CNNM proteins selectively bind to the TRPM7 channel to stimulate divalent cation entry into cells

Magnesium is essential for cellular life, but how it is homeostatically controlled still remains poorly understood. Here, we report that members of CNNM family, which have been controversially implicated in both cellular Mg2+ influx and efflux, selectively bind to the TRPM7 channel to stimulate divalent cation entry into cells. Coexpression of CNNMs with the channel markedly increased uptake of divalent cations, which is prevented by an inactivating mutation to the channel’s pore. Knockout (KO) of Trpm7 in cells or application of the TRPM7 channel inhibitor NS8593 also interfered with CNNM-stimulated divalent cation uptake. Conversely, KO of CNNM3 and CNNM4 in HEK-293 cells significantly reduced TRPM7-mediated divalent cation entry, without affecting TRPM7 protein expression or its cell surface levels. Furthermore, we found that cellular overexpression of phosphatases of regenerating liver (PRLs), known CNNMs binding partners, stimulated TRPM7-dependent divalent cation entry and that CNNMs were required for this activity. Whole-cell electrophysiological recordings demonstrated that deletion of CNNM3 and CNNM4 from HEK-293 cells interfered with heterologously expressed and native TRPM7 channel function. We conclude that CNNMs employ the TRPM7 channel to mediate divalent cation influx and that CNNMs also possess separate TRPM7-independent Mg2+ efflux activities that contribute to CNNMs’ control of cellular Mg2+ homeostasis.

Enhanced metanephric specification to functional proximal tubule enables toxicity screening and infectious disease modelling in kidney organoids

While pluripotent stem cell-derived kidney organoids represent a promising approach for the study of renal disease, renal physiology and drug screening, the proximal nephron remains immature with limited evidence for key functional solute channels. This may reflect early mispatterning of the nephrogenic mesenchyme or insufficient maturation. In this study, prolonged differentiation and modification of media conditions to enhance metanephric nephron progenitor specification resulted in the induction of nephrons containing elongated and aligned proximal nephron segments together with SLC12A1+ loops of Henle. Nephron proximal segments showed superior HNF4A gene and protein expression, as well as upregulation of key functional transporters, including SLC3A1/2, SLC47A1, and SLC22A2. The striking proximo-distal orientation of nephrons was shown to result from localised WNT antagonism originating from the centre of the organoid. Functionality of such transporters was evidenced by albumin and organic cation uptake, as well as appropriate KIM-1 upregulation in response to the nephrotoxicant, cisplatin. PT-enhanced organoids also possessed improved expression of receptors associated with SARS-CoV2 entry, rendering these organoids susceptible to infection and able to support viral replication without co-location of ACE2 and TMPRSS2. These PT-enhanced organoids provide an accurate model with which to study human proximal tubule maturation, inherited and acquired proximal tubular disease, and drug and viral responses.

Roles of TroA and TroR in Metalloregulated Growth and Gene Expression in Treponema denticola

ABSTRACT The availability of divalent metal cations required as cofactors for microbial metabolism is severely limited in the host environment. Bacteria have evolved highly regulated uptake systems to maintain essential metal homeostasis to meet cellular demands while preventing toxicity. The Tro operon (troABCDR), present in all sequenced Treponema spp., is a member of a highly conserved family of ATP-binding cassette transporters involved in metal cation uptake whose expression is controlled by TroR, a DtxR-like cation-responsive regulatory protein. Transcription of troA responds to divalent manganese and iron (T. denticola) or manganese and zinc (T. pallidum), and metal-dependent TroR binding to the troA promoter represses troA transcription. We report here the construction and complementation of defined T. denticola ΔtroR and ΔtroA strains to characterize (i) the role of TroA in metal-dependent T. denticola growth and (ii) the role of TroR in T. denticola gene expression. We show that TroA expression is required for T. denticola growth under iron- and manganese-limited conditions. Furthermore, TroR is required for the transcriptional regulation of troA in response to iron or manganese, and deletion of troR results in significant differential expression of more than 800 T. denticola genes in addition to troA. These results suggest that (i) TroA-mediated cation uptake is important in metal homeostasis in vitro and may be important for Treponema survival in the host environment and (ii) the absence of TroR results in significant dysregulation of nearly one-third of the T. denticola genome. These effects may be direct (as with troA) or indirect due to dysregulation of metal homeostasis. IMPORTANCE Treponema denticola is one of numerous host-associated spirochetes, a group including commensals, pathobionts, and at least one frank pathogen. While most T. denticola research concerns its role in periodontitis, its relative tractability for growth and genetic manipulation make it a useful model for studying Treponema physiology, metabolism, and host-microbe interactions. Metal micronutrient acquisition and homeostasis are highly regulated both in microbial cells and by host innate defense mechanisms that severely limit metal cation bioavailability. Here, we characterized the T. denticola troABCDR operon, the role of TroA-mediated iron and manganese uptake in growth, and the effects of TroR on global gene expression. This study contributes to our understanding of the mechanisms involved in cellular metal homeostasis required for survival in the host environment.

On the pH-dependent strontium adsorption by Al-PILCs, impregnated with carboxylate groups of various acid strengths

The impregnation of a PILC with carboxylate groups is proposed as a means to improve the adsorption properties of the material for strontium ions. The role of solution pH and carboxylate acid strength in the functionalization of the organic groups is probed by base titrations. In comparing the pristine-PILC with PILCs carrying either oxalate or acetate impregnates, enhanced strontium uptake is evidenced by the modified solids, initiated at pH 6 and 8 in the presence of oxalate and acetate, respectively. The effect on uptake is higher in the former case, amounting to an increase in strontium adsorption by a factor between two and three, depending on the pH range. To further elucidate the significance of the carboxylate acid strength in cation uptake phenomena, strontium adsorption isotherms are presented for PILCs carrying acetate, oxalate, malonate and citrate groups. The results demonstrate that, at constant pH, adsorption increases with increasing acid strength

Growth and K Nutrition of Sesame (Sesamum indicum L.) Seedlings as Affected by Balancing Soil Exchangeable Cations Ca, Mg, and K of Continuously Monocropped Soil from Upland Fields Converted Paddy

Growth of sesame is known to be limited by poor K nutrition as a result of imbalance in soil exchangeable cations that cause a competitive ion effect in continuous monocropping from upland fields converted paddy. We hypothesized that balancing soil exchangeable cations will improve the K nutrition and growth of sesame plants. Therefore, the specific objectives of this study were to determine the effect of balancing soil exchangeable cations Ca, Mg, and K of continuously monocropped soils on the growth and cation uptake of sesame seedlings and also identify a suitable source of nutrients for improving K nutrition. A pot experiment was conducted under greenhouse condition in a 3 × 3 factorial design consisting of three levels of balancing treatments i.e. inorganic fertilizer for Ca, Mg, and K, rice husk biochar to increase K content, and the three durations of continuous monocropping soils of one year, two years, and four years from upland fields converted paddy. Balancing soil exchangeable cations was aimed at achieving optimal base saturations (CaO, 75%; MgO, 25%; and K2O, 10%). Results showed that balancing exchangeable cations did not significantly affect growth and cation uptake in the one and two-year soils but significant effect was observed in the four-year soil. Overall, plant height and dry weight increased for the balancing treatments of inorganic fertilizer K and rice husk biochar. Balancing exchangeable cations with biochar was more beneficial than with inorganic fertilizers. The four-year soil’s growth increase was attributed to an increase in K concentration and uptake due to the decrease in the soil Ca/K and Mg/K ratios to that of acceptable levels, which eliminated competitive ion effect as the soil K saturation increased above 5.0%, enhancing sesame growth. Therefore, a balanced soil exchangeable Ca, Mg, and K that eliminates a competitive ion effect will improve sesame growth and K nutrition although future research should focus on ensuring balanced cation rations under field conditions in continuous monocropping.

Quantifying the Acidic and Basic Effects of Vegetable and Herb Species in Peat-based Substrate and Hydroponics

Objectives were 1) to quantify acidic and basic effects on the root zone pH for eight vegetable and herb species grown in peat-based substrate and hydroponic nutrient solution and 2) to determine the applied NH4+:NO3– ratio expected to have a neutral pH reaction for each species during its vegetative growth phase. In one experiment, plants were grown for 33 days in substrate (70% peat:30% perlite by volume), and were fertilized with a nutrient solution containing 7.14 milli-equivalents (mEq)·L–1 N and NH4+:NO3– ratios ranging from 0:100 to 40:60. During the second experiment, the same species were grown in hydroponic nutrient solutions at 7.14 mEq·L–1 N with NH4+:NO3– ratios ranging from 0:100 to 30:70, and data were collected over a 6-day period. In substrate, species increased root zone pH when supplied 0:100 solution, except for cucumber, which did not change substrate pH. Increasing the NH4+:NO3– ratio to 40:60 increased acidity and decreased pH across species. Similar trends were observed in hydroponics, in which the most basic response occurred across species with 0:100, and the most acidic response occurred with 30:70. Arugula was the only species that increased root zone pH with all three NH4+:NO3– ratios in substrate and hydroponics. In substrate and hydroponics, mEq of acidity (negative) or basicity (positive) produced per gram dry weight gain per plant (mEq·g−1) correlated positively with mEq·g−1 net cation minus anion uptake, respectively, in which greater cation uptake resulted in acidity and greater anion uptake resulted in basicity. In hydroponics, the greatest net anion uptake occurred with 0:100, and increasing the NH4+:NO3– ratio increased total cation uptake across species. Cucumber had the most acidic effect and required less than 10% of N as NH4+-N for a neutral pH over time, arugula was the most basic and required more than 20% NH4+-N, and the remaining species had neutral percent NH4+-N between 10% and 20% of N. Increasing the NH4+:NO3– ratio decreased Ca2+ uptake across all species in hydroponics, which could potentially impact tip burn and postharvest quality negatively. Controlling root zone pH in substrate and hydroponic culture requires regular pH monitoring in combination with NH4+:NO3– adjustments and other pH management strategies, such as injecting mineral acid to neutralize irrigation water alkalinity or adjusting the limestone incorporation rate for substrate.

Frontiers and progress in cation-uptake and exchange chemistry of polyoxometalate-based compounds

Reduction-induced cation-uptake in POM-based ionic solids.