Water Regulation Research Articles

Majority of potable water microplastics are smaller than the 20 μm EU methodology limit for consumable water quality

Microplastic (MP) content in nutrition including potable water is unregulated, although MP concentrations in individual brands of bottled water diverge by several orders of magnitude. The EU Directive 2020/2184 on consumable water quality recently proposed methodological approaches to the detection of MPs in potable water in the size range of 20–5000 μm. However, fine MPs in the 1–20 μm range are far more likely to pass the human intestine into blood and organs. To evaluate the impact of this omission on the total number of detected MPs, we determined MP concentrations down to 1 μm in ten different brands of polyethylene terephthalate (PET) bottled water and one tap water sample using automated Raman microspectroscopy. We found that MP concentrations ranged from 19 to 1,154 (n/L) [0.001 to 0.250 μg/L], and although all the investigated samples of bottled water were stored in PET containers, PET accounted only for a small percentage of identified MPs in most samples. Importantly, 98 and 94% of MPs measured less than 20 and 10 μm in diameter, respectively, demonstrating the importance of small MP inclusion in potable water analyses and regulation. The current study presents a protocol to identify MPs down to 1 μm in any type of potable water regardless of hardness, and demonstrates the importance of implementing both negative and positive procedural, quality control measures.

Thinning Improves Large Diameter Timber Cultivation but Undermines Ecosystem Multifunctionality in the Short Term

Implementing thinning practices can enhance the growth of plantation forests and improve soil health. Nevertheless, the impacts of thinning applications on soil quality, large-diameter timber production of Castanopsis hystrix, and ecosystem multifunctionality are poorly understood. Therefore, we chose two sample plots, unthinned (control) and thinned, to investigate productivity and ecosystem multifunctionality after thinning for six years. Results revealed that thinning significantly reduced the soil’s bulk density, enhanced large-diameter timber growth, and undermined ecosystem multifunctionality in the short term compared to control (unthinning) treatment. Compared to the control, the thinning treatment considerably enhanced the soil organic carbon (0–30 cm soil layer) and tree diameter at breast height (20–30 cm), and enhanced shrub leaf nitrogen (N), shrub root N, herb aboveground N, Gram-positive bacteria (0–10 cm soil layer), and Gram-positive bacteria (20–30 cm soil layer) contents by 29.61%, 65.29%, 44.61%, 274.35%, and 323.44%, respectively. Furthermore, the thinning application could improve the N and P resorption efficiency more than control. Furthermore, compared with control, thinning treatment maximized decomposition and nutrient cycling function by 11.81% and 143.40%, respectively. Moreover, total PLFA content significantly impacts carbon stocks, wood production, and water regulation functions. In conclusion, this study underscores the considerable potential of thinning in augmenting large-diameter timber production by stimulating the positive effects of forest stands. These findings provide valuable insights for ecosystem multifunctionality elevation and the judicious application of thinning to improve forestry productivity, facilitating sustainable development in the forestry sector.

Revealing gaps in marine evidence with a natural capital lens.

The natural capital concept positions the natural environment as an asset, crucial for the flow of goods and benefits to humanity. There is a growing trend in applying this concept in marine environmental management in the United Kingdom (UK). This study evaluates six varied marine decisions across England, Scotland and Wales. It focuses on the evidence informing these decisions and the extent to which they represent the complete spectrum of marine natural assets and ecosystem services. We identified a reliance on various evidence types, including consultations, data and statistics, maps and literature reviews. Natural assets such as aquatic resources and energy sources were most frequently evidenced. Fishing was the predominant provisioning ecosystem service benefit. There was a notable gap in evidence on marine habitats' water quality regulation service. Recreation and tourism dominated the cultural ecosystem service evidence, with less focus on indirect uses such as spiritual nature connections. We reveal gaps in the evidence in marine decisions on significant marine ecosystem service benefits. Our study provides additional evidence to an already identified need to fill evidence gaps in marine water regulation and non-use values of the UK's marine environments.This article is part of the discussion meeting issue 'Bending the curve towards nature recovery: building on Georgina Mace's legacy for a biodiverse future'.

Interfacial Water Regulation for Nitrate Electroreduction to Ammonia at Ultralow Overpotentials.

Nitrate electroreduction is promising for achieving effluent waste-water treatment and ammonia production with respect to the global nitrogen balance. However, due to the impeded hydrogenation process, high overpotentials need to be surmounted during nitrate electroreduction, causing intensive energy consumption. Herein, a hydroxide regulation strategy is developed to optimize the interfacial H2O behavior for accelerating the hydrogenation conversion of nitrate to ammonia at ultralow overpotentials. The well-designed Ru─Ni(OH)2 electrocatalyst shows a remarkable energy efficiency of 44.6% at +0.1V versus RHE and a nearly 100% Faradaic efficiency for NH3 synthesis at 0V versus RHE. In situ characterizations and theoretical calculations indicate that Ni(OH)2 can regulate the interfacial H2O structure with a promoted H2O dissociation process and contribute to the spontaneous hydrogen spillover process for boosting NO3 - electroreduction to NH3 at Ru sites. Furthermore, the assembled rechargeable Zn-NO3 -/ethanol battery system exhibits an outstanding long-term cycling stability during the charge-discharge tests with the production of high-value-added ammonium acetate, showing great potential for simultaneously achieving nitrate removal, energy conversion, and chemical synthesis. This work can not only provide a guidance for interfacial H2O regulation in extensive hydrogenation reactions but also inspire the design of a novel hybrid flow battery with multiple functions.

Water regulation weakens the relationship between biodiversity and ecosystem multifunctionality: Insights from a highly managed Chinese lake.

Water diversions can mitigate water scarcities by strategically reallocating water resources. Despite their benefits, these interventions may profoundly affect biodiversity and multiple ecological functions ("multifunctionality") within highly managed lake systems. However, the specific impact of such interventions on the relationship between biodiversity and multifunctionality remains elusive, which limits our grasp of how water regulation shapes the dynamics of managed lake ecosystems. In this study, we investigated the differences in biodiversity and ecological functions between periods with and without water regulation in a highly managed lake, and extended this analysis by calculating diversity and multi-diversity indices and assessing the ecosystem's multifunctionality, with the goal of improving our understanding of how water regulation affects the ecological integrity of such lakes. We found that the diversity and multi-diversity indices both decreased markedly during the regulation period, indicating a negative impact on biodiversity. Conversely, multifunctionality remained stable. To clarify these responses, we developed linear mixed-effects models that incorporated multi-diversity indices and multifunctionality. Our analysis uncovered a robust positive correlation between multi-richness and multifunctionality. However, this relationship weakened during the periods with water regulation. Finally, we established structural equation models based on trophic cascade theory to pinpoint the key biological groups that sustained multifunctionality. We found that plants and omnivorous fish were instrumental in supporting multifunctionality during the non-regulation period, whereas planktivorous fish played a crucial role during the regulation period. These findings significantly enhance our understanding of how water regulation influences both biodiversity and multifunctionality, and provide insights for future management of regulated lakes.

EXPERIMENTAL STUDY OF WATER AND NUTRIENT BALANCE INTERACTIONS IN CLOSED HYDROPONIC SYSTEMS

This article presents a study aimed at modeling the interactions of water and nutrient balances in hydroponic systems, which are increasingly recognized as an environmentally sustainable and highly efficient method for growing plants without soil. Hydroponic systems provide optimal nutrient delivery to plants through a water solution maintained in a closed cycle, reducing water and fertilizer usage compared to traditional agriculture. However, the effectiveness of such systems largely depends on precise regulation of water and nutrient balances to ensure proper plant growth and optimize resource use. The purpose of this study is to develop a comprehensive model that considers the dynamics of water and nutrient environments within the hydroponic system and predicts plants' requirements for essential nutrients. The article examines key parameters that influence nutrient balance in hydroponics, including pH level, concentrations of essential micro- and macronutrients, nutrient uptake rates by plants, evaporation, transpiration, temperature variations, and the nutrient ratio in the solution. Achieving stable water and nutrient balance requires accounting for all these factors and adjusting system parameters according to external conditions and the plants' growth stage. The study’s methodology is based on a combination of mathematical modeling and experimental data, enabling the creation of an adaptive model for the hydroponic system. The article details the model structure, including input variables (temperature, humidity, nutrient concentration), exchange processes (uptake and transpiration), and output variables (optimal nutrient element concentrations). The modeling results facilitate the control and adjustment of the nutrient solution, minimizing water and fertilizer usage while ensuring maximum plant yield. The practical significance of the developed model lies in its applicability for managing hydroponic farms of various scales, from home setups to industrial agricultural enterprises.

Water flow regulation and incentives for conservation: Coca river - ecuadorian Amazon

Conservation and land use policies are key to regulating water in watersheds, influencing water quantity, quality and storage, and mitigating risks such as erosion and flooding. In Ecuador, the Socio Bosque Program (SBP), part of the Good Living Development Plan (2013-2017), focused on conservation without fully considering water benefits. To optimize its implementation, the upper basin of the Coca River, relevant for its hydroelectric potential, was evaluated by analyzing the inclusion of water criteria in the SBP. Four scenarios of land cover and land use change were developed using the Delphi method: Business as Usual (BAU), Strengthening of SPB (SBPS), National Incentive Plan (NIP), and Degradation (DEG). Runoff rates were estimated for each scenario using a land cover and land use model (TerrSet) and a hydrological model (SWAT). The results show that BAU and DEG cause a decrease in lateral and base flow and an increase in surface runoff, while NIP is the only scenario that maintains water regulation. This highlights the need for future incentives to prioritize strategic areas for water regulation in line with watershed management objectives.

Spatio-temporal analysis of scientific research on models to estimate water balance in hydrographic basinsV

Objective: To identify in the world the main models or tools used to evaluate changes in water regulation,through a bibliometric review in the especialized in scientific information, Scopus database, in order to knowthe main variables evaluated that affect the hydrological regime of the basins.Design/methodology/approach: With the help of bibliometric techniques, scientific articles available untilDecember 2022 in the Scopus database were collected using the SUBJAREA (envi) TITLE-ABS-KEY (“waterbalance” AND “land use”) AND (LIMIT-TO (OA, “all”)) AND (LIMIT-TO (DOCTYPE, “ar”)).Results: 407 scientific articles were collected from 1965 to 2022, this scientific production shows an exponentialgrowing trend that was concentrated above all in countries with agricultural traditions such as the UnitedStates (89 publications), China (33), Germany (28), the United Kingdom (25) and Australia (21).Limitations on study/implications: Most of the research was developed on topics associated with land usechange (82 texts), climate change (48), hydrological processes (44), subsoil water (34) and evapotranspirationcalculation (33). The most recurrent models for calculating the water balance were the Direct Water Balance(230 texts) and the Soil and Water Assessment Tool (83).Findings/conclusions: No research development was found in Latin American and African countries, sothese results can help redirect efforts in the research centers of these countries, with the aim of generating newknowledge that helps make decisions about how to improve the efficient use of water in the face of climatechange scenarios and dynamics of land use change

A Case Study on Recycling Industrial Wastewater with Nanofiltration Membrane Separation Technology.

As pressure on water resources intensifies and stringent regulations for groundwater and surface water are enacted, wastewater recycling has emerged as a key research objective for many enterprises. In this study, based on the actual wastewater discharged from Eternal Electronic (Suzhou, China) Co., Ltd., the performance differences in different membrane materials in treating this wastewater were analyzed and compared. The NF90 membrane was ultimately selected as the most suitable choice for treating this wastewater with optimal operating conditions of 150 psi, 25 °C, and 1 L/min, respectively. All the indices of wastewater treatment can satisfy the standard of circulating cooling water. In addition, this work is closely combined with the practical applications by using an HCl solution (pH = 3-4) to clean the fouled membranes, thus effectively solving the membrane fouling in the treatment process. This approach not only satisfies the environmental management requirements of enterprises but also provides valuable insights for similar wastewater treatment projects.

Hydroponic System with Integrator Modified on PI Controller for Nutrient and Water Regulation

— This study developed a hydroponic control system to regulate nutrient concentration (PPM) and water volume using a Proportional-Integral (PI) controller. The PI controller was chosen for its simplicity and effectiveness in systems that do not require fast response times. A trial-and-error method was used to tune the Kp and Ki values, which were implemented in an ATMega328 microcontroller. The system consists of one AC pump and two DC pumps: DC Pump-1 controls the PPM by supplying liquid fertilizer, while DC Pump-2 regulates the water volume. The results showed that modifying the PI controller with an integrator limitation improved performance. For PPM control, a limit of 13000 with Kp = 2 and Ki = 0.01 resulted in a 0.5% error, 56.8-second rise time, and a 4.8% overshot. For water level control, a limit of 12000 with Kp = 100 and Ki = 0.16 yielded a 3.25% error, 2% overshot, and a 65.4-second rise time. The test results demonstrated that the use of AB Mix can be reduced by up to 26% compared to a conventional PID, with an initial PPM value of around 350.

Multiscale Effects of Xylella fastidiosa on Landscape Services

The spread of Xylella fastidiosa since 2013 in the Mediterranean olive groves of the Apulia region has modified the landscape. The aims of this research are focused on the analysis of its effects on the following: (1) Landscape multifunctionality supported by olive groves in terms of landscape service provision; (2) The functional relations among the main Mediterranean land covers in terms of landscape service supply and demand. (3) The landscape fragmentation at different spatial scales. The landscape has completely changed, mainly in those land covers that, in the past, acted as stabilizing factors (croplands and olive groves), which has been replaced by grasslands in 2021. The main effects of Xylella fastidiosa were on the multifunctionality of olive grove landscape in terms of food production, water regulation, carbon sequestration, and pollination, as well as on landscape cultural value. Ecosystem service supply is mainly related to olive groves, tree covers, shrublands, and wetlands. The province of Lecce showed the highest fragmentation, as demonstrated by the number of patches, the mean patch area, and the DIVISION metric, while the province of Brindisi was the least fragmented, with a DIVISION metric similar in 2011 and in 2021. The multiscale assessment of “olive groves” fragmentation has helped in better analyzing the effect of its spatial configuration on the provision of landscape services and in identifying the right spatial scale for each landscape service provision. It is essential to analyze landscape service flow to enlarge the understanding of the ways in which their supply is maintained through a landscape regeneration policy toward the socio-economic–ecological recovery.

Quantification of the Total and Extractable Content of Micro- and Trace Elements in Linden Blossom and Infusions—The Impact of Urban Pollution on Health Risk for Samples from Plovdiv, Bulgaria

Linden (Tilia) is one of the most frequently utilized plants for the preparation of infusions because of its salutary effects, including the reduction in inflammatory processes and pain, alleviation of stress, and lowering of blood pressure. As Linden is a common species in Bulgarian cities, it is frequently used for homemade infusions. The regular consumption of these tea beverages may contribute to the attainment of the recommended daily allowances of certain minerals, but it may also result in the accumulation of toxic elements within the human body. The present study compares the concentrations of essential and toxic elements in linden blossom collected from disparate locations in Plovdiv, Bulgaria, with those of samples gathered in ecologically pristine regions and commercially available products labeled “bio”. Both total element content and extractable forms in infusions were quantified by ICP-MS. The health risk due to consuming infusions was assessed by comparing the water, tea, drinks, and EFSA regulations. The applied cluster analysis divided the samples from the urban area into three groups related to traffic pollution. In spite of the short blooming period, the concentrations of Al, Fe, Pb, V, Cr, Co, Ni, and Cd in the samples from the most polluted areas are increased by a factor of two compared to those from the clean zones.

The Response of Runoff to Land Use Change in the Northeastern Black Soil Region, China

With the intensification of climate change and human activities, the impacts of land use shifts on hydrological processes are becoming more pronounced, especially in regions with complex geographic, geological, and climatic conditions such as the Northeast Black Soil Region, China. This study quantitatively examines the variations in various land use types from 1980 to 2020 by means of a land use transfer matrix, and it incorporates the multi-year average runoff value to mitigate the interference of short-term climate fluctuations on the runoff trend, thereby enhancing the representativeness and stability of the simulation outcomes. The SWAT (Soil and Water Assessment Tool) model is employed to simulate land use alterations in different periods. The findings indicate that the area of farmland increased by 5.34% and the area of grassland decreased by 5.36% over 40 years. The areas of forest land and wetland have fluctuated significantly due to policy interventions and population growth. This study discovers that LUCC has resulted in a marginal increase in annual water yield. For instance, the water yield of paddy fields in 2020 amounts to 92.26 mm/year, which is 0.52–9.42% higher than the historical scenario and exhibits a notable upward trend in summer. Spatial analysis discloses regional disparities, with substantial changes in the hydrological behavior of northern watersheds (such as the Huma River) and southeastern regions (such as the Toudao River). The augmentation of wetland and forest coverage has effectively mitigated peak runoff, especially during extreme rainfall events. Wetlands have manifested strong water regulation capabilities and alleviated the impact of floods. This study quantitatively discloses the complex response pattern of LUCC to runoff by introducing a multi-scale analysis approach, which furnishes a scientific basis for flood risk assessment, land use optimization, and water resource management, and demonstrates the potential for extensive application in other countries and regions with similar climatic and topographic conditions.

Mining and Climate Change Alters Water Storage and Streamflow Dynamics of Northern Peatland‐Dominated Catchments

AbstractThe Hudson Bay Lowlands (HBL) of northern Ontario, a globally significant carbon store, are characterized by expansive peatland complexes of patterned bogs and fens, which play a vital role in regional water regulation. These peatlands are threatened by disturbance from large‐scale resource extraction and projected climate change, both of which have the potential to compromise their ecohydrological function. Field measurements and numerical modeling were used to investigate the hydrological responses of peatlands and downgradient streamflow as a consequence of disturbance from mining and shifts in climate, individually and in combination. Mine dewatering reduced groundwater storage by as much as 150 mm, equivalent to a water table lowering of 75 cm, thereby decreasing annual streamflow by 66% in impacted tributaries. Although the projected increases to precipitation and evapotranspiration due to climate change were approximately balanced, resulting in minor changes to storage, there were pronounced shifts in the temporal patterns of streamflow, with a diminished snowmelt and spring freshet occurring a month earlier. When considering the cumulative impacts of climate change coupled with mining, a potential shift in peatland ecohydrology toward new equilibria is plausible, implying altered water movement across the landscape and compromised ecosystem function. This study emphasizes the critical need for further monitoring and modeling efforts to characterize the thresholds and mechanisms driving these ecohydrological changes. This research will guide future investigations on the implications of disturbance on local and regional hydrologic connectivity and facilitate the protection of peatland ecosystems in the HBL and other northern peatland‐dominated landscapes.

Effects of Water and Nitrogen Regulation on Soil Environment and Crop Growth in a Lycium barbarum||Alfalfa System.

The increasing scarcity of water and soil resources, combined with inefficient water and fertilizer management, poses significant challenges to agriculture in arid regions. This study aimed to determine an optimal water and nitrogen regulation model to alleviate water shortages and improve agricultural productivity and quality. In this study, a two-year experiment was conducted to investigate the effects of varying irrigation and nitrogen levels on the soil environment and crop growth in a Lycium barbarum||alfalfa system (LB||AS). The experiment involved four moisture gradients and four nitrogen application levels (using urea as the nitrogen source). The results indicated that soil moisture decreased during crop development, followed by a slow increase, with significant variation across soil depths. Soil temperature peaked during the fruiting stage of Lycium barbarum in July, decreasing significantly with soil depth. Higher temperatures were recorded in N0 under the same irrigation level and in W3 under the same nitrogen level. Soil organic carbon (SOC) levels increased by 16.24% in W3N0 and by 18.05% in W2N1, compared to W0N3. Easily oxidizable organic carbon (EOC) and soluble organic carbon (DOC) levels exhibited significant variations depending on irrigation and nitrogen treatments. Irrigation and nitrogen had a stronger individual impact on alfalfa height and stem thickness than their combined effects. Water and nitrogen regulation significantly influenced Lycium barbarum yield, its 100-fruit weight, and economic efficiency (p < 0.05). The W0N2 treatment produced the highest yield (3238 kg·ha-1), exceeding other treatments by up to 29.52%. In conclusion, the optimal water-nitrogen regulation model for the LB||AS system is full irrigation (75-85% θfc) with a nitrogen application rate of 300 kg·ha-1. These findings offer critical insights for improving water and nitrogen management strategies in arid regions.

Understanding water-energy-carbon nexus in English and Welsh water industry by assessing eco-productivity of water companies

Understanding the water-energy-carbon nexus in water supply is essential for water regulators and utilities. This study employs a non-radial Data Envelopment Analysis (DEA) model to assess eco-productivity (ecoP) change, a synthetic indicator that integrates carbon emissions, energy costs, and water delivered. It also evaluates its components—eco-efficiency change and eco-technological change—across water companies in England and Wales from 2011 to 2018. The analysis reveals an annual improvement in ecoP of 1.1%, driven by a 2.1% gain in eco-efficiency but offset by a 1.0% decline in technological advancement. The reduction in GHG emissions emerged as the most significant positive contributor, enhancing ecoP by 3.22% annually, while energy costs detracted ecoP by –0.09%. The results underscore the negative impacts of increased water delivery (–1.74%) and the number of connected properties (–1.27%) on ecoP, highlighting the need for demand management policies.

Study on the effects of extreme climate and human activities on the growth mechanisms of Zostera japonica in the Yellow River estuary

Zostera japonica, as one of the major seagrasses in the Yellow River Estuary, plays a critical ecological role, particularly in providing habitat for marine organisms, stabilizing sediment, and contributing significantly to carbon sequestration. In recent years, Zostera japonica seagrass beds have receded extensively due to multiple impacts of natural factors and human activities. This study investigates the complex effects of extreme climate events and human activities on the growth mechanisms of Zostera japonica in the Yellow River Estuary using a combination of field sampling, laboratory analysis, and quantitative calculations. The result shows that there are significant differences in sediment characteristics between the north and south shores, with the south shore having finer sediments and higher nutrient content, which support more robust seagrass growth. The Water and Sediment Regulation Scheme (WSRS) dramatically alters water quality by reducing salinity and increasing turbidity, thus inhibiting photosynthesis and disrupting the physiological functions of Zostera japonica. Additionally, WSRS introduces an increase in heavy metals, which could potentially impact plant health and stress levels. Extreme weather events, particularly Super Typhoon Lekima, further compound these impacts by causing soil erosion, uprooting seagrass beds, and reducing biomass and seed production. The interplay of WSRS, climate change, and anthropogenic activities necessitates integrated management strategies to mitigate adverse effects and enhance habitat resilience. This study underscores the need for specific management strategies, such as controlling heavy metal inflows, implementing sediment stabilization techniques, and regulating freshwater discharge during key growth periods, to mitigate adverse effects and enhance habitat resilience for Zostera japonica in the Yellow River Estuary.

Transformer-Based hyperspectral image analysis for phenotyping drought tolerance in blueberries

Drought-induced stress significantly impacted blueberry production due to the plants’ inefficient water regulation mechanisms to maintain yield and fruit quality under drought stress. Traditional methods of manual phenotyping for drought stress are not only time-consuming but also labor-intensive. To address the need for accurate and large-scale assessment of drought tolerance, we developed a high-throughput phenotyping (HTP) system to capture hyperspectral images of blueberry plants under drought conditions. A novel transformer-based model, LWC-former was introduced to predict leaf water content (LWC) utilizing spectral reflectance from hyperspectral images obtained from the developed HTP system. The LWC-former transformed the spectral reflectance into patch representations and embedded these patches into a lower dimensional to address multicollinearity issues. These patches were then passed to the transformer encoder to learn distributed features, followed by a regression head to predict LWC. To train the model, spectral reflectance data were extracted from hyperspectral images and pre-processed using log(1/R), mean scatter correction (MSC), and mean centering (MC). The results showed that our model achieved a coefficient of determination (R2) of 0.81 on the test dataset. The performance of the proposed model was also compared with TabTransformer, DeepRWC, multilayer perceptron (MLP), partial least squares regression (PLSR), support vector regression (SVR), and random forest (RF), achieving R2 values of 0.65, 0.73, 0.71, 0.47, and 0.58, respectively. The results demonstrated that LWC-former outperformed other deep learning and statistical-based models. The high-throughput phenotyping system effectively facilitated large-scale data collection, while the LWC-former model addressed multicollinearity issues, significantly improving the prediction of LWC. These results demonstrate the potential of our approach for large-scale drought tolerance assessment in blueberries.

Water and chloride as allosteric inhibitors in WNK kinase osmosensing.

Osmotic stress and chloride regulate the autophosphorylation and activity of the WNK1 and WNK3 kinase domains. The kinase domain of unphosphorylated WNK1 (uWNK1) is an asymmetric dimer possessing water molecules conserved in multiple uWNK1 crystal structures. Conserved waters are present in two networks, referred to here as conserved water networks 1 and 2 (CWN1 and CWN2). Here, we show that PEG400 applied to crystals of dimeric uWNK1 induces de-dimerization. Both the WNK1 the water networks and the chloride-binding site are disrupted by PEG400. CWN1 is surrounded by a cluster of pan-WNK-conserved charged residues. Here, we mutagenized these charges in WNK3, a highly active WNK isoform kinase domain, and WNK1, the isoform best studied crystallographically. Mutation of E314 in the Activation Loop of WNK3 (WNK3/E314Q and WNK3/E314A, and the homologous WNK1/E388A) enhanced the rate of autophosphorylation, and reduced chloride sensitivity. Other WNK3 mutants reduced the rate of autophosphorylation activity coupled with greater chloride sensitivity than wild-type. The water and chloride regulation thus appear linked. The lower activity of some mutants may reflect effects on catalysis. Crystallography showed that activating mutants introduced conformational changes in similar parts of the structure to those induced by PEG400. WNK activating mutations and crystallography support a role for CWN1 in WNK inhibition consistent with water functioning as an allosteric ligand.

Water and chloride as allosteric inhibitors in WNK kinase osmosensing

Osmotic stress and chloride regulate the autophosphorylation and activity of the WNK1 and WNK3 kinase domains. The kinase domain of unphosphorylated WNK1 (uWNK1) is an asymmetric dimer possessing water molecules conserved in multiple uWNK1 crystal structures. Conserved waters are present in two networks, referred to here as conserved water networks 1 and 2 (CWN1 and CWN2). Here, we show that PEG400 applied to crystals of dimeric uWNK1 induces de-dimerization. Both the WNK1 the water networks and the chloride-binding site are disrupted by PEG400. CWN1 is surrounded by a cluster of pan-WNK-conserved charged residues. Here, we mutagenized these charges in WNK3, a highly active WNK isoform kinase domain, and WNK1, the isoform best studied crystallographically. Mutation of E314 in the Activation Loop of WNK3 (WNK3/E314Q and WNK3/E314A, and the homologous WNK1/E388A) enhanced the rate of autophosphorylation, and reduced chloride sensitivity. Other WNK3 mutants reduced the rate of autophosphorylation activity coupled with greater chloride sensitivity than wild-type. The water and chloride regulation thus appear linked. The lower activity of some mutants may reflect effects on catalysis. Crystallography showed that activating mutants introduced conformational changes in similar parts of the structure to those induced by PEG400. WNK activating mutations and crystallography support a role for CWN1 in WNK inhibition consistent with water functioning as an allosteric ligand.