Empagliflozin in children with glycogen storage disease-associated inflammatory bowel disease: a prospective, single-arm, open-label clinical trial.

Glycogen storage disease type Ib (GSD-Ib) is a rare inborn error of glycogen metabolism caused by mutations in SLC37A4. Patients with GSD-Ib are at high risk of developing inflammatory bowel disease (IBD). We evaluated the efficacy of empagliflozin, a renal sodium‒glucose cotransporter protein 2 (SGLT2) inhibitor, on colonic mucosal healing in patients with GSD-associated IBD. A prospective, single-arm, open-label clinical trial enrolled eight patients with GSD-associated IBD from Guangdong Provincial People's Hospital in China from July 1, 2022 through December 31, 2023. Eight patients were enrolled with a mean age of 10.34 ± 2.61 years. Four male and four female. The endoscopic features included deep and large circular ulcers, inflammatory hyperplasia, obstruction and stenosis. The SES-CD score significantly decreased at week 48 compared with before empagliflozin. Six patients completed 48 weeks of empagliflozin therapy and endoscopy showed significant improvement or healing of mucosal ulcers, inflammatory hyperplasia, stenosis, and obstruction. One patient had severe sweating that required rehydration and developed a urinary tract infection. No serious or life-threatening adverse events. This study suggested that empagliflozin may promote colonic mucosal healing and reduce hyperplasia, stenosis, and obstruction in children with GSD-associated IBD.

Functional activity of endophytic bacteria G9H01 with high salt tolerance and anti-Magnaporthe oryzae that isolated from saline-alkali-tolerant rice.

It holds significant practical importance to screen and investigate endophytic bacteria with salt-tolerant activity in rice for the development of relevant microbial agents. A total of 179 strains of endophytic bacteria were isolated from 24 samples of salt-tolerant rice seeds, with almost 95 % of these bacteria exhibiting tolerance to a salt content of 2 % (0.34 mol/L). Following the screening process, a bacterium named G9H01 was identified, which demonstrated a salt tolerance of up to 15 % (2.57 mol/L) and resistance to Magnaporthe oryzae, the causal agent of rice blast disease. Phylogenetic analysis confirmed G9H01 as a strain of Bacillus paralicheniformis. The complete genome of G9H01 was sequenced and assembled, revealing a considerable number of genes encoding proteins associated with salt tolerance. Further analysis indicated that G9H01 may alleviate salt stress in a high-salt environment through various mechanisms. These mechanisms include the utilization of proteins such as K+ transporters, antiporters, and Na+/H+ antiporters, which are involved in K+ absorption and Na+ excretion. G9H01 also demonstrated the ability to uptake and accumulate betaine, as well as secrete extracellular polysaccharides. Collectively, these findings suggest that Bacillus paralicheniformis G9H01 has potential as a biocontrol agent, capable of promoting rice growth under saline-alkali-tolerant conditions.

Genome-wide characterization of the CPA gene family in potato and a preliminary functional analysis of its role in NaCl tolerance.

BACKGROUND: The cation/proton antiporter (CPA) superfamily plays a crucial role in regulating ion homeostasis and pH in plant cells, contributing to stress resistance. However, in potato (Solanum tuberosum L.), systematic identification and analysis of CPA genes are lacking. RESULTS: A total of 33 StCPA members were identified and classified into StNHX (n = 7), StKEA (n = 6), and StCHX (n = 20) subfamilies. StCHX owned the highest number of conserved motifs, followed by StKEA and StNHX. The StNHX and StKEA subfamilies owned more exons than StCHX. NaCl stress induced the differentially expression of 19 genes in roots or leaves, among which StCHX14 and StCHX16 were specifically induced in leaves, while StCHX2 and StCHX19 were specifically expressed in the roots. A total of 11 strongly responded genes were further verified by qPCR. Six CPA family members, StNHX1, StNHX2, StNHX3, StNHX5, StNHX6 and StCHX19, were proved to transport Na+ through yeast complementation experiments. CONCLUSIONS: This study provides comprehensive insights into StCPAs and their response to NaCl stress, facilitating further functional characterization.

A mathematical model of ENaC and Slc26a6 regulation by CFTR in salivary gland ducts.

Cystic fibrosis (CF) is a genetic disease caused by the mutations of cystic fibrosis transmembrane conductance regulator (CFTR), the cystic fibrosis transmembrane conductance regulator gene. Cftr is a critical ion channel expressed in the apical membrane of mouse salivary gland striated duct cells. Although Cftr is primarily a Cl- channel, its knockout leads to higher salivary Cl- and Na+ concentrations and lower pH. Mouse experiments show that the activation of Cftr upregulates epithelial Na+ channel (ENaC) protein expression level and Slc26a6 (a 1Cl-:2[Formula: see text] exchanger of the solute carrier family) activity. Experimentally, it is difficult to predict how much the coregulation effects of CFTR contribute to the abnormal Na+, Cl-, and [Formula: see text] concentrations and pH in CF saliva. To address this question, we construct a wild-type mouse salivary gland model and simulate CFTR knockout by altering the expression levels of CFTR, ENaC, and Slc26a6. By reproducing the in vivo and ex vivo final saliva measurements from wild-type and CFTR knockout animals, we obtain computational evidence that ENaC and Slc26a6 activities are downregulated in CFTR knockout in salivary glands.NEW & NOTEWORTHY This paper describes a salivary gland mathematical model simulating the ion exchange between saliva and the salivary gland duct epithelium. The novelty lies in the implementation of CFTR regulating ENaC and Slc26a6 in a CFTR knockout gland. By reproducing the experimental saliva measurements in wild-type and CFTR knockout glands, the model shows that CFTR regulates ENaC and Slc26a6 anion exchanger in salivary glands. The method could be used to understand the various cystic fibrosis phenotypes.

Structural and functional insights into the lipid regulation of human anion exchanger 2.

Anion exchanger 2 (AE2) is an electroneutral Na+-independent Cl-/HCO3- exchanger belongs to the SLC4 transporter family. The widely expressed AE2 participates in a variety of physiological processes, including transepithelial acid-base secretion and osteoclastogenesis. Both the transmembrane domains (TMDs) and the N-terminal cytoplasmic domain (NTD) are involved in regulation of AE2 activity. However, the regulatory mechanism remains unclear. Here, we report a 3.2 Å cryo-EM structure of the AE2 TMDs in complex with PIP2 and a 3.3 Å full-length mutant AE2 structure in the resting state without PIP2. We demonstrate that PIP2 at the TMD dimer interface is involved in the substrate exchange process. Mutation in the PIP2 binding site leads to the displacement of TM7 and further stabilizes the interaction between the TMD and the NTD. Reduced substrate transport activity and conformation similar to AE2 in acidic pH indicating the central contribution of PIP2 to the function of AE2.

Investigation of the functional impact of CHED- and FECD4-associated SLC4A11 mutations in human corneal endothelial cells.

Mutations in the solute linked carrier family 4 member 11 (SLC4A11) gene are associated with congenital hereditary endothelial dystrophy (CHED) and Fuchs corneal endothelial dystrophy type 4 (FECD4), both characterized by corneal endothelial cell (CEnC) dysfunction and/or cell loss leading to corneal edema and visual impairment. In this study, we characterize the impact of CHED-/FECD4-associated SLC4A11 mutations on CEnC function and SLC4A11 protein localization by generating and comparing human CEnC (hCEnC) lines expressing wild type SLC4A11 (SLC4A11WT) or mutant SLC4A11 harboring CHED-/FECD4-associated SLC4A11 mutations (SLC4A11MU). SLC4A11WT and SLC4A11MU hCEnC lines were generated to express either SLC4A11 variant 2 (V2WT and V2MU) or variant 3 (V3WT and V3MU), the two major variants expressed in ex vivo hCEnC. Functional assays were performed to assess cell barrier, proliferation, viability, migration, and NH3-induced membrane conductance. We demonstrate SLC4A11-/- and SLC4A11MU hCEnC lines exhibited increased migration rates, altered proliferation and decreased cell viability compared to SLC4A11WT hCEnC. Additionally, SLC4A11-/- hCEnC demonstrated decreased cell-substrate adhesion and membrane capacitances compared to SLC4A11WT hCEnC. Induction with 10mM NH4Cl led SLC4A11WT hCEnC to depolarize; conversely, SLC4A11-/- hCEnC hyperpolarized and the majority of SLC4A11MU hCEnC either hyperpolarized or had minimal membrane potential changes following NH4Cl induction. Immunostaining of primary hCEnC and SLC4A11WT hCEnC lines for SLC4A11 demonstrated predominately plasma membrane staining with poor or partial colocalization with mitochondrial marker COX4 within a subset of punctate subcellular structures. Overall, our findings suggest CHED-associated SLC4A11 mutations likely lead to hCEnC dysfunction, and ultimately CHED, by interfering with cell migration, proliferation, viability, membrane conductance, barrier function, and/or cell surface localization of the SLC4A11 protein in hCEnC. Additionally, based on their similar subcellular localization and exhibiting similar cell functional profiles, protein isoforms encoded by SLC4A11 variant 2 and variant 3 likely have highly overlapping functional roles in hCEnC.

Functional promiscuity of small multidrug resistance transporters from Staphylococcus aureus, Pseudomonas aeruginosa, and Francisella tularensis.

Small multidrug resistance transporters efflux toxic compounds from bacteria and are a minimal system to understand multidrug transport. Most previous studies have focused on EmrE, the model SMR from Escherichia coli, finding that EmrE has a broader substrate profile than previously thought and that EmrE may perform multiple types of transport, resulting in substrate-dependent resistance or susceptibility. Here, we performed a broad screen to identify potential substrates of three other SMRs: PAsmr from Pseudomonas aeruginosa; FTsmr from Francisella tularensis; and SAsmr from Staphylococcus aureus. This screen tested metabolic differences in E. coli expressing each transporter versus an inactive mutant, for a clean comparison of sequence and substrate-specific differences in transporter function, and identified many substrates for each transporter. In general, resistance compounds were charged, and susceptibility substrates were uncharged, but hydrophobicity was not correlated with phenotype. Two resistance hits and two susceptibility hits were validated via growth assays and IC50 calculations. Susceptibility is proposed to occur via substrate-gated proton leak, and the addition of bicarbonate antagonizes the susceptibility phenotype, consistent with this hypothesis.

Sequential removal of cation/H+ exchangers reveals their additive role in elemental distribution, calcium depletion and anoxia tolerance.

Multiple Arabidopsis H+ /Cation exchangers (CAXs) participate in high-capacity transport into the vacuole. Previous studies have analysed single and double mutants that marginally reduced transport; however, assessing phenotypes caused by transport loss has proven enigmatic. Here, we generated quadruple mutants (cax1-4: qKO) that exhibited growth inhibition, an 85% reduction in tonoplast-localised H+ /Ca transport, and enhanced tolerance to anoxic conditions compared to CAX1 mutants. Leveraging inductively coupled plasma mass spectrometry (ICP-MS) and synchrotron X-ray fluorescence (SXRF), we demonstrate CAX transporters work together to regulate leaf elemental content: ICP-MS analysis showed that the elemental concentrations in leaves strongly correlated with the number of CAX mutations; SXRF imaging showed changes in element partitioning not present in single CAX mutants and qKO had a 40% reduction in calcium (Ca) abundance. Reduced endogenous Ca may promote anoxia tolerance; wild-type plants grown in Ca-limited conditions were anoxia tolerant. Sequential reduction of CAXs increased mRNA expression and protein abundance changes associated with reactive oxygen species and stress signalling pathways. Multiple CAXs participate in postanoxia recovery as their concerted removal heightened changes in postanoxia Ca signalling. This work showcases the integrated and diverse function of H+ /Cation transporters and demonstrates the ability to improve anoxia tolerance through diminishing endogenous Ca levels.

Genome-wide identification of the cation/proton antiporter (CPA) gene family and functional characterization of the key member BnaA05.NHX2 in allotetraploid rapeseed.

Rapeseed (Brassica napus L.) is susceptible to nutrient stresses during growth and development; however, the CPA (cation proton antiporter) family genes have not been identified in B. napus and their biological functions remain unclear. This study was aimed to identify the molecular characteristics of rapeseed CPAs and their transcriptional responses to multiple nutrient stresses. Through bioinformatics analysis, 117 BnaCPAs, consisting of three subfamilies: Na+/H+ antiporter (NHX), K+ efflux antiporter (KEA), and cation/H+ antiporter (CHX), were identified in the rapeseed genome. Transcriptomic profiling showed that BnaCPAs, particularly BnaNHXs, were transcriptionally responsive to diverse nutrient stresses, including Cd toxicity, K starvation, salt stress, NH4+ toxicity, and low Pi. We found that the salt tolerance of the transgenic rapeseed lines overexpressing BnaA05.NHX2 was significantly higher than that of wild type. Subcellular localization showed that BnaA05.NHX2 was localized on the tonoplast, and TEM combined with X-ray energy spectrum analysis revealed that the vacuolar Na+ concentrations of the BnaA05.NHX2-overexpressing rapeseed plants were significantly higher than those of wild type. The findings of this study will provide insights into the complexity of the BnaCPA family and a valuable resource to explore the in-depth functions of CPAs in B. napus.

Genome-Wide Analysis of Cation/Proton Antiporter Family in Soybean (Glycine max) and Functional Analysis of GmCHX20a on Salt Response.

Monovalent cation proton antiporters (CPAs) play crucial roles in ion and pH homeostasis, which is essential for plant development and environmental adaptation, including salt tolerance. Here, 68 CPA genes were identified in soybean, phylogenetically dividing into 11 Na+/H+ exchangers (NHXs), 12 K+ efflux antiporters (KEAs), and 45 cation/H+ exchangers (CHXs). The GmCPA genes are unevenly distributed across the 20 chromosomes and might expand largely due to segmental duplication in soybean. The GmCPA family underwent purifying selection rather than neutral or positive selections. The cis-element analysis and the publicly available transcriptome data indicated that GmCPAs are involved in development and various environmental adaptations, especially for salt tolerance. Based on the RNA-seq data, twelve of the chosen GmCPA genes were confirmed for their differentially expression under salt or osmotic stresses using qRT-PCR. Among them, GmCHX20a was selected due to its high induction under salt stress for the exploration of its biological function on salt responses by ectopic expressing in Arabidopsis. The results suggest that the overexpression of GmCHX20a increases the sensitivity to salt stress by altering the redox system. Overall, this study provides comprehensive insights into the CPA family in soybean and has the potential to supply new candidate genes to develop salt-tolerant soybean varieties.

Three novel SLC37A4 variants in glycogen storage disease type 1b and a literature review.

Glycogen storage disease type 1b (GSD1b) is a rare genetic disorder, resulting from mutations in the SLC37A4 gene located on chromosome 11q23.3. Although the SLC37A4 gene has been identified as the pathogenic gene for GSD1b, the complete variant spectrum of this gene remains to be fully elucidated. In this study, we present three patients diagnosed with GSD1b through genetic testing. We detected five variants of the SLC37A4 gene in these three patients, with three of these mutations (p. L382Pfs*15, p. G117fs*28, and p. T312Sfs*13) being novel variants not previously reported in the literature. We also present a literature review and general overview of the currently reported SLC37A4 gene variants. Our study expands the mutation spectrum of SLC37A4, which may help enable genetic testing to facilitate prompt diagnosis, appropriate intervention, and genetic counseling for affected families.

Slc26a1 is not essential for spermatogenesis and male fertility in mice.

Thousands of genes are expressed in the testis of mice. However, the details about their roles during spermatogenesis have not been well-clarified for most genes. The purpose of this study was to examine the effect of Slc26a1 deficiency on mouse spermatogenesis and male fertility. Slc26a1-knockout (KO) mice were generated using CRISPR/Cas9 technology on C57BL/6J background. We found no obvious differences between Slc26a1-KO and Slc26a1-WT mice in fertility tests, testicular weight, sperm concentrations, or morphology. Histological analysis found that Slc26a1-KO mouse testes had normal germ cell types and mature sperm. These findings indicated that Slc26a1 was dispensable for male fertility in mice. Our results may save time and resources by allowing other researchers to focus on genes that are more meaningful for fertility studies. We also found that mRNAs of two Slc26a family members (Slc26a5 and Slc26a11) were expressed on higher mean levels in Slc26a1-KO total mouse testes, compared to Slc26a1-WT mice. This effect was not found in mouse GC-1 and GC-2 germ cell lines with the Slc26a1 gene transiently knocked down. This result may indicate that a gene compensation phenomenon was present in the testes of Slc26a1-KO mice.

A molecular signature for the G6PC3/SLC37A2/SLC37A4 interactors in glioblastoma disease progression and in the acquisition of a brain cancer stem cell phenotype.

Background: Glycogen plays an important role in glucose homeostasis and contributes to key functions related to brain cancer cell survival in glioblastoma multiforme (GBM) disease progression. Such adaptive molecular mechanism is dependent on the glycogenolytic pathway and intracellular glucose-6-phosphate (G6P) sensing by brain cancer cells residing within those highly hypoxic tumors. The involvement of components of the glucose-6-phosphatase (G6Pase) system remains however elusive. Objective: We questioned the gene expression levels of components of the G6Pase system in GBM tissues and their functional impact in the control of the invasive and brain cancer stem cells (CSC) phenotypes. Methods: In silico analysis of transcript levels in GBM tumor tissues was done by GEPIA. Total RNA was extracted and gene expression of G6PC1-3 as well as of SLC37A1-4 members analyzed by qPCR in four human brain cancer cell lines and from clinically annotated brain tumor cDNA arrays. Transient siRNA-mediated gene silencing was used to assess the impact of TGF-ß-induced epithelial-to-mesenchymal transition (EMT) and cell chemotaxis. Three-dimensional (3D) neurosphere cultures were generated to recapitulate the brain CSC phenotype. Results: Higher expression in G6PC3, SLC37A2, and SLC37A4 was found in GBM tumor tissues in comparison to low-grade glioma and healthy tissue. The expression of these genes was also found elevated in established human U87, U251, U118, and U138 GBM cell models compared to human HepG2 hepatoma cells. SLC37A4/G6PC3, but not SLC37A2, levels were induced in 3D CD133/SOX2-positive U87 neurospheres when compared to 2D monolayers. Silencing of SLC37A4/G6PC3 altered TGF-ß-induced EMT biomarker SNAIL and cell chemotaxis. Conclusion: Two members of the G6Pase system, G6PC3 and SLC37A4, associate with GBM disease progression and regulate the metabolic reprogramming of an invasive and CSC phenotype. Such molecular signature may support their role in cancer cell survival and chemoresistance and become future therapeutic targets.

Structural insights into the conformational changes of BTR1/SLC4A11 in complex with PIP2.

BTR1 (SLC4A11) is a NH3 stimulated H+ (OH-) transporter belonging to the SLC4 family. Dysfunction of BTR1 leads to diseases such as congenital hereditary endothelial dystrophy (CHED) and Fuchs endothelial corneal dystrophy (FECD). However, the mechanistic basis of BTR1 activation by alkaline pH, transport activity regulation and pathogenic mutations remains elusive. Here, we present cryo-EM structures of human BTR1 in the outward-facing state in complex with its activating ligands PIP2 and the inward-facing state with the pathogenic R125H mutation. We reveal that PIP2 binds at the interface between the transmembrane domain and the N-terminal cytosolic domain of BTR1. Disruption of either the PIP2 binding site or protonation of PIP2 phosphate groups by acidic pH can transform BTR1 into an inward-facing conformation. Our results provide insights into the mechanisms of how the transport activity and conformation changes of BTR1 are regulated by PIP2 binding and interaction of TMD and NTD.

Pathogenicity and Function Analysis of Two Novel SLC4A11 Variants in Patients With Congenital Hereditary Endothelial Dystrophy.

Purpose: The purpose of this study was to explore the pathogenicity and function of two novel SLC4A11 variants associated with congenital hereditary endothelial dystrophy (CHED) and to study the function of a SLC4A11 (K263R) mutant in vitro. Methods: Ophthalmic examinations were performed on a 28-year-old male proband with CHED. Whole-exome and Sanger sequencing were applied for mutation screening. Bioinformatics and pathogenicity analysis were performed. HEK293T cells were transfected with the plasmids of empty vector, wild-type SLC4A11, and SLC4A11 (K263R) mutant. The transfected cells were treated with SkQ1. Oxygen consumption, cellular reactive oxygen species (ROS) level, mitochondrial membrane potential, and apoptosis rate were measured. Results: The proband had poor visual acuity with nystagmus since childhood. Corneal foggy opacity was evident in both eyes. Two novel SLC4A11 variants were detected. Sanger sequencing showed that the proband's father and sister carried c.1464-1G>T variant, and the proband's mother and sister carried c.788A>G (p.Lys263Arg) variant. Based on the American College of Medical Genetics (ACMG) guidelines, SLC4A11 c.1464-1G>T was pathogenic, whereas c.788A>G, p.K263R was a variant of undetermined significance. In vitro, SLC4A11 (K263R) variant increased ROS level and apoptosis rate. Decrease in mitochondrial membrane potential and oxygen consumption rate were remarkable. Furthermore, SkQ1 decreased ROS levels and apoptosis rate but increased mitochondrial membrane potential in the transfected cells. Conclusions: Two novel heterozygous pathogenic variants of the SLC4A11 gene were identified in a family with CHED. The missense variant SLC4A11 (K263R) caused mitochondrial dysfunction and increased apoptosis in mutant transfected cells. In addition, SkQ1 presented a protective effect suggesting the anti-oxidant might be a novel therapeutic drug. Translational Relevance: This study verified the pathogenicity of 2 novel variants in the SLC4A11 gene in a CHED family and found an anti-oxidant might be a new drug.

The vacuolar Ca2+ transporter CATION EXCHANGER 2 regulates cytosolic calcium homeostasis, hypoxic signaling, and response to flooding in Arabidopsis thaliana.

Flooding represents a major threat to global agricultural productivity and food security, but plants are capable of deploying a suite of adaptive responses that can lead to short- or longer-term survival to this stress. One cellular pathway thought to help coordinate these responses is via flooding-triggered Ca2+ signaling. We have mined publicly available transcriptomic data from Arabidopsis subjected to flooding or low oxygen stress to identify rapidly upregulated, Ca2+ -related transcripts. We then focused on transporters likely to modulate Ca2+ signals. Candidates emerging from this analysis included AUTOINHIBITED Ca2+ ATPASE 1 and CATION EXCHANGER 2. We therefore assayed mutants in these genes for flooding sensitivity at levels from growth to patterns of gene expression and the kinetics of flooding-related Ca2+ changes. Knockout mutants in CAX2 especially showed enhanced survival to soil waterlogging coupled with suppressed induction of many marker genes for hypoxic response and constitutive activation of others. CAX2 mutants also generated larger and more sustained Ca2+ signals in response to both flooding and hypoxic challenges. CAX2 is a Ca2+ transporter located on the tonoplast, and so these results are consistent with an important role for vacuolar Ca2+ transport in the signaling systems that trigger flooding response.

Characterization of Two Na+(K+, Li+)/H+ Antiporters from Natronorubrum daqingense.

The Na+/H+ antiporter NhaC family protein is a kind of Na+/H+ exchanger from the ion transporter (IT) superfamily, which has mainly been identified in the halophilic bacteria of Bacillus. However, little is known about the Na+/H+ antiporter NhaC family of proteins in the extremely halophilic archaea. In this study, two Na+/H+ antiporter genes, nhaC1 and nhaC2, were screened from the genome of Natronorubrum daqingense based on the gene library and complementation of salt-sensitive Escherichia coli KNabc. A clone vector pUC18 containing nhaC1 or nhaC2 could make KNabc tolerate 0.6 M/0.7 M NaCl or 30 mM/40 mM LiCl and a pH of up to 8.5/9.5, respectively. Functional analysis shows that the Na+(K+, Li+)/H+ antiport activities of NhaC1 and NhaC2 are both pH-dependent in the range of pH 7.0-10.0, and the optimal pH is 9.5. Phylogenetic analysis shows that both NhaC1 and NhaC2 belong to the Na+/H+ antiporter NhaC family of proteins and are significantly distant from the identified NhaC proteins from Bacillus. In summary, we have identified two Na+(K+, Li+)/H+ antiporters from N. daqingense.

The revolution of personalized pharmacotherapies for cystic fibrosis: what does the future hold?

INTRODUCTION: Cystic fibrosis (CF), a potentially fatal genetic disease, is caused by loss-of-function mutations in the gene encoding for the CFTR chloride/bicarbonate channel. Modulator drugs rescuing mutant CFTR traffic and function are now in the clinic, providing unprecedented breakthrough therapies for people with CF (PwCF) carrying specific genotypes. However, several CFTR variants are unresponsive to these therapies. AREA COVERED: We discussed several therapeutic approaches that are under development to tackle the fundamental cause of CF, including strategies targeting defective CFTR mRNA and/or protein expression and function. Alternatively, defective chloride secretion and dehydration in CF epithelia could be restored by exploiting pharmacological modulation of alternative targets, i.e., ion channels/transporters that concur with CFTR to maintain the airway surface liquid homeostasis (e.g., ENaC, TMEM16A, SLC26A4, SLC26A9, and ATP12A). Finally, we assessed progress and challenges in the development of gene-based therapies to replace or correct the mutant CFTR gene. EXPERT OPINION: CFTR modulators are benefiting many PwCF responsive to these drugs, yielding substantial improvements in various clinical outcomes. Meanwhile, the CF therapy development pipeline continues to expand with the development of novel CFTR modulators and alternative therapeutic strategies with the ultimate goal of providing effective therapies for all PwCF in the foreseeable future.

Gene knockout of NHX-3 in the soil nematode Caenorhabditis elegans leads to broad-spectrum compensatory regulation of Na+/H+ exchangers, antiporters, and the V-type H+-ATPase.

Na+/H+ exchangers are directly involved in a variety of an animal's essential physiological processes such as ionoregulation, acid-base regulation, nitrogenous waste excretion, and nutrient absorption. While nine NHX isoforms have been identified in Caenorhabditis elegans, the physiological importance of each isoform is not understood. The current study aimed to further our knowledge of NHX-3 which has previously been suggested to be involved in the movement of ammonia and acid-base equivalents across the nematode's hypodermis. Although NHX-3 knockout mutant nematodes exported H+ and imported Na+ at slower rates than wild-type nematodes, attempts to inhibit the NHX activity of mutant nematodes using amiloride and EIPA caused an unexpected increase in hypodermal H+ export and did not impact Na+ fluxes suggesting that the different H+ and Na+ transport profiles of the nematodes are likely due to compensatory changes in the mutants in response to the NHX-3 knockout, rather than the loss of NHX-3's physiological function. Significant changes in the mRNA expression of 7 other NHX isoforms, 2 Na+/H+ antiporter isoforms, and the V-type H+-ATPase were detected between wild-type and mutant nematodes. Furthermore, mutant nematodes possessed significantly reduced rates of cytochrome C oxidase activity and ammonia excretion rates, indicating the knockout of NHX-3 induced fundamental changes in metabolism that could impact the nematode's need to eliminate metabolic end-products like H+ and ammonia that relate to NHX transport. While C. elegans is a popular genetic model with cheap and accessible commercial mutants, our findings suggest caution in interpretation of results in studies using mutants to study physiological traits and the biological significance of specific transporters.

A Na+/H+ antiporter-encoding salt overly sensitive 1 gene, LpSOS1, involved in positively regulating the salt tolerance in Lilium pumilum.

Lilium pumilum has a strong salt tolerance. However, the molecular mechanism underlying its salt tolerance remains unexplored. Here, LpSOS1 was cloned from L. pumilum and found to be significantly enriched at high NaCl concentrations (100 mM). In tobacco epidermal cells, localization analysis showed that the LpSOS1 protein was primarily located in the plasma membrane. Overexpression of LpSOS1 resulted in up-regulation of salt stress tolerance in Arabidopsis, as indicated by reduced malondialdehyde levels and Na+/K+ ratio, and increased activity of antioxidant reductases (including superoxide dismutase, peroxidase, and catalase). Treatment with NaCl resulted in improved growth, as evidenced by increased biomass, root length, and lateral root growth, in both sos1 mutant (atsos1) and wild-type (WT) Arabidopsis plants that overexpressed LpSOS1, Under NaCl treatment,atsos1 and WT Arabidopsis plants overexpressing LpSOS1 exhibited better growth, with higher biomass, root length, and lateral root quantity, whereas in the absence of LpSOS1 overexpression, the plants of both lines were wilted and chlorotic and even died under salt stress. When exposed to salt stress, the expression of stress-related genes was notably upregulated in the LpSOS1 overexpression line of Arabidopsis as compared to the WT. Our findings indicate that LpSOS1 enhances salt tolerance in plants by regulating ion homeostasis, reducing Na+/K+ ratio, thereby protecting the plasma membrane from oxidative damage caused by salt stress, and enhancing the activity of antioxidant enzymes. Therefore, the increased salt tolerance conferred by LpSOS1 in plants makes it a potential bioresource for breeding salt-tolerant crops. Further investigation into the mechanisms underlying lily's resistance to salt stress would be advantageous and could serve as a foundation for future molecular improvements.