase the chains and sprockets. The SKITTER 2 is capable of walking upside down, righting itself after being overturned, and has the ability to perform in many environments. Applications for this walker include lunar transport or drilling, undersea exploration, and operation in severe surroundings such as arctic temperatures or high radiation

Wireless, battery-free optoelectronic systems as subdermal implants for local tissue oximetry

Monitoring regional tissue oxygenation in animal models and potentially in human subjects can yield insights into the underlying mechanisms of local O2-mediated physiological processes and provide diagnostic and therapeutic guidance for relevant disease states. Existing technologies for tissue oxygenation assessments involve some combination of disadvantages in requirements for physical tethers, anesthetics, and special apparatus, often with confounding effects on the natural behaviors of test subjects. This work introduces an entirely wireless and fully implantable platform incorporating (i) microscale optoelectronics for continuous sensing of local hemoglobin dynamics and (ii) advanced designs in continuous, wireless power delivery and data output for tether-free operation. These features support in vivo, highly localized tissue oximetry at sites of interest, including deep brain regions of mice, on untethered, awake animal models. The results create many opportunities for studying various O2-mediated processes in naturally behaving subjects, with implications in biomedical research and clinical practice.Center for Bio-Integrated Electronics at Northwestern University; Soft and Hybrid Nanotechnology Experimental (SHyNE) Resource (NSF) [ECCS-1542205]; Materials Research Science and Engineering Center [DMR-1720139]; State of Illinois; Northwestern University; Developmental Therapeutics Core at Northwestern University; Robert H. Lurie Comprehensive Cancer Center [NCI CA060553]Open access journalThis item from the UA Faculty Publications collection is made available by the University of Arizona with support from the University of Arizona Libraries. If you have questions, please contact us at [email protected]