Phosphate transporter PHT1;1 as a key determinant of phosphorus acquisition in Arabidopsis natural accessions

To understand the genetic basis in governing phosphorus (P) acquisition, we performed genome-wide association studies (GWAS) on a diversity panel of Arabidopsis thaliana by two primary determinants of P acquisition, phosphate (Pi)-uptake activity and PHOSPHATE TRANSPORTER 1 (PHT1) protein abundance. Association mapping revealed a shared significant peak on chromosome 5 (Chr5) where the PHT1;1/2/3 genes reside, suggesting a strong correlation between the regulation of Pi-uptake activity and PHT1 protein abundance. Genes encoding transcription factors, kinases, and a metalloprotease associated with both traits were also identified. Conditional GWAS followed by statistical analysis of genotype-dependent expression of PHT1;1 and transcription activity assays revealed an epistatic interaction between PHT1;1 and MYB DOMAIN PROTEIN 52 (MYB52) on Chr1. Analyses of F1 hybrids generated by crossing two subgroups of natural accessions carrying specific SNPs associated with PHT1;1 and MYB52 further revealed the strong effects of potential variants on PHT1;1 expression and Pi uptake activity. Notably, the soil P contents in A. thaliana habitats were found to coincide with PHT1;1 haplotype, underscoring how fine-tuning of the activity of P acquisition by natural variants allows plants to adapt to their environments. This study sheds light on the complex regulation of P acquisition and offers a framework to systematically assess the effectiveness of GWAS approaches in the study of quantitative traits.

In Vitro Evaluation of the Anti-Diabetic Potential of Aqueous Acetone Helichrysumpetiolare Extract (AAHPE) with Molecular Docking Relevance in Diabetes Mellitus

Diabetes mellitus (DM) is a chronic metabolic condition that can lead to significant complications and a high fatality rate worldwide. Efforts are ramping up to find and develop novel α-glucosidase and α-amylase inhibitors that are both effective and potentially safe. Traditional methodologies are being replaced with new techniques that are less complicated and less time demanding; yet, both the experimental and computational strategies are viable and complementary in drug discovery and development. As a result, this study was conducted to investigate the in vitro anti-diabetic potential of aqueous acetone Helichrysum petiolare and B.L Burtt extract (AAHPE) using a 2-NBDG, 2-(N-(7-Nitrobenz-2-oxa-1,3-diazol-4-yl) amino)-2-deoxy-d-glucose uptake assay. In addition, we performed molecular docking of the flavonoid constituents identified and quantified by liquid chromatography-mass spectrometry (LC-MS) from AAHPE with the potential to serve as effective and safe α-amylase and α-glucosidase inhibitors, which are important in drug discovery and development. The results showed that AAHPE is a potential inhibitor of both α-amylase and α-glucosidase, with IC50 values of 46.50 ± 6.17 (µg/mL) and 37.81 ± 5.15 (µg/mL), respectively. This is demonstrated by a significant increase in the glucose uptake activity percentage in a concentration-dependent manner compared to the control, with the highest AAHPE concentration of 75 µg/mL of glucose uptake activity being higher than metformin, a standard anti-diabetic drug, in the insulin-resistant HepG2 cell line. The molecular docking results displayed that the constituents strongly bind α-amylase and α-glucosidase while achieving better binding affinities that ranged from ΔG = −7.2 to −9.6 kcal/mol (compared with acarbose ΔG = −6.1 kcal/mol) for α-amylase, and ΔG = −7.3 to −9.0 kcal/mol (compared with acarbose ΔG = −6.3 kcal/mol) for α-glucosidase. This study revealed the potential use of the H. petiolare plant extract and its phytochemicals, which could be explored to develop potent and safe α-amylase and α-glucosidase inhibitors to treat postprandial glycemic levels in diabetic patients.

The cellular prion protein interacts with and promotes the activity of Na,K-ATPases

The prion protein (PrP) is best known for its ability to cause fatal neurodegenerative diseases in humans and animals. Here, we revisited its molecular environment in the brain using a well-developed affinity-capture mass spectrometry workflow that offers robust relative quantitation. The analysis confirmed many previously reported interactions. It also pointed toward a profound enrichment of Na,K-ATPases (NKAs) in proximity to cellular PrP (PrPC). Follow-on work validated the interaction, demonstrated partial co-localization of the ATP1A1 and PrPC, and revealed that cells exposed to cardiac glycoside (CG) inhibitors of NKAs exhibit correlated changes to the steady-state levels of both proteins. Moreover, the presence of PrPC was observed to promote the ion uptake activity of NKAs in a human co-culture paradigm of differentiated neurons and glia cells, and in mouse neuroblastoma cells. Consistent with this finding, changes in the expression of 5’-nucleotidase that manifest in wild-type cells in response to CG exposure can also be observed in untreated PrPC-deficient cells. Finally, the endoproteolytic cleavage of the glial fibrillary acidic protein, a hallmark of late-stage prion disease, can also be induced by CGs, raising the prospect that a loss of NKA activity may contribute to the pathobiology of prion diseases.

Deltaproteobacterium Strain KaireiS1, a Mesophilic, Hydrogen-Oxidizing and Sulfate-Reducing Bacterium From an Inactive Deep-Sea Hydrothermal Chimney

A novel deltaproteobacterial, mesophilic, hydrogen-oxidizing, and sulfate-reducing bacterium (strain KaireiS1) was highly enriched from an inactive chimney located in the active zone of the Kairei hydrothermal vent field (Central Indian Ridge) in the Indian Ocean. Based on 16S rRNA gene analyses, strain KaireiS1 is the currently only cultured representative of a cluster of uncultured Deltaproteobacteria, positioned within the Desulfobulbaceae family, between the Desulfobulbus genus and the “Cable Bacteria.” A facultative autotrophic lifestyle of KaireiS1 is indicated by its growth in the absence of organic compounds, measurements of CO2-fixation rates, and activity measurements of carbon monoxide dehydrogenase, the key enzyme of the reductive Acetyl-CoA pathway. Apart from hydrogen, strain KaireiS1 can also use propionate, lactate, and pentadecane as electron donors. However, the highest cell numbers were reached when grown autotrophically with molecular hydrogen. Hydrogen uptake activity was found in membrane and soluble fractions of cell-free extracts and reached up to 2,981±129 nmol H2*min−1*mg−1 of partially purified protein. Commonly, autotrophic sulfate-reducing bacteria from the Deltaproteobacteria class, thriving in hydrothermal vent habitats are described as thermophiles. Given its physiological characteristics and specific isolation source, strain KaireiS1 demonstrates a previously unnoticed potential for microbial sulfate reduction by autotrophs taking place at moderate temperatures in hydrothermal vent fields.

Regulation of Copper Metabolism by Nitrogen Utilization in Saccharomyces cerevisiae

To understand the relationship between carbon or nitrogen utilization and iron homeostasis, we performed an iron uptake assay with several deletion mutants with partial defects in carbon or nitrogen metabolism. Among them, some deletion mutants defective in carbon metabolism partially and the MEP2 deletion mutant showed lower iron uptake activity than the wild type. Mep2 is known as a high-affinity ammonia transporter in Saccharomyces cerevisiae. Interestingly, we found that nitrogen starvation resulted in lower iron uptake activity than that of wild-type cells without downregulation of the genes involved in the high-affinity iron uptake system FET3/FTR1. However, the gene expression of FRE1 and CTR1 was downregulated by nitrogen starvation. The protein level of Ctr1 was also decreased by nitrogen starvation, and addition of copper to the nitrogen starvation medium partially restored iron uptake activity. However, the expression of MAC1, which is a copper-responsive transcriptional activator, was not downregulated by nitrogen starvation at the transcriptional level but was highly downregulated at the translational level. Mac1 was downregulated dramatically under nitrogen starvation, and treatment with MG132, which is an inhibitor of proteasome-dependent protein degradation, partially attenuated the downregulation of Mac1. Taken together, these results suggest that nitrogen starvation downregulates the high-affinity iron uptake system by degrading Mac1 in a proteasome-dependent manner and eventually downregulates copper metabolism.

The Effect of Water Level in Rice Cropping System on Phosphorus Uptake Activity of Pup1 in a Pup1+Sub1 Breeding Line

Pyramiding useful QTLs into an elite variety is a promising strategy to develop tolerant varieties against multiple abiotic stresses. However, some QTLs may not be functionally compatible when they are introgressed into the same variety. Here, we tested the functional compatibility of Pup1 and Sub1, major QTLs for tolerance to phosphorus (P)-deficiency and submergence conditions, respectively. Phenotypic analysis revealed that IR64-Pup1+Sub1 (IPS) plants harboring both Pup1 and Sub1 QTLs show significant tolerance to submerged conditions, similarly to IR64-Sub1, while IPS failed to tolerate P deficiency and mild drought conditions; only IR64-Pup1 showed P deficiency tolerance. In submerged conditions, Sub1A and OsPSTOL1, major genes for Sub1 and Pup1 QTLs, respectively, were expressed at the same levels as in IPS and IR64-Sub1 and in IPS and IR64-Pup1, respectively. On the other hand, in P-non-supplied condition, crown root number, root length, and OsPSTOL1 expression level were significantly lower in IPS compared to those of IR64-Pup1. However, there was no significant difference in P content between IPS and IR64-Pup1. These results imply that Pup1 does not compromise Sub1 function in submerged condition, while Sub1 suppresses Pup1 function in P-non-supplied condition, possibly by regulating the transcript level of Pup1. In conclusion, Pup1 and Sub1 are regarded as functionally compatible under submergence condition but not under P-non-supplied condition. Further study is needed to elucidate the functional incompatibility of Pup1 and Sub1 QTLs in IPS under P-non-supplied condition.

A Simple Fluorescence Assay for Cystine Uptake via the xCT in Cells using Selenocystine and a Fluorescent Probe

ABSTRACTThe cystine/glutamate antiporter (xCT) is a crucial transporter that maintains cellular redox balance by regulating intracellular glutathione synthesis via cystine uptake. However, no robust and simple method to determine the cystine uptake activity of xCT is currently available. We have developed a method to measure the xCT activity via the reaction of selenocysteine and fluorescein O,O′-diacrylate (FOdA). Selenocystine, a cystine analog, is transported into cells through xCT on the cell membrane. The amount of the transported selenocystine was then determined by a reaction using tris(2-carboxyethyl)phosphine (TCEP) and FOdA in a weak acidic buffer at pH 6. Using this method, the cystine uptake activity of xCT in various cells, and the inhibitory efficiency of xCT inhibitors, were evaluated.

ZMK1 Is Involved in K+ Uptake and Regulated by Protein Kinase ZmCIPK23 in Zea mays

Potassium (K+) is one of essential mineral elements for plant growth and development. K+ channels, especially AKT1-like channels, play crucial roles in K+ uptake in plant roots. Maize is one of important crops; however, the K+ uptake mechanism in maize is little known. Here, we report the physiological functions of K+ channel ZMK1 in K+ uptake and homeostasis in maize. ZMK1 is a homolog of Arabidopsis AKT1 channel in maize, and mainly expressed in maize root. Yeast complementation experiments and electrophysiological characterization in Xenopus oocytes indicated that ZMK1 could mediate K+ uptake. ZMK1 rescued the low-K+-sensitive phenotype of akt1 mutant and enhanced K+ uptake in Arabidopsis. Overexpression of ZMK1 also significantly increased K+ uptake activity in maize, but led to an oversensitive phenotype. Similar to AKT1 regulation, the protein kinase ZmCIPK23 interacted with ZMK1 and phosphorylated the cytosolic region of ZMK1, activating ZMK1-mediated K+ uptake. ZmCIPK23 could also complement the low-K+-sensitive phenotype of Arabidopsis cipk23/lks1 mutant. These findings demonstrate that ZMK1 together with ZmCIPK23 plays important roles in K+ uptake and homeostasis in maize.