Revealing Assembly of a Pore-Forming Complex Using Single-Cell Kinetic Analysis and Modeling

Many biological processes depend on the sequential assembly of protein complexes. However, studying the kinetics of such processes by direct methods is often not feasible. As an important class of such protein complexes, pore-forming toxins start their journey as soluble monomeric proteins, and oligomerize into transmembrane complexes to eventually form pores in the target cell membrane. Here, we monitored pore formation kinetics for the well-characterized bacterial pore-forming toxin aerolysin in single cells in real time to determine the lag times leading to the formation of the first functional pores per cell. Probabilistic modeling of these lag times revealed that one slow and seven equally fast rate-limiting reactions best explain the overall pore formation kinetics. The model predicted that monomer activation is the rate-limiting step for the entire pore formation process. We hypothesized that this could be through release of a propeptide and indeed found that peptide removal abolished these steps. This study illustrates how stochasticity in the kinetics of a complex process can be exploited to identify rate-limiting mechanisms underlying multistep biomolecular assembly pathways.

Carbon-Based Ocean Productivity and Phytoplankton Physiology from Space

Ocean biogeochemical and ecosystem processes are linked by net primary production (NPP) in the ocean's surface layer, where inorganic carbon is fixed by photosynthetic processes. Determinations of NPP are necessarily a function of phytoplankton biomass and its physiological status, but the estimation of these two terms from space has remained an elusive target. Here we present new satellite ocean color observations of phytoplankton carbon (C) and chlorophyll (Chl) biomass and show that derived Chl:C ratios closely follow anticipated physiological dependencies on light, nutrients, and temperature. With this new information, global estimates of phytoplankton growth rates (mu) and carbon-based NPP are made for the first time. Compared to an earlier chlorophyll-based approach, our carbon-based values are considerably higher in tropical oceans, show greater seasonality at middle and high latitudes, and illustrate important differences in the formation and demise of regional algal blooms. This fusion of emerging concepts from the phycological and remote sensing disciplines has the potential to fundamentally change how we model and observe carbon cycling in the global oceans.

Oxygen Production and Carbon Sequestration in an Upwelling Coastal Margin

We examined high-resolution cross-shelf distributions of particulate organic carbon (POC) and dissolved O(2) during the upwelling season off the Oregon coast. Oxygen concentrations were supersaturated in surface waters, and hypoxic in near-bottom waters, with greatly expanded hypoxic conditions late in the season. Simplified time-dependent mass balances on cross-shelf integrated concentrations of these two parameters, found the following: ( 1) The average net rate of photosynthesis generated 2.1 mmol O(2) m(-3) d(-1) and ( 2) essentially none of the corresponding net carbon fixation of 1.4 mmol m(-3) d(-1) could be accounted for in the observed standing stocks of POC. After examining other possible sinks for carbon, we conclude that most of the net production is being exported to the adjacent deep ocean. A simplified POC budget suggests that about a quarter of the export is via alongshore advection, and the remainder is due to some other process. We propose a simplistic conceptual model of across-shelf transport in which POC sinks to the bottom boundary layer where it comes into contact with mineral ballast material but is kept in suspension by high turbulence. When upwelling conditions ease, the BBL waters move seaward, carrying the suspended, ballasted POC with it where it sinks rapidly into the deep ocean at the shelf break. This suggests a mechanism whereby the duration and frequency of upwelling events and relaxations can determine the extent to which new carbon produced by photosynthesis in the coastal ocean is exported to depth rather than being respired on the shelf.

High-Frequency in Situ Optical Measurements During a Storm Event: Assessing Relationships Between Dissolved Organic Matter, Sediment Concentrations, and Hydrologic Processes

Dissolved organic matter (DOM) dynamics during storm events has received considerable attention in forested watersheds, but the extent to which storms impart rapid changes in DOM concentration and composition in highly disturbed agricultural watersheds remains poorly understood. In this study, we used identical in situ optical sensors for DOM fluorescence (FDOM) with and without filtration to continuously evaluate surface water DOM dynamics in a 415 km(2) agricultural watershed over a 4 week period containing a short-duration rainfall event. Peak turbidity preceded peak discharge by 4 h and increased by over 2 orders of magnitude, while the peak filtered FDOM lagged behind peak turbidity by 15 h. FDOM values reported using the filtered in situ fluorometer increased nearly fourfold and were highly correlated with dissolved organic carbon (DOC) concentrations (r(2) = 0.97), providing a highly resolved proxy for DOC throughout the study period. Discrete optical properties including specific UV absorbance (SUVA(254)), spectral slope (S(290-350)), and fluorescence index (FI) were also strongly correlated with in situ FDOM and indicate a shift toward aromatic, high molecular weight DOM from terrestrially derived sources during the storm. The lag of the peak in FDOM behind peak discharge presumably reflects the draining of watershed soils from natural and agricultural landscapes. Field and experimental evidence showed that unfiltered FDOM measurements underestimated filtered FDOM concentrations by up to similar to 60% at particle concentrations typical of many riverine systems during hydrologic events. Together, laboratory and in situ data provide insights into the timing and magnitude of changes in DOM quantity and quality during storm events in an agricultural watershed, and indicate the need for sample filtration in systems with moderate to high suspended sediment loads.

A Comparison of Hydrographically and Optically Derived Mixed Layer Depths

Efforts to understand and model the dynamics of the upper ocean would be significantly advanced given the ability to rapidly determine mixed layer depths (MLDs) over large regions. Remote sensing technologies are an ideal choice for achieving this goal. This study addresses the feasibility of estimating MLDs from optical properties. These properties are strongly influenced by suspended particle concentrations, which generally reach a maximum at pycnoclines. The premise therefore is to use a gradient in beam attenuation at 660 nm (c660) as a proxy for the depth of a particle-scattering layer. Using a global data set collected during World Ocean Circulation Experiment cruises from 1988-1997, six algorithms were employed to compute MLDs from either density or temperature profiles. Given the absence of published optically based MLD algorithms, two new methods were developed that use c660 profiles to estimate the MLD. Intercomparison of the six hydrographically based algorithms revealed some significant disparities among the resulting MLD values. Comparisons between the hydrographical and optical approaches indicated a first-order agreement between the MLDs based on the depths of gradient maxima for density and c660. When comparing various hydrographically based algorithms, other investigators reported that inherent fluctuations of the mixed layer depth limit the accuracy of its determination to 20 m. Using this benchmark, we found a similar to 70% agreement between the best hydrographical-optical algorithm pairings.

Observations of the Sensitivity of Beam Attenuation to Particle Size in a Coastal Bottom Boundary Layer

The goal of this study was to test the hypothesis that the aggregated state of natural marine particles constrains the sensitivity of optical beam attenuation to particle size. An instrumented bottom tripod was deployed at the 12-m node of the Martha's Vineyard Coastal Observatory to monitor particle size distributions, particle size-versus-settling-velocity relationships, and the beam attenuation coefficient (c(p)) in the bottom boundary layer in September 2007. An automated in situ filtration system on the tripod collected 24 direct estimates of suspended particulate mass (SPM) during each of five deployments. On a sampling interval of 5 min, data from a Sequoia Scientific LISST 100x Type B were merged with data from a digital floc camera to generate suspended particle volume size distributions spanning diameters from approximately 2 mu m to 4 cm. Diameter-dependent densities were calculated from size-versus-settling-velocity data, allowing conversion of the volume size distributions to mass distributions, which were used to estimate SPM every 5 min. Estimated SPM and measured c(p) from the LISST 100x were linearly correlated throughout the experiment, despite wide variations in particle size. The slope of the line, which is the ratio of c(p) to SPM, was 0.22 g m(-2). Individual estimates of c(p):SPM were between 0.2 and 0.4 g m(-2) for volumetric median particle diameters ranging from 10 to 150 mu m. The wide range of values in c(p):SPM in the literature likely results from three factors capable of producing factor-of-two variability in the ratio: particle size, particle composition, and the finite acceptance angle of commercial beam-transmissometers.

Carbon-Based Primary Productivity Modeling With Vertically Resolved Photoacclimation

Net primary production (NPP) is commonly modeled as a function of chlorophyll concentration (Chl), even though it has been long recognized that variability in intracellular chlorophyll content from light acclimation and nutrient stress confounds the relationship between Chl and phytoplankton biomass. It was suggested previously that satellite estimates of backscattering can be related to phytoplankton carbon biomass (C) under conditions of a conserved particle size distribution or a relatively stable relationship between C and total particulate organic carbon. Together, C and Chl can be used to describe physiological state (through variations in Chl:C ratios) and NPP. Here, we fully develop the carbon-based productivity model (CbPM) to include information on the subsurface light field and nitracline depths to parameterize photoacclimation and nutrient stress throughout the water column. This depth-resolved approach produces profiles of biological properties (Chl, C, NPP) that are broadly consistent with observations. The CbPM is validated using regional in situ data sets of irradiance-derived products, phytoplankton chlorophyll: carbon ratios, and measured NPP rates. CbPM-based distributions of global NPP are significantly different in both space and time from previous Chl-based estimates because of the distinction between biomass and physiological influences on global Chl fields. The new model yields annual, areally integrated water column production of similar to 52 Pg C a(-1) for the global oceans.

Distributions and Variability of Particulate Organic Matter in a Coastal Upwelling System

In this study we examined the spatial and temporal variability of particulate organic material (POM) off Oregon during the upwelling season. High-resolution vertical profiling of beam attenuation was conducted along two cross-shelf transects. One transect was located in a region where the shelf is relatively uniform and narrow (off Cascade Head (CH)); the second transect was located in a region where the shelf is shallow and wide (off Cape Perpetua (CP)). In addition, water samples were collected for direct analysis of chlorophyll, particulate organic carbon (POC), and particulate organic nitrogen (PON). Beam attenuation was highly correlated with POC and PON. Striking differences in distribution patterns and characteristics of POM were observed between CH and CP. Off CH, elevated concentrations of chlorophyll and POC were restricted to the inner shelf and were highly variable in time. The magnitude of the observed short-term temporal variability was of the same order as that of the seasonal variability reported in previous studies. Elevated concentrations of nondegraded chlorophyll and POM were observed near the bottom. Downwelling and rapid sinking are two mechanisms by which phytoplankton cells can be delivered to the bottom before being degraded. POM may be then transported across the shelf via the benthic nepheloid layer. Along the CP transect, concentrations of POM were generally higher than they were along the CH transect and extended farther across the shelf. Characteristics of surface POM, namely, C: N ratios and carbon: chlorophyll ratios, differed between the two sites. These differences can be attributed to differences in shelf circulation.

Destellos de un nuevo sujeto : Los conflictos obreros en los saladeros y el nacimiento de la clase obrera entrerriana (1854-1868)

El objetivo del trabajo es demostrar la existencia de conflictos obreros en Entre Ríos, a partir de las luchas salariales en los saladeros entre 1854 y 1868. Colateralmente, cuestionamos las fechas de génesis del movimiento obrero argentino. La reconstrucción de las luchas se realizó con una gran variedad de fuentes: crónicas de viajeros, documentación oficial y, principalmente, el análisis epistolar de los establecimientos saladeriles de Justo José de Urquiza. A partir de la evidencia empírica, que muestra los enfrentamientos entre los trabajadores y patrones, sostenemos la existencia de un proletariado en formación. Se analizarán las estrategias de luchas y los mecanismos patronales para superar las situaciones planteadas

Las formas del salario en la agricultura pampeana : Su rol en el disciplinamiento, el aumento de la productividad y el abaratamiento de la fuerza de trabajo

Las transformaciones socio-económicas, políticas y gremiales acaecidas a fines de los '70 inauguraron un período desfavorable para las condiciones de trabajo de los obreros agrícolas. Se desarrolló una tendencia hacia la disminución de los tiempos requeridos por cada tarea, la discontinuación del proceso productivo, la estacionalización de la demanda de empleo y una mayor especialización de la fuerza de trabajo. La fragmentación extrema del proceso productivo derivó así en el astillamiento del ciclo laboral de los trabajadores en términos de ingresos y de empleadores. En este contexto, fueron logrando imponerse y naturalizarse distintas formas y niveles de remuneración para cada una de las tareas agrícolas, aun en los casos en que un trabajador las realizara todas para un mismo patrón como peón permanente, y singularmente en los casos de los obreros temporarios que se enfrentan a distintos empleadores para desarrollar cada una de las labores con las que construyen su ciclo laboral. Nos proponemos analizar el rol que la combinación de estas formas y niveles salariales ha venido cumpliendo para el abaratamiento de la fuerza de trabajo agrícola en su conjunto, así como en su estímulo a una mayor productividad y en la frustración de conflictos abiertos o a gran escala entre esta fracción de la clase obrera rural y sus patrones, a pesar de las condiciones de trabajo desfavorables que verificamos en este estudio

Destellos de un nuevo sujeto : Los conflictos obreros en los saladeros y el nacimiento de la clase obrera entrerriana (1854-1868)

El objetivo del trabajo es demostrar la existencia de conflictos obreros en Entre Ríos, a partir de las luchas salariales en los saladeros entre 1854 y 1868. Colateralmente, cuestionamos las fechas de génesis del movimiento obrero argentino. La reconstrucción de las luchas se realizó con una gran variedad de fuentes: crónicas de viajeros, documentación oficial y, principalmente, el análisis epistolar de los establecimientos saladeriles de Justo José de Urquiza. A partir de la evidencia empírica, que muestra los enfrentamientos entre los trabajadores y patrones, sostenemos la existencia de un proletariado en formación. Se analizarán las estrategias de luchas y los mecanismos patronales para superar las situaciones planteadas

Destellos de un nuevo sujeto : Los conflictos obreros en los saladeros y el nacimiento de la clase obrera entrerriana (1854-1868)

El objetivo del trabajo es demostrar la existencia de conflictos obreros en Entre Ríos, a partir de las luchas salariales en los saladeros entre 1854 y 1868. Colateralmente, cuestionamos las fechas de génesis del movimiento obrero argentino. La reconstrucción de las luchas se realizó con una gran variedad de fuentes: crónicas de viajeros, documentación oficial y, principalmente, el análisis epistolar de los establecimientos saladeriles de Justo José de Urquiza. A partir de la evidencia empírica, que muestra los enfrentamientos entre los trabajadores y patrones, sostenemos la existencia de un proletariado en formación. Se analizarán las estrategias de luchas y los mecanismos patronales para superar las situaciones planteadas

Las formas del salario en la agricultura pampeana : Su rol en el disciplinamiento, el aumento de la productividad y el abaratamiento de la fuerza de trabajo

Las transformaciones socio-económicas, políticas y gremiales acaecidas a fines de los '70 inauguraron un período desfavorable para las condiciones de trabajo de los obreros agrícolas. Se desarrolló una tendencia hacia la disminución de los tiempos requeridos por cada tarea, la discontinuación del proceso productivo, la estacionalización de la demanda de empleo y una mayor especialización de la fuerza de trabajo. La fragmentación extrema del proceso productivo derivó así en el astillamiento del ciclo laboral de los trabajadores en términos de ingresos y de empleadores. En este contexto, fueron logrando imponerse y naturalizarse distintas formas y niveles de remuneración para cada una de las tareas agrícolas, aun en los casos en que un trabajador las realizara todas para un mismo patrón como peón permanente, y singularmente en los casos de los obreros temporarios que se enfrentan a distintos empleadores para desarrollar cada una de las labores con las que construyen su ciclo laboral. Nos proponemos analizar el rol que la combinación de estas formas y niveles salariales ha venido cumpliendo para el abaratamiento de la fuerza de trabajo agrícola en su conjunto, así como en su estímulo a una mayor productividad y en la frustración de conflictos abiertos o a gran escala entre esta fracción de la clase obrera rural y sus patrones, a pesar de las condiciones de trabajo desfavorables que verificamos en este estudio

Destellos de un nuevo sujeto : Los conflictos obreros en los saladeros y el nacimiento de la clase obrera entrerriana (1854-1868)

El objetivo del trabajo es demostrar la existencia de conflictos obreros en Entre Ríos, a partir de las luchas salariales en los saladeros entre 1854 y 1868. Colateralmente, cuestionamos las fechas de génesis del movimiento obrero argentino. La reconstrucción de las luchas se realizó con una gran variedad de fuentes: crónicas de viajeros, documentación oficial y, principalmente, el análisis epistolar de los establecimientos saladeriles de Justo José de Urquiza. A partir de la evidencia empírica, que muestra los enfrentamientos entre los trabajadores y patrones, sostenemos la existencia de un proletariado en formación. Se analizarán las estrategias de luchas y los mecanismos patronales para superar las situaciones planteadas

Las formas del salario en la agricultura pampeana : Su rol en el disciplinamiento, el aumento de la productividad y el abaratamiento de la fuerza de trabajo

Las transformaciones socio-económicas, políticas y gremiales acaecidas a fines de los '70 inauguraron un período desfavorable para las condiciones de trabajo de los obreros agrícolas. Se desarrolló una tendencia hacia la disminución de los tiempos requeridos por cada tarea, la discontinuación del proceso productivo, la estacionalización de la demanda de empleo y una mayor especialización de la fuerza de trabajo. La fragmentación extrema del proceso productivo derivó así en el astillamiento del ciclo laboral de los trabajadores en términos de ingresos y de empleadores. En este contexto, fueron logrando imponerse y naturalizarse distintas formas y niveles de remuneración para cada una de las tareas agrícolas, aun en los casos en que un trabajador las realizara todas para un mismo patrón como peón permanente, y singularmente en los casos de los obreros temporarios que se enfrentan a distintos empleadores para desarrollar cada una de las labores con las que construyen su ciclo laboral. Nos proponemos analizar el rol que la combinación de estas formas y niveles salariales ha venido cumpliendo para el abaratamiento de la fuerza de trabajo agrícola en su conjunto, así como en su estímulo a una mayor productividad y en la frustración de conflictos abiertos o a gran escala entre esta fracción de la clase obrera rural y sus patrones, a pesar de las condiciones de trabajo desfavorables que verificamos en este estudio