Extracellular pH sensing by plant cell-surface peptide-receptor complexes.

The extracellular pH is a vital regulator of various biological processes in plants. However, how plants perceive extracellular pH remains obscure. Here, we report that plant cell-surface peptide-receptor complexes can function as extracellular pH sensors. We found that pattern-triggered immunity (PTI) dramatically alkalinizes the acidic extracellular pH in root apical meristem (RAM) region, which is essential for root meristem growth factor 1 (RGF1)-mediated RAM growth. The extracellular alkalinization progressively inhibits the acidic-dependent interaction between RGF1 and its receptors (RGFRs) through the pH sensor sulfotyrosine. Conversely, extracellular alkalinization promotes the alkaline-dependent binding of plant elicitor peptides (Peps) to its receptors (PEPRs) through the pH sensor Glu/Asp, thereby promoting immunity. A domain swap between RGFR and PEPR switches the pH dependency of RAM growth. Thus, our results reveal a mechanism of extracellular pH sensing by plant peptide-receptor complexes and provide insights into the extracellular pH-mediated regulation of growth and immunity in the RAM.

Dual Sensing of Physiologic pH and Calcium by EFCAB9 Regulates Sperm Motility.

Varying pH of luminal fluid along the female reproductive tract is a physiological cue that modulates sperm motility. CatSper is a sperm-specific, pH-sensitive calcium channel essential for hyperactivated motility and male fertility. Multi-subunit CatSper channel complexes organize linear Ca2+ signaling nanodomains along the sperm tail. Here, we identify EF-hand calcium-binding domain-containing protein 9 (EFCAB9) as a bifunctional, cytoplasmic machine modulating the channel activity and the domain organization of CatSper. Knockout mice studies demonstrate that EFCAB9, in complex with the CatSper subunit, CATSPERζ, is essential for pH-dependent and Ca2+-sensitive activation of the CatSper channel. In the absence of EFCAB9, sperm motility and fertility is compromised, and the linear arrangement of the Ca2+ signaling domains is disrupted. EFCAB9 interacts directly with CATSPERζ in a Ca2+-dependent manner and dissociates at elevated pH. These observations suggest that EFCAB9 is a long-sought, intracellular, pH-dependent Ca2+ sensor that triggers changes in sperm motility.

TCR signaling promotes formation of an STS1-Cbl-b complex with pH-sensitive phosphatase activity that suppresses T cell function in acidic environments.

T cell responses are inhibited by acidic environments. T cell receptor (TCR)-induced protein phosphorylation is negatively regulated by dephosphorylation and/or ubiquitination, but the mechanisms underlying sensitivity to acidic environments are not fully understood. Here, we found that TCR stimulation induced a molecular complex of Cbl-b, an E3-ubiquitin ligase, with STS1, a pH-sensitive unconventional phosphatase. The induced interaction depended upon a proline motif in Cbl-b interacting with the STS1 SH3 domain. STS1 dephosphorylated Cbl-b interacting phosphoproteins. The deficiency of STS1 or Cbl-b diminished the sensitivity of T cell responses to the inhibitory effects of acid in an autocrine or paracrine manner in vitro or in vivo. Moreover, the deficiency of STS1 or Cbl-b promoted T cell proliferative and differentiation activities in vivo and inhibited tumor growth, prolonged survival, and improved T cell fitness in tumor models. Thus, a TCR-induced STS1-Cbl-b complex senses intra- or extra-cellular acidity and regulates T cell responses, presenting a potential therapeutic target for improving anti-tumor immunity.

pH sensing at the intersection of tissue homeostasis and inflammation.

pH is tightly maintained at cellular, tissue, and systemic levels, and altered pH - particularly in the acidic range - is associated with infection, injury, solid tumors, and physiological and pathological inflammation. However, how pH is sensed and regulated and how it influences immune responses remain poorly understood at the tissue level. Applying conceptual frameworks of homeostatic and inflammatory circuitries, we categorize cellular and tissue components engaged in pH regulation, drawing parallels from established cases in physiology. By expressing various intracellular (pHi) and extracellular pH (pHe)-sensing receptors, the immune system may integrate information on tissue and cellular states into the regulation of homeostatic and inflammatory programs. We introduce the novel concept of resistance and adaptation responses to rationalize pH-dependent immunomodulation intertwined with homeostatic equilibrium and inflammatory control. We discuss emerging challenges and opportunities in understanding the immunological roles of pH sensing, which might reveal new strategies to combat inflammation and restore tissue homeostasis.

Protein Phosphorylation Orchestrates Acclimations of Arabidopsis Plants to Environmental pH.

Environment pH (pHe) is a key parameter dictating a surfeit of conditions critical to plant survival and fitness. To elucidate the mechanisms that recalibrate cytoplasmic and apoplastic pH homeostasis, we conducted a comprehensive proteomic/phosphoproteomic inventory of plants subjected to transient exposure to acidic or alkaline pH, an approach that covered the majority of protein-coding genes of the reference plant Arabidopsis thaliana. Our survey revealed a large set-of so far undocumented pHe-dependent phospho-sites, indicative of extensive post-translational regulation of proteins involved in the acclimation to pHe. Changes in pHe altered both electrogenic H+ pumping via P-type ATPases and H+/anion co-transport processes, putatively leading to altered net trans-plasma membrane translocation of H+ ions. In pH 7.5 plants, the transport (but not the assimilation) of nitrogen via NRT2-type nitrate and AMT1-type ammonium transporters was induced, conceivably to increase the cytosolic H+ concentration. Exposure to both acidic and alkaline pH resulted in a marked repression of primary root elongation. No such cessation was observed in nrt2.1 mutants. Alkaline pH decreased the number of root hairs in the wild type but not in nrt2.1 plants, supporting a role of NRT2.1 in developmental signaling. Sequestration of iron into the vacuole via alterations in protein abundance of the vacuolar iron transporter VTL5 was inversely regulated in response to high and low pHe, presumptively in anticipation of associated changes in iron availability. A pH-dependent phospho-switch was also observed for the ABC transporter PDR7, suggesting changes in activity and, possibly, substrate specificity. Unexpectedly, the effect of pHe was not restricted to roots and provoked pronounced changes in the shoot proteome. In both roots and shoots, the plant-specific TPLATE complex components AtEH1 and AtEH2-essential for clathrin-mediated endocytosis-were differentially phosphorylated at multiple sites in response to pHe, indicating that the endocytic cargo protein trafficking is orchestrated by pHe.

Role of pH-sensing receptors in colitis.

Low pH in the gut is associated with severe inflammation, fibrosis, and colorectal cancer (CRC) and is a hallmark of active inflammatory bowel disease (IBD). Subsequently, pH-sensing mechanisms are of interest for the understanding of IBD pathophysiology. Tissue hypoxia and acidosis-two contributing factors to disease pathophysiology-are linked to IBD, and understanding their interplay is highly relevant for the development of new therapeutic options. One member of the proton-sensing G protein-coupled receptor (GPCR) family, GPR65 (T-cell death-associated gene 8, TDAG8), was identified as a susceptibility gene for IBD in a large genome-wide association study. In response to acidic extracellular pH, GPR65 induces an anti-inflammatory response, whereas the two other proton-sensing receptors, GPR4 and GPR68 (ovarian cancer G protein-coupled receptor 1, OGR1), mediate pro-inflammatory responses. Here, we review the current knowledge on the role of these proton-sensing receptors in IBD and IBD-associated fibrosis and cancer, as well as colitis-associated cancer (CAC). We also describe emerging small molecule modulators of these receptors as therapeutic opportunities for the treatment of IBD.

Automatic segmentation of lysosomes and analysis of intracellular pH with Radachlorin photosensitizer and FLIM.

We report application of the fluorescence lifetime imaging microscopy (FLIM) for analysis of distributions of intracellular acidity using a chlorin-e6 based photosensitizer Radachlorin. An almost two-fold increase of the photosensitizer fluorescence lifetime in alkaline microenvironments as compared to acidic ones allowed for clear distinguishing between acidic and alkaline intracellular structures. Clusterization of a phasor plot calculated from fits of the FLIM raw data by two Gaussian distributions provided accurate automatic segmentation of lysosomes featuring acidic contents. The approach was validated in colocalization experiments with LysoTracker fluorescence in living cells of four established lines. The dependence of photosensitizer fluorescence lifetime on microenvironment acidity allowed for estimation of pH inside the cells, except for the nuclei, where photosensitizer does not penetrate. The developed method is promising for combined application of the photosensitizer for both photodynamic treatment and diagnostics.

Jasmonates Regulate Auxin-Mediated Root Growth Inhibition in Response to Rhizospheric pH in Arabidopsis thaliana.

Rhizospheric pH, an important environmental cue, severely impacts plant growth and fitness, therefore, has emerged as a major determinant of crop productivity. Despite numerous attempts, the key questions related to plants response against rhizospheric pH remains largely elusive. The present study provides a mechanistic framework for rhizospheric pH-mediated root growth inhibition (RGI). Utilizing various genetic resources combined with pharmacological agents and high-resolution confocal microscopy, the study provides direct evidence for the involvement of jasmonates and auxin in rhizospheric pH-mediated RGI. We show that auxin maxima at root tip is tightly regulated by the rhizospheric pH. In contrast, jasmonates (JAs) abundance inversely correlates with rhizospheric pH. Furthermore, JA-mediated regulation of auxin maxima through GRETCHEN HAGEN 3 (GH3) family genes explains the pattern of RGI observed over the range of rhizospheric pH. Our findings revealed auxin as the key regulator of RGI during severe pH conditions, while JAs antagonistically regulate auxin response against rhizospheric pH.

Benzo-Fused BOPAM Fluorophores: Synthesis, Post-functionalization, Photophysical Properties and Acid sensing Applications.

A novel category of asymmetric boron chromophores with the attachment of two BF2 moieties denoted as BOPAM has been successfully synthesized via a one-pot three-step reaction starting from N-phenylbenzothioamide. This synthetic route results in the production of [a] and [b]benzo-fused BOPAMs along with post-functionalization of the [a]benzo-fused BOPAMs. The photophysical properties of these compounds have been systematically investigated through steady-state absorption and fluorescence emission measurements in solvents at both ambient and cryogenic temperatures, as well as in the solid state. Computational methods have been employed to elucidate the emissive characteristics of the benzo-fused BOPAMs, revealing distinctive photophysical attributes, including solvent-dependent fluorescence intensity. Remarkably, certain BOPAM derivatives exhibit noteworthy photophysical phenomena, such as the induction of off-on fluorescence emission under specific solvent conditions and the manifestation of intermolecular charge transfer states in solid-state matrices. Through post-functionalization strategies involving the introduction of electron-donating groups onto the [a]benzo-fused BOPAM scaffold, an intramolecular charge transfer (ICT) pathway is activated, leading to substantial fluorescence quenching via non-radiative decay processes. Notably, one [a]benzo-fused BOPAM variant exhibits a pronounced fluorescence enhancement upon exposure to acidic conditions, thereby underscoring its potential utility in pH-sensing applications.

Advances in pH Sensing: From Traditional Approaches to Next-Generation Sensors in Biological Contexts.

pH has been considered one of the paramount factors in bodily functions because most cellular tasks exclusively rely on precise pH values. In this context, the current techniques for pH sensing provide us with the futuristic insight to further design therapeutic and diagnostic tools. Thus, pH-sensing (electrochemically and optically) is rapidly evolving toward exciting new applications and expanding researchers' interests in many chemical contexts, especially in biomedical applications. The adaptation of cutting-edge technology is subsequently producing the modest form of these biosensors as wearable devices, which are providing us the opportunity to target the real-time collection of vital parameters, including pH for improved healthcare systems. The motif of this review is to provide insight into trending tech-based systems employed in real-time or in-vivo pH-responsive monitoring. Herein, we briefly go through the pH regulation in the human body to help the beginners and scientific community with quick background knowledge, recent advances in the field, and pH detection in real-time biological applications. In the end, we summarize our review by providing an outlook; challenges that need to be addressed, and prospective integration of various pH in vivo platforms with modern electronics that can open new avenues of cutting-edge techniques for disease diagnostics and prevention.

Green Synthesis of Highly Fluorescent NCQDs: A Comprehensive Study on Synthesis, Characterization, Photophysical Properties, pH Sensing, Heavy Metal Detection, and Solvatochromic Behavior through Hydrothermal Method.

Solvatochromic studies in conjunction with NCQDs and analysis of material at different pH levels provide valuable insights about the process of metal ion sensing. Metal ion sensing holds significant importance in various fields like environment monitoring, biomedical diagnostics and various industrial purpose. The detection of metal ions by mixing the nitrogen-doped quantum dots (NCQDs) in the solvent at different pH levels for the analysis of the photoluminescence spectra is the unique property to achieve selective metal ion detection. In present study, the synthesis of NCQDs was performed by the use of flowers of Tecoma stans. The synthesis of NCQDs to best of our knowledge using flowers of Tecoma stans as natural carbon source via hydrothermal process has been done for the first time. The NCQDs exhibit absorption bands ranging from 190 to 450 nm, with the energy band gap varying from 3.55 to 5.42 eV when mixed with different solvent such as, 1-4 dioxane, acetone, acetonitrile, ethyl- acetate, ethanol, methanol and toluene. The fluorescence spectra exhibited highly intense range from approximately 390 to 680 nm across various solvents. XRD analysis further confirmed the crystalline nature of the particles with an average size of 6.96 nm. Different peak positions of the FTIR spectra support functional groups having C-H stretching, C = O (carbonyl) stretching, and C = C stretching vibrations. In the study a notable solvatochromic shift was observed, indicating sensitivity to change in solvent polarity. Additionally, the investigation of the ratio of ground to excited state dipole moment based on solvatochromic shift yielded a value of 3.30. This provide valuable information about optical and electronic properties of NCQDs. Overall, our study sheds light on the unique properties of NCQDs synthesized from Tecoma stans flowers and their potential applications in metal ion sensing, pH probing, and solvent polarity studies.

Fluorescence Spectra of Prototropic Forms of Fluorescein and Some Derivatives and Their Potential Use for Calibration-Free pH Sensing.

Fluorescence pH sensing has proven to be efficient but with the drawback that molecules photobleach, requiring frequent calibrations. Double-emission peak molecules allow ratiometric measurements and theoretically avoid calibration. However, they are often expensive and fragile and usually have very low quantum yields. Single emission peaks such as fluorescein and derivatives are inexpensive and have very high quantum yields. Because they are single emission peaks, the pH is assumed to be derived from the ratio of emitted intensities at measured pH and at high pH values, i.e., they require frequent calibration. However, the shape of their single emitted peak evolves slightly with pH. In this paper, we first demonstrate a simple method to calculate the emission spectrum shape of each prototropic form of fluorescein (and derivatives) as well as the values of the pKas. A complete model of the evolution of the emission spectrum shape with pH is then constructed. Second, we evaluate the potential of these molecules for pH sensing by fitting the experimental spectra with the complete emission model. The method is applied to fluorescein, FITC and FAM. Depending on the molecule, pH can be measured from pH 1.9 to pH 7.3 with standard deviations between 0.06 and 0.08 pH units. Estimating pH and pKas from shape instead of intensity allows calibration-free measurements even with single-emission peak molecules.

Hell's Gate Globin-I from Methylacidiphilum infernorum Displays a Unique Temperature-Independent pH Sensing Mechanism Utililized a Lipid-Induced Conformational Change.

Hell's Gate globin-I (HGb-I) is a thermally stable globin from the aerobic methanotroph Methylacidiphilium infernorum. Here we report that HGb-I interacts with lipids stoichiometrically to induce structural changes in the heme pocket, changing the heme iron distal ligation coordination from hexacoordinate to pentacoordinate. Such changes in heme geometry have only been previously reported for cytochrome c and cytoglobin, linked to apoptosis regulation and enhanced lipid peroxidation activity, respectively. However, unlike cytoglobin and cytochrome c, the heme iron of HGb-I is altered by lipids in ferrous as well as ferric oxidation states. The apparent affinity for lipids in this thermally stable globin is highly pH-dependent but essentially temperature-independent within the range of 20-60 °C. We propose a mechanism to explain these observations, in which lipid binding and stability of the distal endogenous ligand are juxtaposed as a function of temperature. Additionally, we propose that these coupled equilibria may constitute a mechanism through which this acidophilic thermophile senses the pH of its environment.

A novel anthocyanin indicator film with rosmarinic acid copigmentation having enhanced stability and pH indicator ability for monitoring pork freshness.

BACKGROUND: Anthocyanin-based pH-sensing films have been widely fabricated for potential application in monitoring food freshness. However, the color fading of anthocyanins limits their application for the food industry due to their low stability. In addition, the color sensitivity and pH indicator ability of anthocyanin-based films currently available are not satisfied and need to be improved. RESULTS: Chitosan/xanthan gum (CX)-based colorimetric films with addition of purple cabbage anthocyanin (PAN) and different amounts of rosmarinic acid (RA) were fabricated. RA copigmentation in chitosan/xanthan gum-purple cabbage anthocyanin-rosmarinic acid (CX-P-RA) films significantly improved the stability and pH response sensitivity of PAN, and the combined copigmentation of RA and xanthan gum exhibited an additive effect. The addition of RA significantly improved the tensile strength and elongation at break, thermal stability, antioxidant and antibacterial activities of CX-P-RA films. Moreover, addition of RA enhanced the pH sensitivity and colorimetry of CX-P-RA films, which exhibited a good response to different pH values. CX-P-RA2 film was tested to monitor the freshness of pork. It showed visible color changes during the storage of pork. In addition, the ∆E of CX-P-RA2 film was highly correlated with changes in total volatile basic nitrogen in pork (R2 = 0.951). CONCLUSION: These results indicated that CX-P-RA2 film can be used as a pH-sensing indicator with good stability and high sensitivity for real-time monitoring of pork freshness. © 2023 Society of Chemical Industry.

Allosteric coupling between transmembrane segment 4 and the selectivity filter of TALK1 potassium channels regulates their gating by extracellular pH.

Opening of two-pore domain K+ channels (K2Ps) is regulated by various external cues, such as pH, membrane tension, or temperature, which allosterically modulate the selectivity filter (SF) gate. However, how these cues cause conformational changes in the SF of some K2P channels remains unclear. Herein, we investigate the mechanisms by which extracellular pH affects gating in an alkaline-activated K2P channel, TALK1, using electrophysiology and molecular dynamics (MD) simulations. We show that R233, located at the N-terminal end of transmembrane segment 4, is the primary pHo sensor. This residue distally regulates the orientation of the carbonyl group at the S1 potassium-binding site through an interacting network composed of residues on transmembrane segment 4, the pore helix domain 1, and the SF. Moreover, in the presence of divalent cations, we found the acidic pH-activated R233E mutant recapitulates the network interactions of protonated R233. Intriguingly, our data further suggested stochastic coupling between R233 and the SF gate, which can be described by an allosteric gating model. We propose that this allosteric model could predict the hybrid pH sensitivity in heterodimeric channels with alkaline-activated and acidic-activated K2P subunits.

Mechanisms of Chemosensory Transduction in the Carotid Body.

The mammalian carotid body (CB) is a polymodal chemoreceptor, which is activated by blood-borne stimuli, most notably hypoxia, hypercapnia and acidosis, thus ensuring an appropriate cellular response to changes in physical and chemical parameters of the blood. The glomus cells are considered the CB chemosensory cells and the initial site of chemoreceptor transduction. However, the molecular mechanisms by which they detect changes in blood chemical levels and how these changes lead to transmitter release are not yet well understood. Chemotransduction mechanisms are by far best described for oxygen and acid/carbon dioxide sensing. A few testable hypotheses have been postulated including a direct interaction of oxygen with ion channels in the glomus cells (membrane hypothesis), an indirect interface by a reversible ligand like a heme (metabolic hypothesis), or even a functional interaction between putative oxygen sensors (chemosome hypothesis) or the interaction of lactate with a highly expressed in the CB atypical olfactory receptor, Olfr78, (endocrine model). It is also suggested that sensory transduction in the CB is uniquely dependent on the actions and interactions of gaseous transmitters. Apparently, oxygen sensing does not utilize a single mechanism, and later observations have given strong support to a unified membrane model of chemotransduction.

Advances in the design and use of carbon dots for analytical and biomedical applications.

Carbon dots (CDs) have garnered significant interest for their potential use in multiple applications due to their size, fluorescent properties, high photostability, low toxicity and biocompatibility. CDs can be tailored for specific needs, as they can be synthesized with diverse precursors and techniques for functionalization. Since the applications of CDs are rapidly expanding, this review highlights recent developments in this burgeoning field. Specifically, we describe advances in CD synthesis tailored for applications that include pH and temperature sensing, biochemical analysis, and bioimaging. We also discuss various challenges and practical solutions that will drive CD-based research forward. Challenges include the lack of standardized synthesis and purification methods for CDs, the lack of clarity regarding their mechanism of action, and procedural flaws in their applications. In conclusion, we provide recommendations for collaboration among disciplines to bridge existing knowledge gaps and address the key challenges required for CDs to be fully commercialized.

Nanodiamond surface as a photoluminescent pH sensor.

A systematic spectroscopic characterization of highly homogeneous water suspensions of 'buckydiamonds' comprising sp3cubic nanodiamond (ND) core covered with disordered sp2shell densely decorated with oxygen-containing groups demonstrates the excitation-wavelength-dependent photoluminescence (PL) given by at least four types of specific structures on the ND surface (hydroxyl, C=O containing ketones, carboxylic anhydrides, and carboxyl groups). PL properties of NDs suspensions possess concentration-dependent behavior revealing tendency of NDs to agglomerate. PL of NDs has been found to be strongly sensitive to pH of the environment in wide range of pH values, i.e. 2-11. We disclosed the mechanisms of pH sensitivity of the 'buckydiamond' and proved that it can serve as all-optical sensor of tiny pH variations suitable for further exploitation for pH sensing locally in the area where NDs have been delivered for any purpose, e.g. bioimaging or therapeutic needs.

Facile and Green Synthesis of Carbon Dots from Melia Azedarach Leaves for pH Sensing and Cell Imaging.

Carbon dots (CDs) have preeminent application prospects as a new star in the nanomaterials field. In this work, a green and facile method to synthesize the blue-emitting CDs was proposed with Melia azedarach leaves as the carbon precursors. Using nature materials without other expensive reagents and instruments, the processes were simple and environmental-friendly. The CDs had high fluorescence quantum yield (11.8%) and excellent luminescence properties. The size of them were among 1.5-2.5 nm and the emission spectrum exhibited a strong peak at 460 nm when excited at 380 nm. Additionally, the CDs were stable in most ions but sensitive to different pH values. As a result, a pH sensor was established for the detection of pH with a linear range of 3-10 pH. Moreover, it was demonstrated that the synthesized CDs had extremely low cytotoxicity. Due to their low toxicity and good biocompatibility, they entered into the A549 cells successfully for cell imaging.

Hill-type pH probes.

Sensitive detection of the minute and yet pathologically significant pH variation is important and in fact challenging for the conventional pH probes following the Henderson-Hasselbalch equation, i.e., HH-type probes. A paradigm shift to Hill-type pH probes is ongoing. Bestowed by their positive cooperative acid-base chemistry, their pH-responsive profile follows the Hill equation, which exhibits a narrower acid/base transition width than HH-type probes and warrants a higher detection sensitivity. A polymer-based Hill-type pH-responsive material was first developed. More recently, there emerged several distinct small-molecular approaches to achieve Hill-type pH-responsive profiles. They complement the polymer-based sensing materials in applications where membrane permeability is a concern. In this trends article, we rationalize the molecular origins of their positive cooperativity in pH sensing and highlight some interesting proof-of-concept applications. We also discussed future directions of this dynamic research area.