Genetic Encoding of Phosphorylated Amino Acids into Proteins.

Reversible phosphorylation is a fundamental mechanism for controlling protein function. Despite the critical roles phosphorylated proteins play in physiology and disease, our ability to study individual phospho-proteoforms has been hindered by a lack of versatile methods to efficiently generate homogeneous proteins with site-specific phosphoamino acids or with functional mimics that are resistant to phosphatases. Genetic code expansion (GCE) is emerging as a transformative approach to tackle this challenge, allowing direct incorporation of phosphoamino acids into proteins during translation in response to amber stop codons. This genetic programming of phospho-protein synthesis eliminates the reliance on kinase-based or chemical semisynthesis approaches, making it broadly applicable to diverse phospho-proteoforms. In this comprehensive review, we provide a brief introduction to GCE and trace the development of existing GCE technologies for installing phosphoserine, phosphothreonine, phosphotyrosine, and their mimics, discussing both their advantages as well as their limitations. While some of the technologies are still early in their development, others are already robust enough to greatly expand the range of biologically relevant questions that can be addressed. We highlight new discoveries enabled by these GCE approaches, provide practical considerations for the application of technologies by non-GCE experts, and also identify avenues ripe for further development.

An impact evaluation of conservation investments targeting long-distance migratory species.

We evaluated the impact of a philanthropic program investing in the conservation of sites along the Pacific Americas Flyway, which spans >16,000 km of coastline and is used by millions of shorebirds. Using a quasi-experimental, mixed methods approach, we estimated what would have happened to shorebird populations at 17 wintering sites without the sustained and additional investment they received. We modeled shorebird populations across the entire flyway and at sites with and without investment. Combining shorebird abundance estimates with a land-cover classification model, we used the synthetic control method to create counterfactuals for shorebird trends at the treatment sites. We found no evidence of an overall effect across three outcome variables. Species- and site-level treatment effects were heterogeneous, with a few cases showing evidence of a positive effect, including a site with a high level of overall investment. Results suggest six shorebirds declined across the entire flyway, including at many Latin American sites. However, the percentage of flyway populations present at the sites remained stable, and the percentage at the treatment sites was higher (i.e., investment sites) than at control sites. Multiple mechanisms behind our results are possible, including that investments have yet to mitigate impacts and negative impacts at other sites are driving declines at the treatment sites. A limitation of our evaluation is the sole focus on shorebird abundance and the lack of data that prohibits the inclusion of other outcome variables. Monitoring infrastructure is now in place to design a more robust and a priori shorebird evaluation framework across the entire flyway. With this framework, it will prove easier to prioritize limited dollars to result in the most positive conservation outcomes.

Evaluación del impacto de la inversión para la conservación enfocada en especies migratorias de largo recorrido Resumen Evaluamos el impacto de un programa filantrópico que invierte en la conservación de sitios a lo largo de la Ruta Migratoria Pacífico-Américas, la cual abarca >16,000 km de la línea costera y millones de aves playeras la usan. Estimamos con una estrategia cuasiexperimental y de métodos mixtos lo que habría pasado con las poblaciones de estas aves en 17 sitios invernales sin la inversión adicional y continua que recibieron. Modelamos estas poblaciones en toda la ruta y en sitios con y sin inversión. Combinamos las estimaciones de aves playeras con el modelo de clasificación de la cobertura del suelo y usamos el método de control sintético para crear contrafactuales para las tendencias de las aves playeras en sitios de tratamiento. No encontramos evidencia alguna de un efecto generalizado en las tres variables de los resultados. Los efectos del tratamiento de especies y de sitio fueron heterogéneos, con unos cuantos casos que mostraron evidencia de un efecto positivo, incluido un sitio con un nivel elevado de inversión general. Los resultados sugieren que seis especies de aves playeras declinaron a lo largo de toda la ruta, incluyendo en varios sitios de América Latina. Sin embargo, el porcentaje de poblaciones de la ruta presentes en los sitios permaneció estable y el porcentaje en los sitios de tratamiento (sitios de inversión) fue más elevado que en los sitios control. Muchos mecanismos son posibles detrás de nuestros resultados, incluidas las inversiones que todavía no han mitigado impactos y los impactos negativos en otros sitios que están causando las declinaciones en los sitios de tratamiento. Una limitación en nuestra evaluación es el enfoque único en la abundancia de aves playeras y la falta de datos que impiden la inclusión de otras variables de los resultados. El monitoreo de la infraestructura ahora está en una posición en la que puede diseñar un marco de evaluación más robusto y a priori de las aves playeras a lo largo de toda la ruta. Con este marco, será más fácil priorizar los dólares limitados para que los resultados de conservación sean lo más positivos posible.

Sampling environmental DNA from trees and soil to detect cryptic arboreal mammals.

Environmental DNA (eDNA) approaches to monitoring biodiversity in terrestrial environments have largely focused on sampling water bodies, potentially limiting the geographic and taxonomic scope of eDNA investigations. We assessed the performance of two strictly terrestrial eDNA sampling approaches to detect arboreal mammals, a guild with many threatened and poorly studied taxa worldwide, within two central New Jersey (USA) woodlands. We evaluated species detected with metabarcoding using two eDNA collection methods (tree bark vs. soil sampling), and compared the performance of two detection methods (qPCR vs. metabarcoding) within a single species. Our survey, which included 94 sampling events at 21 trees, detected 16 species of mammals, representing over 60% of the diversity expected in the area. More DNA was found for the 8 arboreal versus 8 non-arboreal species detected (mean: 2466 vs. 289 reads/sample). Soil samples revealed a generally similar composition, but a lower diversity, of mammal species. Detection rates for big brown bat were 3.4 × higher for qPCR over metabarcoding, illustrating the enhanced sensitivity of single-species approaches. Our results suggest that sampling eDNA from on and around trees could serve as a useful new monitoring tool for cryptic arboreal mammal communities globally.

Combining surface and soil environmental DNA with artificial cover objects to improve terrestrial reptile survey detection.

Reptiles are increasingly of conservation concern due to their susceptibility to habitat loss, emerging disease, and harvest in the wildlife trade. However, reptile populations are often difficult to monitor given the frequency of crypsis in their life history. This difficulty has left uncertain the conservation status of many species and the efficacy of conservation actions unknown. Environmental DNA (eDNA) surveys consistently elevate the detection rate of species they are designed to monitor, and while their use is promising for terrestrial reptile conservation, successes in developing such surveys have been sparse. We tested the degree to which inclusion of surface and soil eDNA sampling into conventional artificial-cover methods elevates the detection probability of a small, cryptic terrestrial lizard, Scincella lateralis. The eDNA sampling of cover object surfaces with paint rollers elevated per sample detection probabilities for this species 4-16 times compared with visual surveys alone. We readily detected S. lateralis eDNA under cover objects up to 2 weeks after the last visual detection, and at some cover objects where no S. lateralis were visually observed in prior months. With sufficient sampling intensity, eDNA testing of soil under cover objects produced comparable per sample detection probabilities as roller surface methods. Our results suggest that combining eDNA and cover object methods can considerably increase the detection power of reptile monitoring programs, allowing more accurate estimates of population size, detection of temporal and spatial changes in habitat use, and tracking success of restoration efforts. Further research into the deposition and decay rates of reptile eDNA under cover objects, as well as tailored protocols for different species and habitats, is needed to bring the technique into widespread use.

El interés por la conservación de los reptiles es cada vez mayor debido a su susceptibilidad ante la pérdida del hábitat, enfermedades emergentes y la captura para el mercado de fauna. Sin embargo, las poblaciones de reptiles son difíciles de monitorear por lo frecuente que es la cripsis en sus historias de vida. Esta dificultad deja incierto el estado de conservación de muchas especies y desconocida la eficacia de las acciones de conservación. Los censos de ADN ambiental (DNAa) elevan sistemáticamente la tasa de detección de las especies que monitorean, y aunque su uso es prometedor para la conservación de los reptiles terrestres, han sido escasos los éxitos en el desarrollo de dichos censos. Analizamos el grado al que la inclusión del muestreo de DNAa superficial y del suelo a los métodos convencionales de cobertura artificial eleva la probabilidad de detección de una pequeña lagartija terrestre críptica: Scincella lateralis. El muestreo de DNAa de las superficies con cobertura de objetos con rodillos de pintura elevó las probabilidades de detección por muestra para esta especie 4-16 veces más que los censos visuales. Detectamos fácilmente el DNAa de S. lateralis bajo los objetos de cubierta hasta dos semanas después de la última detección visual y en algunos objetos de cubierta en donde no se había observado en los meses previos a S. lateralis. Con suficiente intensidad de muestreo, el análisis de DNAa del suelo bajo objetos de cubierta produjo probabilidades de detección por muestra comparables como métodos de rodillo superficial. Nuestros resultados sugieren que la combinación del DNAa y los métodos de objetos de cobertura puede incrementar considerablemente el poder de detección de los programas de monitoreo de reptiles, lo que permite estimaciones más precisas del tamaño poblacional, detección de los cambios espaciales y temporales en el uso de hábitat y el éxito de rastreo de los esfuerzos de restauración. Además, se necesita la investigación sobre las tasas de depósito y descomposición del DNAa de reptiles bajo objetos de cubierta, así como los protocolos hechos para diferentes especies y hábitats, para que la técnica entre al uso difundido.

Mapping shifts in spatial synchrony in grassland birds to inform conservation planning.

Spatial synchrony, defined as the correlated fluctuations in abundance of spatially separated populations, can be caused by regional fluctuations in natural and anthropogenic environmental population drivers. Investigations into the geography of synchrony can provide useful insight to inform conservation planning efforts by revealing regions of common population drivers and metapopulation extinction vulnerability. We examined the geography of spatial synchrony and decadal changes in these patterns for grassland birds in the United States and Canada, which are experiencing widespread and persistent population declines. We used Bayesian hierarchical models and over 50 years of abundance data from the North American Breeding Bird Survey to generate population indices within a 2° latitude by 2° longitude grid. We computed and mapped mean local spatial synchrony for each cell (mean detrended correlation of the index among neighboring cells), along with associated uncertainty, for 19 species in 2, 26-year periods, 1968-1993 and 1994-2019. Grassland birds were predicted to increase in spatial synchrony where agricultural intensification, climate change, or interactions between the 2 increased. We found no evidence of an overall increase in synchrony among grassland bird species. However, based on the geography of these changes, there was considerable spatial heterogeneity within species. Averaging across species, we identified clusters of increasing spatial synchrony in the Prairie Pothole and Shortgrass Prairie regions and a region of decreasing spatial synchrony in the eastern United States. Our approach has the potential to inform continental-scale conservation planning by adding an additional layer of relevant information to species status assessments and spatial prioritization of policy and management actions. Our work adds to a growing literature suggesting that global change may result in shifting patterns of spatial synchrony in population dynamics across taxa with broad implications for biodiversity conservation.

La sincronía espacial, definida como las fluctuaciones correlacionadas de la abundancia de poblaciones separadas espacialmente, puede deberse a las fluctuaciones regionales de los conductores ambientales naturales y antropogénicos de las poblaciones. Los análisis de la geografía de las sincronías pueden proporcionar conocimiento útil para guiar los esfuerzos de planeación de la conservación al revelar regiones de conductores comunes de población y la vulnerabilidad de extinción de las metapoblaciones. Examinamos la geografía de la sincronía espacial y los cambios por década en estos patrones para las aves de pastizal en los Estados Unidos y Canadá, las cuales están experimentando declinaciones poblacionales extensas y persistentes. Usamos modelos bayesianos jerárquicos y más de 50 años de datos de abundancia tomados del Censo Norteamericano de Aves Reproductoras para generar índices poblacionales dentro una cuadrícula de 2° de latitud por 2° de longitud. Calculamos y mapeamos la sincronía espacial local media para cada celda (la correlación con deducción de tendencia de la media del índice entre las celdas vecinas), además de la incertidumbre asociada, para 19 especies en dos períodos de 26 años: 1968 - 1993 y 1994 - 2019. Se predijo que las aves de pastizal aumentarían su sincronía espacial en donde aumentarán la intensificación agrícola, el cambio climático o las interacciones entre ambos. No encontramos evidencia de un aumento generalizado de la sincronía entre las especies de aves de pastizal. Sin embargo, con base en la geografía de estos cambios, hubo una heterogeneidad espacial considerable entre las especies. Como promedio entre las especies, identificamos módulos de sincronía espacial en aumento en las regiones de Prairie Pothole y Shortgrass Prairie así como una región de sincronía espacial en declive al este de los Estados Unidos. Nuestra estrategia tiene el potencial para guiar a la planeación de la conservación a escala continental al añadir una capa adicional de información relevante a los análisis del estado de las especies y a la priorización espacial de las acciones de manejo y de políticas. Nuestro trabajo contribuye a una creciente literatura que sugiere que el cambio mundial podría derivar en patrones cambiantes de sincronía espacial en las dinámicas poblacionales de muchos taxones con implicaciones amplias para la conservación de la biodiversidad. Mapeo de los Cambios en la Sincronía Espacial de las Aves de Pastizal para Guiar la Planeación de la Conservación.

In situ differentiation of iridophore crystallotypes underlies zebrafish stripe patterning.

Skin color patterns are ubiquitous in nature, impact social behavior, predator avoidance, and protection from ultraviolet irradiation. A leading model system for vertebrate skin patterning is the zebrafish; its alternating blue stripes and yellow interstripes depend on light-reflecting cells called iridophores. It was suggested that the zebrafish's color pattern arises from a single type of iridophore migrating differentially to stripes and interstripes. However, here we find that iridophores do not migrate between stripes and interstripes but instead differentiate and proliferate in-place, based on their micro-environment. RNA-sequencing analysis further reveals that stripe and interstripe iridophores have different transcriptomic states, while cryogenic-scanning-electron-microscopy and micro-X-ray diffraction identify different crystal-arrays architectures, indicating that stripe and interstripe iridophores are different cell types. Based on these results, we present an alternative model of skin patterning in zebrafish in which distinct iridophore crystallotypes containing specialized, physiologically responsive, organelles arise in stripe and interstripe by in-situ differentiation.

Fate plasticity and reprogramming in genetically distinct populations of Danio leucophores.

Understanding genetic and cellular bases of adult form remains a fundamental goal at the intersection of developmental and evolutionary biology. The skin pigment cells of vertebrates, derived from embryonic neural crest, are a useful system for elucidating mechanisms of fate specification, pattern formation, and how particular phenotypes impact organismal behavior and ecology. In a survey of Danio fishes, including the zebrafish Danio rerio, we identified two populations of white pigment cells-leucophores-one of which arises by transdifferentiation of adult melanophores and another of which develops from a yellow-orange xanthophore or xanthophore-like progenitor. Single-cell transcriptomic, mutational, chemical, and ultrastructural analyses of zebrafish leucophores revealed cell-type-specific chemical compositions, organelle configurations, and genetic requirements. At the organismal level, we identified distinct physiological responses of leucophores during environmental background matching, and we showed that leucophore complement influences behavior. Together, our studies reveal independently arisen pigment cell types and mechanisms of fate acquisition in zebrafish and illustrate how concerted analyses across hierarchical levels can provide insights into phenotypes and their evolution.

Rainbow peacock spiders inspire miniature super-iridescent optics.

Colour produced by wavelength-dependent light scattering is a key component of visual communication in nature and acts particularly strongly in visual signalling by structurally-coloured animals during courtship. Two miniature peacock spiders (Maratus robinsoni and M. chrysomelas) court females using tiny structured scales (~ 40 × 10 µm2) that reflect the full visual spectrum. Using TEM and optical modelling, we show that the spiders' scales have 2D nanogratings on microscale 3D convex surfaces with at least twice the resolving power of a conventional 2D diffraction grating of the same period. Whereas the long optical path lengths required for light-dispersive components to resolve individual wavelengths constrain current spectrometers to bulky sizes, our nano-3D printed prototypes demonstrate that the design principle of the peacock spiders' scales could inspire novel, miniature light-dispersive components.

Hyperspectral imaging of snow algae and green algae from aeroterrestrial habitats.

Snow algae and green algae living in aeroterrestrial habitats are ideal objects to study adaptation to high light irradiation. Here, we used a detailed description of the spectral properties as a proxy for photo-acclimation/protection in snow algae (Chlamydomonas nivalis, Chlainomonas sp. and Chloromonas sp.) and charophyte green algae (Zygnema sp., Zygogonium ericetorum and Klebsormidium crenulatum). The hyperspectral microscopic mapping and imaging technique allowed us to acquire total absorption spectra of these microalgae in the waveband of 400-900nm. Particularly in Chlamydomonas nivalis and Chlainomonas sp., a high absorbance between 400-550nm was observed, due to naturally occurring secondary carotenoids; in Chloromonas sp. and in the charopyhte algae this high absorbance was missing, the latter being close relatives to land plants. To investigate if cellular water loss has an influence on the spectral properties, the cells were plasmolysed in sorbitol or desiccated at ambient air. While in snow algae, these treatments did hardly change the spectral properties, in the charopyhte algae the condensation of the cytoplasm and plastids increased the absorbance in the lower waveband of 400-500nm. These changes might be ecologically relevant and photoprotective, as aeroterrestrial algae are naturally exposed to occasional water limitation, leading to desiccation, which are conditions usually occurring together with higher irradiation.

Effects of CO2-induced pH reduction on the exoskeleton structure and biophotonic properties of the shrimp Lysmata californica.

The anticipated effects of CO2-induced ocean acidification on marine calcifiers are generally negative, and include dissolution of calcified elements and reduced calcification rates. Such negative effects are not typical of crustaceans for which comparatively little ocean acidification research has been conducted. Crustaceans, however, depend on their calcified exoskeleton for many critical functions. Here, we conducted a short-term study on a common caridean shrimp, Lysmata californica, to determine the effect of CO2-driven reduction in seawater pH on exoskeleton growth, structure, and mineralization and animal cryptic coloration. Shrimp exposed to ambient (7.99 ± 0.04) and reduced pH (7.53 ± 0.06) for 21 days showed no differences in exoskeleton growth (percent increase in carapace length), but the calcium weight percent of their cuticle increased significantly in reduced pH conditions, resulting in a greater Ca:Mg ratio. Cuticle thickness did not change, indicating an increase in the mineral to matrix ratio, which may have mechanical consequences for exoskeleton function. Furthermore, there was a 5-fold decrease in animal transparency, but no change in overall shrimp coloration (red). These results suggest that even short-term exposure to CO2-induced pH reduction can significantly affect exoskeleton mineralization and shrimp biophotonics, with potential impacts on crypsis, physical defense, and predator avoidance.

Bio-Inspired Structural Colors Produced via Self-Assembly of Synthetic Melanin Nanoparticles.

Structural colors arising from interactions of light with submicron scale periodic structures have been found in many species across all taxa, serving multiple biological functions including sexual signaling, camouflage, and aposematism. Directly inspired by the extensive use of self-assembled melanosomes to produce colors in avian feathers, we set out to synthesize and assemble polydopamine-based synthetic melanin nanoparticles in an effort to fabricate colored films. We have quantitatively demonstrated that synthetic melanin nanoparticles have a high refractive index and broad absorption spanning across the UV-visible range, similar to natural melanins. Utilizing a thin-film interference model, we demonstrated the coloration mechanism of deposited films and showed that the unique optical properties of synthetic melanin nanoparticles provide advantages for structural colors over other polymeric nanoparticles (i.e., polystyrene colloidal particles).

Capillary foams: stabilization and functionalization of porous liquids and solids.

Liquid foams are two-phase systems in which a large volume of gas is dispersed as bubbles in a continuous liquid phase. These foams are ubiquitous in nature. In addition, they are found in industrial applications, such as pharmaceutical formulation, food processing, wastewater treatment, construction, and cosmetics. Recently, we reported a new type of foam material, capillary foam, which is stabilized by the synergistic action of particles and a small amount of an immiscible secondary liquid. In this study, we explore in more detail the foam preparation routes. To illustrate some of the potential applications, we create vividly colored wet and dried foams, which are difficult to prepare using traditional methods, and load-bearing porous solids. The combined action of particles and immiscible secondary fluid confers exceptional stability to capillary foams and many options for functionalization, suggesting a wide range of possible applications.

High-density lipoprotein apolipoprotein A-I kinetics in obesity.

OBJECTIVE: Low plasma concentrations of high-density lipoprotein (HDL)-cholesterol and apolipoprotein A-I (apoA-I) are independent predictors of coronary artery disease and are often associated with obesity and the metabolic syndrome. However, the underlying kinetic determinants of HDL metabolism are not well understood. RESEARCH METHODS AND PROCEDURES: We pooled data from 13 stable isotope studies to investigate the kinetic determinants of apoA-I concentrations in lean and overweight-obese individuals. We also examined the associations of HDL kinetics with age, sex, BMI, fasting plasma glucose, fasting insulin, Homeostasis Model Assessment score, and concentrations of apoA-I, triglycerides, HDL-cholesterol and low-density lipoprotein-cholesterol. RESULTS: Compared with lean individuals, overweight-obese individuals had significantly higher HDL apoA-I fractional catabolic rate (0.21+/-0.01 vs. 0.33+/-0.01 pools/d; p<0.001) and production rate (PR; 11.3+/-4.4 vs. 15.8+/-2.77 mg/kg per day; p=0.001). In the lean group, HDL apoA-I PR was significantly associated with apoA-I concentration (r=0.455, p=0.004), whereas in the overweight-obese group, both HDL apoA-I fractional catabolic rate (r=-0.396, p=0.050) and HDL apoA-I PR (r=0.399, p=0.048) were significantly associated with apoA-I concentration. After adjustment for fasting insulin or Homeostasis Model Assessment score, HDL apoA-I PR was an independent predictor of apoA-I concentration. DISCUSSION: In overweight-obese subjects, hypercatabolism of apoA-I is paralleled by an increased production of apoA-I, with HDL apoA-I PR being the stronger determinant of apoA-I concentration. This could have therapeutic implications for the management of dyslipidemia in individuals with low plasma HDL-cholesterol.