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Transgenic technology for mosquitoes is now more than two decades old, and a wide array of control sequences have been described for regulating gene expression in various life stages or specific tissues. Despite this, comparatively little attention has been paid to the development and validation of other transgene-regulating elements, especially 3ʹ-untranslated regions (3ʹUTRs). As a consequence, the same regulatory sequences are often used multiple times in a single transgene array, potentially leading to instability of transgenic effector genes. To increase the repertoire of characterized 3ʹUTRs available for genetics-based mosquito control, we generated fifteen synthetic sequences based on the base composition of the widely used SV40 3ʹUTR sequence, and tested their ability to contribute to the expression of reporter genes EGFP or luciferase. Transient transfection in mosquito cells identified nine candidate 3ʹUTRs that conferred moderate to strong gene expression. Two of these were engineered into the mosquito genome through CRISPR/Cas9-mediated site-specific insertion and compared to the original SV40 3ʹUTR. Both synthetic 3ʹUTRs were shown to successfully promote transgene expression in all mosquito life stages (larva, pupa and adults), similar to the SV40 3ʹUTR, albeit with differences in intensity. Thus, the synthetic 3ʹUTR elements described here are suitable for regulating transgene expression in Ae. aegypti, and provide valuable alternatives in the design of multi-gene cassettes. Additionally, the synthetic-scramble approach we validate here could be used to generate additional functional 3ʹUTR elements in this or other organisms.

Background. Bahia de la Paz, Mexico have seasonal blooms of planktonic prey that support a range of marine species including whale sharks (Rhincodon typus). Goals. This study includes the first sampling cycle that covers all climatic seasons, to know changes in the abundance and composition of zooplankton in the coastal waters off the El Mogote Sandbar, in Bahia de La Paz, BCS, Mexico. Methods. A total of 96 plankton samples were collected at predetermined field stations, spanning the entire length of the El Mogote from June 2016 to May 2017. Results. Total biomass was significantly different between seasons (Kruskal-Wallis chi-squared = 29.993, p = 0.000001385), with the lowest biomass recorded in spring (176.28 ml 100 m-3, 10.1%); and highest observed for winter (703.04 ml 100 m-3, 40.4%). Zooplankton composition was largely comprised of two main taxonomic groups: Copepoda (396,234 organisms100 m-3, 43.1%) and Chaetognatha (196,986 organisms100 m-3, 21.4%). The total number of individuals (sum of all taxonomic groups) per season showed significant differences (Kruskal-Wallis chi-squared = 29.78, p = 0.000001533) with the highest concentrations recorded in winter (292,429 organisms100 m-3, 31.8%) and lowest in spring (162,995 organisms100 m-3; 17.7%). Conclusions. This information provides the first work focused solely in the El Mogote area as a main foraging site for whale sharks, with clear changes in plankton communities with respect to shifting climatic seasons, and the importance of certain taxonomic groups to the diet of whale sharks.

With the size of the biopharmaceutical market exponentially increasing, there is an aligned growth in the importance of data-rich analyses, not only to assess drug product safety but also to assist drug development driven by the deeper understanding of structure/function relationships. In monoclonal antibodies, many functions are regulated by N-glycans present in the constant region of the heavy chains and their mechanisms of action are not completely known. The importance of their function focuses analytical research efforts on the development of robust, accurate and fast methods to support drug development and quality control. Released N-glycan analysis is considered as the gold standard for glycosylation characterisation; however, it is not the only method for quantitative analysis of glycoform heterogeneity. In this study, ten different analytical workflows for N-glycan analysis were compared using four monoclonal antibodies. While observing good comparability between the quantitative results generated, it was possible to appreciate the advantages and disadvantages of each technique and to summarise all the observations to guide the choice of the most appropriate analytical workflow according to application and the desired depth of data generated., Graphical abstract Image 1, Highlights • Comparative analysis of N-glycans present on four monoclonal antibodies using ten analytical methods reported. • Data presented to allow for impartial selection of appropriate analytical strategy depending on depth of information desired and ease of approach. • Methods included intact and subunit strategies and glycopeptide characterisation compared to released N-glycan as the current gold standard.

Monoclonal antibodies (mAbs) and related products undergo a wide range of modifications, many of which can often be directly associated to culture conditions during upstream processing. Ideally, such conditions should be monitored and fine-tuned based on real-time or close to real-time information obtained by the assessment of the product quality attribute (PQA) profile of the biopharmaceutical produced, which is the fundamental idea of process analytical technology. Therefore, methods that are simple, quick and robust, but sufficiently powerful, to allow for the generation of a comprehensive picture of the PQA profile of the protein of interest are required. A major obstacle for the analysis of proteins directly from cultures is the presence of impurities such as cell debris, host cell DNA, proteins and small-molecule compounds, which usually requires a series of capture and polishing steps using affinity and ion-exchange chromatography before characterization can be attempted. In the current study, we demonstrate direct coupling of protein A affinity chromatography with native mass spectrometry (ProA-MS) for development of a robust method that can be used to generate information on the PQA profile of mAbs and related products in as little as 5 min. The developed method was applied to several samples ranging in complexity and stability, such as simple and more complex monoclonal antibodies, as well as cysteine-conjugated antibody-drug conjugate mimics. Moreover, the method demonstrated suitability for the analysis of protein amounts of

Mosquito gene editing has become routine in several laboratories with the establishment of systems such as transcription-activator-like effector nucleases (TALENs), zinc-finger nucleases (ZFNs), and homing endonucleases (HEs). More recently, clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) technology has offered an easier and cheaper alternative for precision genome engineering. Following nuclease action, DNA repair pathways will fix the broken DNA ends, often introducing indels. These out-of-frame mutations are then used for understanding gene function in the target organisms. A drawback, however, is that mutant individuals carry no dominant marker, making identification and tracking of mutant alleles challenging, especially at scales needed for many experiments. High-resolution melt analysis (HRMA) is a simple method to identify variations in nucleic acid sequences and utilizes PCR melting curves to detect such variations. This post-PCR analysis method uses fluorescent double-stranded DNA-binding dyes with instrumentation that has temperature ramp control data capture capability and is easily scaled to 96-well plate formats. Described here is a simple workflow using HRMA for the rapid detection of CRISPR/Cas9-induced indels and the establishment of mutant lines in the mosquito Ae. aegypti. Critically, all steps can be performed with a small amount of leg tissue and do not require sacrificing the organism, allowing genetic crosses or phenotyping assays to be performed after genotyping.

Ovaries of bluefin tuna, Thunnus thynnus, from organisms measuring 247.62 ± 3.54 cm fork length collected within the Mexican Exclusive Economic Zone in the Gulf of Mexico, were examined. Th...

Migration is a critical strategy in maintaining populations, and pathways used by individuals lend insight into habitat quality and connectivity. Yet sustaining migration among large-ranging wildlife poses a challenge for conservation, particularly among landscapes that include a diverse matrix of land tenure. Such is the case in the Northern Great Plains (NGP), a sagebrush (Artemisia spp.) steppe and grassland ecosystem that is home to the longest-ever recorded migrations by both pronghorn (Antilocapra americana) and greater sage-grouse (Centrocercus urophasianus). Here, we identify migratory pathways for both species, and measure the ability of current conservation and policy to maintain cross-taxa migration in the face of continued cultivation. Migratory behavior was similar between species in their timing and duration of migration, and in their use of stopovers along the way. Large and intact private and public working lands largely underpinned migratory pathways, whereas protected areas provided another 5% of habitats. Most pathways for sage-grouse were within state- and federally-designated sage-grouse Core Areas, which contain regulatory caps on anthropogenic disturbance on public lands and help guide conservation efforts; these benefits extended to over half of pathways used by pronghorn. Among private lands, both species largely migrated through intact grazing lands, including many that were already perpetually protected from cultivation with conservation easements. Optimization of remaining private parcels provides managers with a spatial tool to prioritize private-lands conservation, and suggests that comprehensive conservation of shared migratory pathways for pronghorn and sage-grouse in the NGP is within reach of completion given the ongoing pace of conservation.

Genetics-based pest management processes, including the sterile insect technique, are an effective method for the control of some pest insects. However, current SIT methods are not directly transferable to many important pest insect species due to the lack of genetic sexing strains. Genome editing is revolutionizing the way we conduct genetics in insects, including in Tribolium castaneum, an important genetic model and agricultural pest. We identified orthologues of β2Tubulin, Rad50-ATPase and enolase in T. castaneum. Using RT-PCR, we confirmed that these genes are predominantly expressed in the testis. PiggyBac-based transformation of T. castaneum cis-regulatory regions derived from Tc-β2t, Tc-rad50 or Tc-eno resulted in EGFP expression specifically in the T. castaneum testis. Additionally, we determined that each of these regulatory regions regulates EGFP expression in different cell types of the male gonad. Cis-regulatory regions from Tc-β2t produced EGFP expression throughout spermatogenesis and also in mature sperms; Tc-rad50 resulted in expression only in the haploid spermatid, while Tc-eno expressed EGFP in late spermatogenesis. In summary, the regulatory cis-regions characterized in this study are not only suited to study male gonadal function but could be used for development of transgenic sexing strains that produce one sex in pest control strategies.

Among the most economically significant agricultural crops, the species of the Brassicaceae family stand out as one of the most popular in human food. Cabbage leaf (Brassica oleracea var. acephala) is an essential ingredient in many regional dishes and is an excellent nutritional source for children, young and old. However, Brazilian production has been suffering from the incidence of pests and diseases that affect the crop, being the black rot of the cruciferous (BRC), the main cause of economic losses of its producers. Given this context, the objective is to build a theoretical framework by reviewing the literature on the agroecological management of BRC in cabbage leaf, based on the contribution of scientific knowledge to the preservation of agrobiodiversity in rural and traditional family communities. Entre los cultivos agrícolas de mayor importancia económica, la especie de la familia Brassicaceae se destaca como una de las más populares en la alimentación humana. La hoja de col (Brassica oleracea var. acephala) es un ingrediente esencial en muchos platos regionales y es una excelente fuente nutricional para niños, jóvenes y mayores. Sin embargo, la producción brasileña viene sufriendo la incidencia de plagas y enfermedades que afectan al cultivo, siendo la podredumbre negra de las crucíferas (PNC), la principal causa de pérdidas económicas de sus productores. Ante este contexto, el objetivo es construir un marco teórico mediante la revisión de la literatura sobre el manejo agroecológico de PNC en hoja de col, basado en el aporte del conocimiento científico a la preservación de la agrobiodiversidad en comunidades familiares rurales y tradicionales. Dentre as culturas agrícolas de maior expressividade econômica, as espécies da família Brassicaceae destacam-se como uma das mais populares na alimentação humana. A couve-manteiga (Brassica oleracea var. acephala) é um ingrediente essencial em diversos pratos regionais, sendo uma excelente fonte nutricional para crianças, jovens e idosos. Entretanto, a produção brasileira vem sofrendo com a incidência de pragas e doenças que acometem a lavoura, sendo a podridão negra das crucíferas (PNC), a principal causa de perdas econômicas de seus produtores. Diante deste contexto, objetiva-se a construção de um arcabouço teórico mediante revisão de literatura sobre o manejo fitotécnico e fitossanitário agroecológico de cultivares de couve comum, na premissa de agregar ao conhecimento científico ecológico na preservação da agrobiodiversidade em comunidades familiares rurais e tradicionais.

We studied the habitat selection of pronghorn (Antilocapra americana) during seasonal migration; an important period in an animal’s annual cycle associated with broad-scale movements. We further decompose our understanding of migration habitat itself as the product of both broad- and fine-scale behavioral decisions and take a multi-scale approach to assess pronghorn spring and fall migration across the transboundary Northern Sagebrush Steppe region. We used a hierarchical habitat selection framework to assess a suite of natural and anthropogenic features that have been shown to influence selection patterns of pronghorn at both broad (migratory neighborhood) and fine (migratory pathway) scales. We then combined single-scale predictions into a scale-integrated step selection function (ISSF) map to assess its effectiveness in predicting migration route habitat. During spring, pronghorn selected for native grasslands, areas of high forage productivity (NDVI), and avoided human activity (i.e., roads and oil and natural gas wells). During fall, pronghorn selected for native grasslands, larger streams and rivers, and avoided roads. We detected avoidance of paved roads, unpaved roads, and wells at broad spatial scales, but no response to these features at fine scales. In other words, migratory pronghorn responded more strongly to anthropogenic features when selecting a broad neighborhood through which to migrate than when selecting individual steps along their migratory pathway. Our results demonstrate that scales of migratory route selection are hierarchically nested within each other from broader (second-order) to finer scales (third-order). In addition, we found other variables during particular migratory periods (i.e., native grasslands in spring) were selected for across scales indicating their importance for pronghorn. The mapping of ungulate migration habitat is a topic of high conservation relevance. In some applications, corridors are mapped according to telemetry location data from a sample of animals, with the assumption that the sample adequately represents habitat for the entire population. Our use of multi-scale modelling to predict resource selection during migration shows promise and may offer another relevant alternative for use in future conservation planning and land management decisions where telemetry-based sampling is unavailable or incomplete.

Raw milk is a food with great consumption and economic value in Brazil. However, is susceptible of contamination by pathogenic bacteria. The aimed of this study was to evaluate the quality of in natura milk based on microbiological in three dairy farms, somatic cells counting (SCC), bacterial counting and his physical-chemical composition. Were made the following microbiological analysis: counting of mesophilic and psychrotrophic bacteria, coliforms at 30 ºC, coliforms at 45 ºC, Staphylococcus spp., Listeria spp., and SCC. The physical-chemical analysis was fat, protein, lactose, total solids, urea, and casein. There was no evidence of Salmonella spp. and Escherichia coli were identified in any samples. In accordance to the microbiological standards established by Normative Instruction 76 only coliforms 30 ºC and 45 ºC counts were above the standards. There was a significant difference (p≤0.05) between the three farms studied regarding most microbiological aspects. Also, was observed difference (p≤0.05) for most of physical-chemical aspects. Overall, the milk produced in the regions of Alagoas State fails to meet just a constant criterion in the current legislation. La leche cruda es un alimento de gran consumo y valor económico en Brasil. Sin embargo, es susceptible a la contaminación por bacterias patógenas. El objetivo de este estudio fue evaluar la calidad de la leche in natura en base al recuento de células somáticas (CCS), el recuento de bacterias y su composición físico-química. Se realizaron los siguientes análisis microbiológicos: recuento de bacterias mesófilas y psicotróficas, coliformes a 30 ºC, coliformes a 45 ºC, Staphylococcus spp., Listeria spp. y CCS. Los análisis físico-químicos fueron de grasa, proteína, lactosa, sólidos totales, urea y caseína. No se identificaron pruebas de Salmonella spp. y Escherichia coli en ninguna muestra. De acuerdo con las normas microbiológicas establecidas por la Instrucción Normativa Nº 76, sólo los recuentos de coliformes a 30 ºC y 45 ºC estaban por encima de las normas. Hubo una diferencia significativa (p≤0,05) entre las tres granjas estudiadas en cuanto a la mayoría de los aspectos microbiológicos. Además, se observó una diferencia (p≤0,05) en la mayoría de los aspectos físico-químicos. En general, la leche producida en las regiones del estado de Alagoas no puede cumplir con un solo criterio de la legislación vigente. O leite cru é um alimento com grande consumo e valor econômico no Brasil. No entanto, é susceptível de contaminação por bactérias patogênicas. O objetivo deste estudo foi avaliar a qualidade do leite in natura com base em contagem de células somáticas (CCS), contagem bacteriana e sua composição físico-química. Foram feitas as seguintes análises microbiológicas: contagem de bactérias mesofílicas e psicrotróficas, coliformes a 30 ºC, coliformes a 45 ºC, Staphylococcus spp., Listeria spp. e CCS. As análises físico-químicas foram gordura, proteína, lactose, sólidos totais, uréia e caseína. Não houve evidência de Salmonella spp. e Escherichia coli em nenhuma amostra. De acordo com os padrões microbiológicos estabelecidos pela Instrução Normativa nº 76, apenas as contagens de coliformes a 30 ºC e 45 ºC estavam acima dos padrões. Houve uma diferença significativa (p≤0,05) entre as três fazendas estudadas quanto à maioria dos aspectos microbiológicos. Além disso, foi observada diferença (p≤0,05) para a maioria dos aspectos físico-químicos. Em geral, o leite produzido nas regiões do estado de Alagoas não consegue atender apenas a um critério constante da legislação vigente.

The survey of new benthic substrata is expected to yield new records of diatom taxa. A particular type of substratum is the skin of the Stone scorpionfish Scorpaena mystes Jordan & Starks, 1895 that, because of its benthic form of life, represents a potential colonizing surface for various organisms, including epizoic and opportunistic diatom taxa (epibiotic community). Thus, a floristic survey of diatoms was carried out by sampling the epibiotic community from S. mystes specimens collected in the east coast of Baja California Sur (Gulf of California). Thirty diatoms are new records for Mexican littorals. Twelve of these taxa had not been hitherto recorded for American coasts. Micrographs, and morphometric and distribution data on these taxa are provided.

The whale shark is the world’s largest filter feeding shark and holds international protection as an endangered species. It is known to form seasonal aggregations linked to blooms of zooplanktonic organisms. The aim of this study was to investigate whether whale shark foraging behaviors are related to the density and/or concentration of available prey items. Zooplankton samples were obtained between October-2016 and March-2017 from Bahia de La Paz in the absence (n =32) and presence (n = 12) of foraging whale sharks. Results indicated that zooplankton biomass was 1.6 times significantly greater (t = -3.21, p 0.05). The dominant taxonomic groups were copepods and chaetognaths with no significant differences found in the mean number of copepods (t = -1.81, p > 0.05), chaetognaths (t = -1.70, p > 0.05), euphausiids (t = -0.12, p > 0.05) and decapods (t = -1.80, p > 0.05), between the presence and absence of feeding whale sharks. In conclusion, whale sharks feeding behaviors occur during higher concentrations of zooplankton biomass and the species may potentially target dense patches of both copepods and chaetognaths.

Charge sensitive separation methods such as ion exchange chromatography (CEX) and capillary electrophoresis (CE) have recently been coupled to mass spectrometry to facilitate high resolution profiling of proteoforms present within the charge variant profile of complex biopharmaceuticals. Here we apply pH gradient cation exchange chromatography and microfluidic capillary electrophoresis using the ZipChip platform for comparative characterization of the monoclonal antibody Cetuximab. Cetuximab harbors four glycans per molecule, two on each heavy chain, of which the Fab glycans have been reported to be complex and multiply sialylated. The presence of these extra glycosylation sites in the variable region of the molecule causes significant charge variant and glycan heterogeneity, which makes comprehensive analysis on the intact protein level challenging. Both pH gradient CEX-MS and CE-MS were found to be powerful for the separation of Cetuximab charge variants with eight major peaks being baseline resolved using both separation platforms. Informative native-like mass spectra were collected for each charge variant peak using both platforms that facilitated deconvolution and further analysis. The total proteoform coverage was exceptionally high with >100 isoforms identified and relatively quantified with CEX-MS, in case of CE-MS the proteoform coverage was >200. A deep insight into the heterogeneity of Cetuximab was unveiled, the high level of sensitivity achievable enables the implementation of the presented technologies even at early stages of the biopharmaceutical development platform, such as in developability assessment, process development and also for monitoring process consistency.

Length-weight relationships (LWRs) were estimated for Pacific blue marlin ( Makaira nigricans ), striped marlin ( Kajikia audax ), sailfish ( Istiophorus platypterus ), dolphinfish ( Coryphaena hippurus ) and yellowfin tuna ( Thunnus albacares ). These five fish species are key top predators of the pelagic ecosystem and are economically important resources in the sport fishing of Baja California Sur, Mexico. Overall, 53,359 length and weight records were analyzed for the period 1990-2015. LWRs were significant ( P < 0.05), with b -values ranging from 2.52 to 3.13.

The roosterfish is a coastal pelagic species that inhabits shallow waters of the Eastern Pacific Ocean. Despite its ecological and economic importance within the region, there are few studies on its basic biology, especially those focused on age and individual growth. Therefore, the objective of this study was to estimate roosterfish age and individual growth parameters through counting growth increments of sagittal otoliths. The work focused on organisms caught off Baja California Sur, Mexico (BCS) by both the sport and artisanal fishing fleets from 2010–2017. A total of 266 organisms were sampled during the study period, ranging in size from 5.2–133 cm fork length. A model was generated to describe the relationship between otolith radius (OR) and fork length (FL) of the fish, where FL = -43.3 + 44.59 OR. The periodicity of otolith growth increment formation was annual, and this study found roosterfish age estimates to range from 0 to 8 years old, with most individuals ranging from 1–2 years of age. Based on the fitting of the data to three individual growth models (Gompertz, Logistic and von Bertalanffy, VBGF), the VBGF best fit the individual growth curve for roosterfish. The VBGF parameter of L ∞ = 131.53, k = 0.35 y t 0 = -0.23 supports previous studies and suggests that roosterfish are very fast growing in the first years of life.

The exposure of wood to elevated temperatures during hot water extraction (HWE) has been shown to significantly influence a variety of physical and chemical properties of the wood. Aspen (Populus tremuloides) and hemlock (Tsuga canadensis) strands underwent HWE at 160 °C for four different periods of time. The weight loss (WL) associated with the processes ranged from 2.9 to 17.3% for the aspen, and from 2.9 to 12.8% for the hemlock. The higher WL associated with the hardwood was attributed to higher hemicellulose content in the hardwood than in the softwood as well as differences in chemical composition. Substantial increases in cell wall hardness (up to 34%) and modulus of elasticity (MOE) (up to 28%) were obtained at the intermediate extraction conditions. Extended exposure conditions resulted in decreases. The axial MOE of strands showed significant differences only in aspen. These changes are hypothesized to be related, in part, to changes in cellulose crystallinity.

Few studies have examined differential responses of partially migratory ungulates to human development or activity, where some individuals in a population migrate and others do not. Yet understanding how animals with different movement tactics respond to anthropogenic disturbance is key to sustaining global ungulate migrations. We examined seasonal resource selection of a partially migratory population of pronghorn (Antilocapra americana) in the Northern Sagebrush Steppe of Alberta, Saskatchewan, and Montana from 2003 to 2011. We developed step‐selection functions (SSF) for migrant and resident pronghorn during the summer and winter at two spatial scales (second order and third order) and then integrated SSFs across scales to estimate pronghorn responses to fences and subsequent habitat loss from these features while accounting for responses to other resource use. Both migrant and resident pronghorn showed the strongest responses to natural and anthropogenic features at the second order and weaker responses at the third order. Selection responses of migrant and residents differed the most in response to normalized difference vegetation index, topography, and anthropogenic features. Seasonally, selection for intermediate greenness was strongest in summer, whereas avoidance of roads strongly influenced winter resource selection of both tactics. Both migrant and resident pronghorn showed strong avoidance of fencing at both spatial scales during summer and winter. Model predictions with complete removal of fences from the landscape (i.e., natural conditions) predicted an increase in the area of high‐quality habitat of 16–38%. In contrast, doubling fence density on the landscape decreased the amount of high‐quality habitat by 1–11% and increased low‐quality habitat by 13–21%. Our results suggest that pronghorn winter and summer ranges can be improved by reducing the density of fences on the landscape, or mitigation measures to enhance fence crossings, to alleviate the indirect loss of habitat for this important endemic prairie species.

Fibrosis, the pathological excess of fibroblast activity, is a significant health issue that hinders the function of many organs in the body, in some cases fatally. However, the severity of fibrosis-derived conditions depends on both the positioning of fibrotic affliction, and the microscopic patterning of fibroblast-deposited matrix proteins within afflicted regions. Variability in an individual’s manifestation of a type of fibrosis is an important factor in explaining differences in symptoms, optimum treatment and prognosis, but a need forex vivoprocedures and a lack of experimental control over conflating factors has meant this variability remains poorly understood. In this work, we present a computational methodology for the generation of patterns of fibrosis microstructure, demonstrating the technique using histological images of four types of cardiac fibrosis. Our generator and automated tuning method prove flexible enough to capture each of these very distinct patterns, allowing for rapid generation of new realisations for high-throughput computational studies. We also demonstrate via simulation, using the generated fibrotic patterns, the importance of micro-scale variability by showing significant differences in electrophysiological impact even within a single class of fibrosis.

Aedes aegypti is a critical vector for transmitting Zika, dengue, chikungunya, and yellow fever viruses to humans. Genetic strategies to limit mosquito survival based upon sex distortion or disruption of development may be valuable new tools to control Ae. aegypti populations. We identified six genes with expression limited to pupal development ; osi8 and osi11 (Osiris protein family), CPRs and CPF (cuticle protein family), and stretchin (a muscle protein). Heritable CRISPR/Cas9-mediated gene knockout of these genes did not reveal any defects in pupal development. However, stretchin-null mutations (strnΔ35/Δ41) resulted in flightless mosquitoes with an abnormal open wing posture. The inability of adult strnΔ35/Δ41 mosquitoes to fly restricted their escape from aquatic rearing media following eclosion , and substantially reduced adult survival rates. Transgenic strains which contain the EGFP marker gene under the control of strn regulatory regions (0.8 kb, 1.4 kb, and 2.2 kb upstream, respectively), revealed the gene expression pattern of strn in muscle-like tissues in the thorax during late morphogenesis from L 4 larvae to young adults. We demonstrated that Ae. aegypti pupae-specific strn is critical for adult mosquito flight capability and a key late-acting lethal target for mosquito-borne disease control.

Brown rot fungi utilize iron as part of a chelator-mediated Fenton (CMF) reaction during wood biodegradation. Research suggests these fungi reduce Fe3+ to promote oxygen radical generation resulting in depolymerization of the wood cell wall. High levels of Mn are also found in wood decayed by brown rot fungi. However, little is known about the oxidation states of Fe and Mn during the decay process. X-ray absorption near edge spectroscopy (XANES) can be used to examine metal oxidation states and coordination chemistry. XANES experiments were conducted on wood decayed by Gloeophyllum trabeum over 2–8 weeks with results showing that Mn2+ and Fe3+ predominated for metal oxidation states. However, Fe2+ was present at sites of greater fungal growth In certain cases, the μXANES measurements showed that the fraction of Fe2+ in the wood samples was as high as 50%. Localized areas of reduced iron corresponded with areas of greater fungal hyphal mass which is in agreement with how brown rot fungi decay wood via the CMF reaction. The limited change in oxidation state of Mn observed in wood with active fungal activity suggests that the role of manganese in CMF biodegradation chemistry should be further explored.

Nearly 400 million years of evolution and field-testing by the natural world has given humans thousands of wood types, each with unique structure–property relationships to study, exploit, and ideally, to manipulate, but the slow growth of trees makes them a recalcitrant experimental system. Variations in wood features of two genotypes of peach (Prunus persica L.) trees, wild-type and crinkle-leaf, were examined to elucidate the nature of weak wood in crinkle-leaf trees. Crinkle-leaf is a naturally-occurring mutation in which wood strength is altered in conjunction with an easily observed ‘crinkling’ of the leaves’ surface. Trees from three vigor classes (low growth rate, average growth rate, and high growth rate) of each genotype were sampled. No meaningful tendency of dissimilarities among the different vigor classes was found, nor any pattern in features in a genotype-by-vigor analysis. Wild-type trees exhibited longer vessels and fibers, wider rays, and slightly higher specific gravity. Neither cell wall mechanical properties measured with nanoindentation nor cell wall histochemical properties were statistically or observably different between crinkle-leaf and wild-type wood. The crinkle-leaf mutant has the potential to be a useful model system for wood properties investigation and manipulation if it can serve as a field-observable vegetative marker for altered wood properties.

Background. In recent years (since 2012) there has been an increase in the commercial exploitation of Scorpaena mystes Jordan & Starks, 1985 in the central area of the Gulf of California and there is no information on the basic biology of this species that serve as a basis to determine the current population status and, in the future, assess any changes associated with exploitation and environmental factors. Goals. Determine the length-weight (LWR) and length-length (LLR) relationships, and the relative condition factor of S. mystes. Methods. Fish specimens (n = 258) were collected monthly from May 2015 through April 2016 in waters off Santa Rosalia, Baja California Sur. Each fish was measured for standard length (SL) and total length (TL), and weighed (W). Results. LWR and LLR were highly significant (p

Un especimen herido de Carcharodon carcharias : examen prospectivo del costo de sus cacerias Aunque culturalmente se percibe al gran tiburon blanco ( Carcharodon carcharias ) como un depredador invulnerable, en el presente se reporta la muerte de un especimen sub-adulto atrapado en una red de pesca; dicho especimen se encontraba herido por la espina de una raya pelagica que penetro la columna vertebral del tiburon, lo que posiblemente mermo su capacidad para nadar. El evento sugiere que los tiburones blancos pueden sufrir lesiones e incluso muerte, mientras cazan presas peligrosas; ello genera preguntas del como los individuos de la especie toman decisiones de caza. Las teorias de “Forrajeo optimo” y “Busqueda de recursos con riesgo de depredacion” combinadas con tecnologia de seguimiento, permitirian responder este tipo de preguntas.

The mechanisms by which colicins, protein toxins produced by Escherichia coli, kill other E. coli, have become much better understood in recent years. Most colicins initially bind to an outer membrane protein receptor, and then search for a separate nearby outer membrane protein translocator that serves as a pathway into target cells. Many colicins use the outer membrane porin, OmpF, as that translocator, while using a different primary receptor. Colicin N is unique among known colicins in that only OmpF had been identified as being required for uptake of the colicin and it was presumed to somehow serve as both receptor and translocator. Genetic screens also identified a number of genes required for lipopolysaccharide (LPS) synthesis as uniquely required for killing by colicin N, but not by other colicins. In this issue of Molecular Microbiology, Johnson et al. show that the receptor-binding domain of colicin N binds to LPS, and does not require OmpF for that binding. LPS of a minimal length is required for binding, explaining the requirement for specific elements of the LPS biosynthetic pathway. For colicin N, the receptor-binding domain does not recognize a protein, but rather the most abundant component of the outer membrane itself, LPS.

The role of ions in the fungal decay process of lignocellulose biomaterials, and more broadly fungal metabolism, has implications for diverse research disciplines ranging from plant pathology and forest ecology, to carbon sequestration. Despite the importance of ions in fungal decay mechanisms, the spatial distribution and quantification of ions in lignocellulosic cell walls and fungal hyphae during decay is not known. Here we employ synchrotron-based X-ray fluorescence microscopy (XFM) to map and quantify physiologically relevant ions, such as K, Ca, Mn, Fe, and Zn, in wood being decayed by the model brown rot fungus Serpula lacrymans. Two-dimensional XFM maps were obtained to study the ion spatial distributions from mm to submicron length scales in wood, fungal hyphae with the dried extracellular matrix (ECM) from the fungus, and Ca oxalate crystals. Three-dimensional ion volume reconstructions were also acquired of wood cell walls and hyphae with ECM. Results show that the fungus actively transports some ions, such as Fe, into the wood and controls the distribution of ions at both the bulk wood and cell wall length scales. These measurements provide new insights into the movement of ions during decay and illustrate how synchrotron-based XFM is uniquely suited study these ions.

Introduction: Use of thrombolysis in the management of pulmonary embolism (PE) is currently based on the European Society of Cardiology guidelines which use shock and right ventricular (RV) dysfunction as indications (Konstantinides, S.V. et al . European Heart Journal 2014;35:3033-3080). Thrombolysis in the absence of cardiovascular compromise remains controversial. Aim: To identify whether respiratory failure should be an indication for thrombolysis in PE. Methods: A retrospective single-centre cohort study. PE cases were identified from CT pulmonary angiogram reports over a 12 month period. Thrombolysed cases were identified from medical records. Results: 280 cases of PE were diagnosed between 01/12/2014-30/11/2015. 7 were thrombolysed. At the time of thrombolysis, mean respiratory rate was 28 (data incomplete for 2). 6 had arterial blood gas sampling, revealing type 1 respiratory failure in all. PaO 2 was 100bpm. 86% had CT or echocardiographic evidence of RV dysfunction. Serum troponin was raised in 71%. 3 cases died within 30 days of thrombolysis. The remaining cases survived to discharge and remain alive at the time of reporting (at least 3 months survival). Conclusions: Current guidelines exclusively use cardiovascular parameters to guide thrombolysis in PE. In this study, more than half of the thrombolysed cases were not hypotensive whilst all were hypoxic and tachypnoeic. Our experience suggests that greater emphasis should be placed on markers of respiratory compromise when risk stratifying in PE and that further research is needed to guide clinicians9 use of thrombolysis outside of currently accepted scenarios.

Colicins are protein toxins made by Escherichia coli to kill related bacteria that compete for scarce resources. All colicins must cross the target cell outer membrane in order to reach their intracellular targets. Normally, the first step in the intoxication process is the tight binding of the colicin to an outer membrane receptor protein via its central receptor-binding domain. It is shown here that for one colicin, E1, that step, although it greatly increases the efficiency of killing, is not absolutely necessary. For colicin E1, the second step, translocation, relies on the outer membrane/transperiplasmic protein TolC. The normal role of TolC in bacteria is as an essential component of a family of tripartite drug and toxin exporters, but for colicin E1, it is essential for its import. Colicin E1 and some N-terminal translocation domain peptides had been shown previously to bind in vitro to TolC and occlude channels made by TolC in planar lipid bilayer membranes. Here, a set of increasingly shorter colicin E1 translocation domain peptides was shown to bind to Escherichia coli in vivo and protect them from subsequent challenge by colicin E1. A segment of only 21 residues, the “TolC box,” was thereby defined; that segment is essential for colicin E1 cytotoxicity and for binding of translocation domain peptides to TolC. IMPORTANCE The Escherichia coli outer membrane/transperiplasmic protein TolC is normally an essential component of the bacterium's tripartite drug and toxin export machinery. The protein toxin colicin E1 instead uses TolC for its import into the cells that it kills, thereby subverting its normal role. Increasingly shorter constructs of the colicin's N-terminal translocation domain were used to define an essential 21-residue segment that is required for both colicin cytotoxicity and for binding of the colicin's translocation domain to bacteria, in order to protect them from subsequent challenge by active colicin E1. Thus, an essential TolC binding sequence of colicin E1 was identified and may ultimately lead to the development of drugs to block the bacterial drug export pathway.

Of the steps involved in the killing of Escherichia coli by colicins, binding to a specific outer-membrane receptor was the best understood and earliest characterized. Receptor binding was believed to be an indispensable step in colicin intoxication, coming before the less well-understood step of translocation across the outer membrane to present the killing domain to its target. In the process of identifying the translocator for colicin Ia, I created chimaeric colicins, as well as a deletion missing the entire receptor-binding domain of colicin Ia. The normal pathway for colicin Ia killing was shown to require two copies of Cir: one that serves as the primary receptor and a second copy that serves as translocator. The novel Ia colicins retain the ability to kill E. coli, even in the absence of receptor binding, as long as they can translocate via their Cir translocator. Experiments to determine whether colicin M uses a second copy of its receptor, FhuA, as its translocator were hampered by precipitation of colicin M chimaeras in inclusion bodies. Nevertheless, I show that receptor binding can be bypassed for killing, as long as a translocation pathway is maintained for colicin M. These experiments suggest that colicin M, unlike colicin Ia, may normally use a single copy of FhuA as both its receptor and its translocator. Colicin E1 can kill in the absence of receptor binding, using translocation through TolC.

Anthrax toxin consists of three proteins: lethal factor (LF), edema factor (EF), and protective antigen (PA). This last forms a heptameric channel, (PA63)7, in the host cell’s endosomal membrane, allowing the former two (which are enzymes) to be translocated into the cytosol. (PA63)7 incorporated into planar bilayer membranes forms a channel that translocates LF and EF, with the N terminus leading the way. The channel is mushroom-shaped with a cap containing the binding sites for EF and LF, and an ∼100 Å–long, 15 Å–wide stem. For proteins to pass through the stem they clearly must unfold, but is secondary structure preserved? To answer this question, we developed a method of trapping the polypeptide chain of a translocating protein within the channel and determined the minimum number of residues that could traverse it. We attached a biotin to the N terminus of LFN (the 263-residue N-terminal portion of LF) and a molecular stopper elsewhere. If the distance from the N terminus to the stopper was long enough to traverse the channel, streptavidin added to the trans side bound the N-terminal biotin, trapping the protein within the channel; if this distance was not long enough, streptavidin did not bind the N-terminal biotin and the protein was not trapped. The trapping rate was dependent on the driving force (voltage), the length of time it was applied, and the number of residues between the N terminus and the stopper. By varying the position of the stopper, we determined the minimum number of residues required to span the channel. We conclude that LFN adopts an extended-chain configuration as it translocates; i.e., the channel unfolds the secondary structure of the protein. We also show that the channel not only can translocate LFN in the normal direction but also can, at least partially, translocate LFN in the opposite direction.

Filamentous inclusions made of α-synuclein are found in nerve cells and glial cells in a number of human neurodegenerative diseases, including Parkinson disease, dementia with Lewy bodies, and multiple system atrophy. The assembly and spreading of these inclusions are likely to play an important role in the etiology of common dementias and movement disorders. Both α-synuclein and the homologous β-synuclein are abundantly expressed in the central nervous system; however, β-synuclein is not present in the pathological inclusions. Previously, we observed a poor correlation between filament formation and the presence of residues 73–83 of α-synuclein, which are absent in β-synuclein. Instead, filament formation correlated with the mean β-sheet propensity, charge, and hydrophilicity of the protein (global physicochemical properties) and β-strand contiguity calculated by a simple algorithm of sliding averages (local physicochemical property). In the present study, we rendered β-synuclein fibrillogenic via one set of point mutations engineered to enhance global properties and a second set engineered to enhance predominantly β-strand contiguity. Our findings show that the intrinsic physicochemical properties of synucleins influence their fibrillogenic propensity via two distinct but overlapping modalities. The implications for filament formation and the pathogenesis of neurodegenerative diseases are discussed.

Two closely related scaffolds were identified through an uHTS campaign as desirable starting points for the development of Rho-Kinase (ROCK) inhibitors. Here, we describe our hit-to-lead evaluation process which culminated in the rapid discovery of potent leads such as 22 which successfully demonstrated an early in vivo proof of concept for anti-hypertensive activity.

Colicin Ia, a channel-forming bactericidal protein, uses the outer membrane protein, Cir, as its primary receptor. To kill E. coli, it must cross this membrane. The crystal structure of Ia receptor-binding domain bound to Cir, a 22-stranded plugged β-barrel protein, suggests that the plug does not move. Therefore, another pathway is needed for the colicin to cross the outer membrane, but no “second receptor” has ever been identified for TonB-dependent colicins, such as Ia. We show that if the receptor-binding domain of colicin Ia is replaced by that of colicin E3, this chimera effectively kills cells, provided they have the E3 receptor (BtuB), Cir, and TonB. This is consistent with wild-type Ia using one Cir as its primary receptor (BtuB in the chimera) and a second Cir as the translocation pathway for its N-terminal translocation (T) domain and its channel-forming C-terminal domain. Deletion of colicin Ia’s receptor-binding domain results in a protein that kills E. coli, albeit less effectively, provided they have Cir and TonB. We show that purified T-domain competes with Ia and protects E. coli from being killed by it. Thus, in addition to binding to colicin Ia’s receptor-binding domain, Cir also binds weakly to its translocation domain.

We used a geographic information system (GIS) and logistic regression to investigate relationships between geomorphology and Castor canadensis (North American beaver) impoundment of lower-order, blackwater streams of a southeastern landscape. Using GIS, we divided streams into 632 500-m reaches and measured a set of geomorphic variables for each reach. Beavers were most likely to impound stream reaches crossed by roads with a gradient of ≈ 0.6 to 1.2% and watershed sizes of ≈ 2500 ha; reaches with watershed sizes 5000 ha were almost completely avoided. Gradient and road crossings contributed little to discrimination among impounded and unimpounded reaches, suggesting these variables had relatively small influences on beaver impoundment when compared to stream size. Our results indicate that GIS and geomorphic variables can be used to model the impoundment of streams over larger areas (e.g., the proportion of third-order watersheds impounded), but are less accurate at predicting th...

Colicin Ia is a 69 kDa protein that kills susceptible Escherichia coli cells by binding to a specific receptor in the outer membrane, colicin I receptor (70 kDa), and subsequently translocating its channel forming domain across the periplasmic space, where it inserts into the inner membrane and forms a voltage-dependent ion channel. We determined crystal structures of colicin I receptor alone and in complex with the receptor binding domain of colicin Ia. The receptor undergoes large and unusual conformational changes upon colicin binding, opening at the cell surface and positioning the receptor binding domain of colicin Ia directly above it. We modelled the interaction with full-length colicin Ia to show that the channel forming domain is initially positioned 150 A above the cell surface. Functional data using full-length colicin Ia show that colicin I receptor is necessary for cell surface binding, and suggest that the receptor participates in translocation of colicin Ia across the outer membrane.

The Pim family of Ser/Thr kinases has been implicated in the process of lymphomagenesis and cell survival. Known substrates of Pim kinases are few and poorly characterized. In this study we set out to identify novel Pim-2 substrates using the Kinase Substrate Tracking and Elucidation (KESTREL) approach. Two potential substrates, eukaryotic initiation factor 4B (eIF4B) and apoptosis inhibitor 5 (API-5), were identified from rat thymus extracts. Sequence comparison of the Pim-2 kinase phosphorylation sites of eIF4B and mouse BAD, the only other known Pim-2 substrate, revealed conserved amino acids preceding the phosphorylated serine residue. Stepwise replacement of the conserved residues produced a consensus sequence for Pim kinase recognition: RXRHXS. Pim-1 and Pim-2 catalyzed the phosphorylation of this recognition sequence 20-fold more efficiently than the original (K/R-K/R-R-K/R-L-S/T-a; a = small chain amino acid) Pim-1 phosphorylation site. The identification of the novel Pim kinase consensus sequence provides a more sensitive and versatile peptide based assay for screening modulators of Pim kinase activity.

A 3300-base segment of Escherichia coli chromosomal DNA, cloned into pBR322, will complement a methionine auxotroph in which the lesion is a defective methionyl-tRNA synthetase with a much reduced affinity for methionine. Crude extracts of these transformants contain elevated levels of a protein which has a subunit molecular weight of 66000, methionyl-tRNA synthetase aminoacylation activity in vitro and which cross-reacts with anti-(methionyl-tRNA synthetase) antibodies. This polypeptide is very slightly larger than the well-characterised and crystallised tryptic fragment of methionyl-tRNA synthetase. A DNA sequence of 1750 residues at one end of the cloned insert codes for a non-terminated open reading frame in which we can locate a large number of methionyl-tRNA synthetase tryptic and chymotryptic peptides. We have also sequenced 300 nucleotides upstream of this coding segment where we find a large invert repeat in the putative methionyl-tRNA synthetase promoter region.

A novel inhibitor of p38 mitogen-activated protein kinase (p38), CMPD1, identified by high-throughput screening, is characterized herein. Unlike the p38 inhibitors described previously, this inhibitor is substrate selective and noncompetitive with ATP. In steady-state kinetics experiments, CMPD1 was observed to prevent the p38alpha-dependent phosphorylation (K(i)(app) = 330 nM) of the splice variant of mitogen-activated protein kinase-activated protein kinase 2 (MK2a) that contains a docking domain for p38alpha and p38beta, but it did not prevent the phosphorylation of ATF-2 (K(i)(app) > 20 microM). In addition to kinetic studies, isothermal titration calorimetry and surface plasmon resonance experiments were performed to elucidate the mechanism of inhibition. While isothermal titration calorimetry analysis indicated that CMPD1 binds to p38alpha, CMPD1 was not observed to compete with ATP for p38alpha, nor was it able to interrupt the binding of p38alpha to MK2a observed by surface plasmon resonance. Therefore, deuterium exchange mass spectrometry (DXMS) was employed to study the p38alpha.CMPD1 inhibitory complex, to provide new insight into the mechanism of substrate selective inhibition. The DXMS data obtained for the p38alpha.CMPD1 complex were compared to the data obtained for the p38alpha.MK2a complex and a p38alpha.active site binding inhibitor complex. Alterations in the DXMS behavior of both p38alpha and MK2a were observed upon complex formation, including but not limited to the interaction between the carboxy-terminal docking domain of MK2a and its binding groove on p38alpha. Alterations in the D(2)O exchange of p38alpha produced by CMPD1 suggest that the substrate selective inhibitor binds in the vicinity of the active site of p38alpha, resulting in perturbations to regions containing nucleotide binding pocket residues, docking groove residues (E160 and D161), and a Mg(2+) ion cofactor binding residue (D168). Although the exact mechanism of substrate selective inhibition by this novel inhibitor has not yet been disclosed, the results suggest that CMPD1 binding in the active site region of p38alpha induces perturbations that may result in the suboptimal positioning of substrates and cofactors in the transition state, resulting in selective inhibition of p38alpha activity.

Signaling through the CD40 receptor activates diverse molecular pathways in a variety of immune cell types. To study CD40 signaling complexes in B cells, we produced soluble CD40 cytoplasmic domain multimers that translocate across cell membranes and engage intracellular CD40 signaling pathways. As visualized by fluorescence microscopy, rapid transduction of recombinant Antennapedia-isoleucine zipper (Izip)-CD40 cytoplasmic domain fusion protein (Antp-CD40) occurred in both the DND39 B cell line and human tonsillar B cells. Upon cellular entry, Antp-CD40 activated NF-κB-dependent transcription, induced proteolytic processing of p100 to the p52/NF-κB2 subunit, and increased expression of CD80 and CD54 on the surface of B cells. Antp-CD40 transduction of B cells did not, however, activate detectable levels of p38 mitogen-activated protein kinase or c-Jun N-terminal kinase and did not up-regulate CD95 expression. Analysis of Antp-CD40 complexes recovered from transduced B cells revealed that Antp-CD40 associated with endogenous TRAF3 and Ku proteins. Multimerization of Antp-CD40, or extensive clustering of transmembrane CD40, diminished the disruptive effect of the T254A mutation in the TRAF2/3 binding site of the CD40 cytoplasmic domain. Taken together, these results indicate that Antp-CD40 mimics some of the natural CD40 signaling pathways in B cells by assembling partially functional signaling intermediates that do not require plasma membrane localization. We present a novel approach for delivering pre-activated, soluble receptor cytoplasmic domains into cells and recovering intact signaling complexes for molecular analysis.

The discovery of novel 5,7-disubstituted[1,6]naphthyridines as potent inhibitors of Spleen Tyrosine Kinase (SYK) is discussed. The SAR reveals the necessity for a 7-aryl group with preference towards para substitution and that this in combination with 5-aminoalkylamino substituents further improved the potency of the compounds. The initial SAR as well as a survey of the other positions is discussed in detail.

An imidazo[4,5-h]isoquinolin-7,9-dione (1) was identified as an adenosine 5'-triphosphate competitive inhibitor of lck by high throughput screening. Initial structure-activity relationship studies identified the dichlorophenyl ring and the imide NH as important pharmacophores. A binding model was constructed to understand how 1 binds to a related kinase, hck. These results suggested that removing the gem-dimethyl group and flattening the ring would enhance activity. This was realized by converting 1 to the imidazo[4,5-h]isoquinolin-9-one (20), resulting in an 18-fold improvement in potency against lck and a 50-fold increase in potency in a cellular assay.

tau gene mutations cause frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP-17). Here we have used Xenopus oocyte maturation as an indicator of microtubule function. We show that wild-type four-repeat Tau protein inhibits maturation in a concentration-dependent manner, whereas three-repeat Tau has no effect. Of the seven four-repeat Tau proteins with FTDP-17 mutations tested, five (G272V, DeltaK280, P301L, P301S, and V337M) failed to interfere significantly with oocyte maturation, demonstrating a greatly reduced ability to interact with microtubules. One mutant protein (R406W) almost behaved like wild-type Tau, and one (S305N) inhibited maturation more strongly than wild-type Tau. With the exception of R406W, wild-type Tau and all the mutants studied were similarly phosphorylated during the Xenopus oocyte maturation, and this was independent of their effects on this process. Data obtained with R406W and S305N may be related to charge changes (phosphorylation and basic amino acids). Our results demonstrate variable effects of FTDP-17 mutations on microtubules in an intact cell situation. Those findings establish Xenopus oocyte maturation as a system allowing the study of the functional effects of tau gene mutations in a quantitative manner.

Coding region and intronic mutations in the tau gene cause frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP-17). We have previously reported that ABalphaC, a major form of protein phosphatase 2A (PP2A) in brain, binds tightly to tau protein in vitro and is a major tau phosphatase in vivo. Using in vitro assays, we show here that the FTDP-17 mutations G272V, DeltaK280, P301L, P301S, S305N, V337M, G389R, and R406W inhibit by approximately 20-95% the binding of recombinant three-repeat and four-repeat tau isoforms to the ABalphaC holoenzyme and the AC core enzyme of PP2A. Reduction in binding was maximal for tau proteins with the G272V, DeltaK280, and V337M mutations. We also show that tau protein can be specifically coimmunoprecipitated with endogenous PP2A from both rat brain and transfected cell extracts. It is significant that, by using similar coimmunoprecipitation assays, we show that all FTDP-17 mutations tested, including the N279K mutation, alter the ability of tau to associate with cellular PP2A. Taken together, these results indicate that FTDP-17 mutations induce a significant decrease in the binding affinity of tau for PP2A in vivo. We propose that altered protein-protein interactions between PP2A and tau may contribute to FTDP-17 pathogenesis.