RESUMEN

In photosystem II (PSII), D1-Tyr246 and D2-Tyr244 are symmetrically located at the binding site of the bicarbonate ligand of the non-heme Fe complex. Here, we investigated the role of the symmetrically arranged tyrosine pair, D1-Tyr246 and D2-Tyr244, in the function of PSII, by generating four chloroplast mutants of PSII from Chlamydomonas reinhardtii: D1-Y246F, D1-Y246T, D2-Y244F, and D2-Y244T. The mutants exhibited altered photoautotrophic growth, reduced PSII protein accumulation, and impaired O2-evolving activity. Flash-induced fluorescence yield decay kinetics indicated a significant slowdown in electron transfer from QA•- to QB in all mutants. Bicarbonate reconstitution resulted in enhanced O2-evolving activity, suggesting destabilization of bicarbonate binding in the mutants. Structural analyses based on a quantum mechanical/molecular mechanical approach identified the existence of a water channel that leads to incorporation of bulk water molecules and destabilization of the bicarbonate binding site. The water intake channels, crucial for bicarbonate stability, exhibited distinct paths in the mutants. These findings shed light on the essential role of the tyrosine pair in maintaining bicarbonate stability and facilitating efficient electron transfer in native PSII.

RESUMEN

Nature has ingeniously developed specialized water transporters that effectively reject ions, including protons, while transporting water across membranes. These natural water channels, known as aquaporins (AQPs), have inspired the creation of Artificial Water Channels (AWCs). However, replicating superfast water transport with synthetic molecular structures that exclude salts and protons is a challenging task. This endeavor demands the coexistence of a suitable water-binding site and a selective filter for precise water transportation. Here, we present small-molecule hydrazides 1b-1d that self-assemble into a rosette-type nanochannel assembly through intermolecular hydrogen bonding and π-π stacking interactions, and selectively transport water molecules across lipid bilayer membranes. The experimental analysis demonstrates notable permeability rates for the 1c derivative, enabling approximately 3.18 × 108 water molecules to traverse the channel per second. This permeability rate is about one order of magnitude lower than that of AQPs. Of particular significance, the 1c ensures exclusive passage of water molecules while effectively blocking salts and protons. MD simulation studies confirmed the stability and water transport properties of the water channel assembly inside the bilayer membranes at ambient conditions.

RESUMEN

Aquaporins (AQPs) are a family of water permeable channels expressed on the plasma membrane with AQP5 being the major channel expressed in several human tissues including salivary and lacrimal glands. Anti-AQP5 autoantibodies have been observed in patients with Sjögren's syndrome who are characterised by dryness of both salivary and lacrimal glands, and they have been implicated in the underlying mechanisms of glandular dysfunction. AQP5 is formed by six transmembrane helices linked with three extracellular and two intracellular loops. Develop antibodies against membrane protein extracellular loops can be a challenge due to the difficulty in maintaining these proteins as recombinant in their native form. Therefore, in this work we aimed to generate an efficient stable-transfected cell line overexpressing human AQP5 (CHO-K1/AQP5) to perform primarily cell-based phage display biopanning experiments to develop new potential recombinant antibodies targeting AQP5. We also showed that the new CHO-K1/AQP5 cell line can be used to study molecular mechanisms of AQP5 sub-cellular trafficking making these cells a useful tool for functional studies.

Asunto(s)

Acuaporina 5 , Cricetulus , Acuaporina 5/metabolismo , Acuaporina 5/genética , Células CHO , Humanos , Animales , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/genética , Anticuerpos/metabolismo , Biblioteca de Péptidos

RESUMEN

Aquaporins (AQPs), also known as water channels, appear to be particularly promising in maintaining male reproductive potential. Therefore, this study aimed to determine the presence of classical AQPs in the bovine (Bos taurus) reproductive system and analyze changes in their expression with age using immunohistochemistry and Western blotting. Of the six classical AQPs, AQP0, AQP1, AQP4, AQP5 and AQP6 were detected, while AQP2 was absent. In the testis, AQP0 was visible in Leydig cells in selected animals, while AQP1 was found in myoid cells surrounding the seminiferous tubules of mature individuals. This characteristic expression patterns of AQP0, limited only to certain bulls, is difficult to explain unequivocally. It is possible that AQP0 expression in cattle is subject to individual variability or changes in response to specific physiological conditions. In the caput and corpus epididymis, AQP0 showed weak expression in epithelial cells of immature animals and stronger expression in basal and principal cells of reproductive bulls. In all animals, AQP1 was present on the apical surface of epithelial cells in the initial segment of the caput epididymis. AQP4, AQP5 and AQP6 were identified in principal and basal cells along the entire epididymis of reproductive bulls. The abundance of AQP4 and AQP6 increased from the caput to the cauda epididymis with the growth and development of the animals. In all males, AQP4, AQP5 and AQP6 were observed in epithelial cells of the vas deferens, and their expression in this section increased with age. In conclusion, the abundance and distribution of the classical AQPs in various cell types and parts of the male reproductive system indicate their crucial role in maintaining water homeostasis, which is essential for normal reproductive function in cattle.

Asunto(s)

Acuaporinas , Animales , Masculino , Bovinos , Acuaporinas/metabolismo , Acuaporinas/genética , Epidídimo/metabolismo , Genitales Masculinos/metabolismo , Testículo/metabolismo , Inmunohistoquímica

RESUMEN

3-Phenyl-2-(thio-phen-3-yl)-2,3-di-hydro-4H-pyrido[3,2-e][1,3]thia-zin-4-one (C17H12N2OS2, 1) and 2-(1H-indol-3-yl)-3-phenyl-2,3-di-hydro-4H-pyrido[3,2-e][1,3]thia-zin-4-one 0.438-hydrate (C21H15N3OS·0.438H2O, 2) crystallize in space groups P21/n and C2/c, respectively. The asymmetric unit in each case is comprised of two parent mol-ecules, albeit of mixed chirality in the case of 1 and of similar chirality in 2 with the enanti-omers occupying the neighboring asymmetric units. Structure 2 also has water mol-ecules (partial occupancies) that form continuous channels along the b -axis direction. The thia-zine rings in both structures exhibit an envelope conformation. Inter-molecular inter-actions in 1 are defined only by C-H⋯O and C-H⋯N hydrogen bonds between crystallographically independent mol-ecules. In 2, hydrogen bonds of the type N-H⋯O between independent mol-ecules and C-H⋯N(π) type, and π-π stacking inter-actions between the pyridine rings of symmetry-related mol-ecules are observed.

RESUMEN

Photosystem II (PSII) is the water-plastoquinone photo-oxidoreductase central to oxygenic photosynthesis. PSII has been extensively studied for its ability to catalyze light-driven water oxidation at a Mn4CaO5 cluster called the oxygen-evolving complex (OEC). Despite these efforts, the complete reaction mechanism for water oxidation by PSII is still heavily debated. Previous mutagenesis studies have investigated the roles of conserved amino acids, but these studies have lacked a direct structural basis that would allow for a more meaningful interpretation. Here, we report a 2.14-Å resolution cryo-EM structure of a PSII complex containing the substitution Asp170Glu on the D1 subunit. This mutation directly perturbs a bridging carboxylate ligand of the OEC, which alters the spectroscopic properties of the OEC without fully abolishing water oxidation. The structure reveals that the mutation shifts the position of the OEC within the active site without markedly distorting the Mn4CaO5 cluster metal-metal geometry, instead shifting the OEC as a rigid body. This shift disturbs the hydrogen-bonding network of structured waters near the OEC, causing disorder in the conserved water channels. This mutation-induced disorder appears consistent with previous FTIR spectroscopic data. We further show using quantum mechanics/molecular mechanics methods that the mutation-induced structural changes can affect the magnetic properties of the OEC by altering the axes of the Jahn-Teller distortion of the Mn(III) ion coordinated to D1-170. These results offer new perspectives on the conserved water channels, the rigid body property of the OEC, and the role of D1-Asp170 in the enzymatic water oxidation mechanism.

Asunto(s)

Dominio Catalítico , Complejo de Proteína del Fotosistema II , Agua , Complejo de Proteína del Fotosistema II/metabolismo , Complejo de Proteína del Fotosistema II/química , Complejo de Proteína del Fotosistema II/genética , Agua/metabolismo , Agua/química , Oxidación-Reducción , Mutación , Microscopía por Crioelectrón , Manganeso/metabolismo , Manganeso/química

RESUMEN

Perivascular spaces (PVSs) as the anatomical basis of the glymphatic system, are increasingly recognized as potential imaging biomarkers of neurological conditions. However, it is not clear whether enlarged PVSs are associated with alcohol-related brain damage (ARBD). We aimed to investigate the effect of long-term alcohol exposure on dyslipidemia and the glymphatic system in ARBD. We found that patients with ARBD exhibited significantly enlargement of PVSs in the frontal cortex and basal ganglia, as well as a notable increased levels of total cholesterol (TC) and triglycerides (TG). The anatomical changes of the glymphatic drainage system mentioned above were positively associated with TC and TG. To further explore whether enlarged PVSs affects the function of the glymphatic system in ARBD, we constructed long alcohol exposure and high fat diet mice models. The mouse model of long alcohol exposure exhibited increased levels of TC and TG, enlarged PVSs, the loss of aquaporin-4 polarity caused by reactive astrocytes and impaired glymphatic drainage function which ultimately caused cognitive deficits, in a similar way as high fat diet leading to impairment in glymphatic drainage. Our study highlights the contribution of dyslipidemia due to long-term alcohol abuse in the impairment of the glymphatic drainage system.

RESUMEN

Aquaporin 2 (AQP2) is a vasopressin (VP)-regulated water channel in the renal collecting duct. Phosphorylation and ubiquitylation of AQP2 play an essential role in controlling the cellular abundance of AQP2 and its accumulation on the plasma membrane in response to VP. Cullin-RING ubiquitin ligases (CRLs) are multisubunit E3 ligases involved in ubiquitylation and degradation of their target proteins, eight of which are expressed in the collecting duct. Here, we used an established cell model of the collecting duct (mpkCCD14 cells) to study the role of cullins in modulating AQP2. Western blotting identified Cul-1 to Cul-5 in mpkCCD14 cells. Treatment of cells for 4 h with a pan-cullin inhibitor (MLN4924) decreased AQP2 abundance, prevented a VP-induced reduction in AQP2 Ser261 phosphorylation, and attenuated VP-induced plasma membrane accumulation of AQP2 relative to the vehicle. AQP2 ubiquitylation levels were significantly higher after MLN4924 treatment compared with controls, and they remained higher despite VP treatment. Cullin inhibition increased ERK1/2 activity, a kinase that regulates AQP2 Ser261 phosphorylation, and VP-induced reductions in ERK1/2 phosphorylation were absent during MLN4924 treatment. Furthermore, the greater Ser261 phosphorylation and reduction in AQP2 abundance during MLN4924 treatment were attenuated during ERK1/2 inhibition. MLN4924 increased intracellular calcium levels via calcium release-activated calcium channels, inhibition of which abolished MLN4924 effects on Ser261 phosphorylation and AQP2 abundance. In conclusion, CRLs play a vital role in mediating some of the effects of VP to increase AQP2 plasma membrane accumulation and AQP2 abundance. Whether modulation of cullin activity can contribute to body water homeostasis requires further studies.NEW & NOTEWORTHY Aquaporin 2 (AQP2) is essential for body water homeostasis and is regulated by the antidiuretic hormone vasopressin. The posttranslational modification ubiquitylation is a key regulator of AQP2 abundance and plasma membrane localization. Here we demonstrate that cullin-RING E3 ligases play a vital role in mediating some of the effects of vasopressin to increase AQP2 abundance and plasma membrane accumulation. The results suggest that manipulating cullin activity could be a novel strategy to alter kidney water handling.

Asunto(s)

Acuaporina 2 , Proteínas Cullin , Ciclopentanos , Túbulos Renales Colectores , Pirimidinas , Ubiquitinación , Acuaporina 2/metabolismo , Proteínas Cullin/metabolismo , Animales , Túbulos Renales Colectores/metabolismo , Túbulos Renales Colectores/efectos de los fármacos , Túbulos Renales Colectores/enzimología , Ubiquitinación/efectos de los fármacos , Fosforilación , Ratones , Vasopresinas/metabolismo , Vasopresinas/farmacología , Línea Celular , Membrana Celular/metabolismo , Membrana Celular/efectos de los fármacos , Ubiquitina-Proteína Ligasas/metabolismo , Calcio/metabolismo

RESUMEN

Aquaporin (AQP) is a water channel protein from the family of transmembrane proteins which facilitates the movement of water across the cell membrane. It is ubiquitous in nature, however the understanding of the water transport mechanism, especially for AQPs in microbes adapted to low temperatures, remains limited. AQP also has been recognized for its ability to be used for water filtration, but knowledge of the biochemical features necessary for its potential applications in industrial processes has been lacking. Therefore, this research was conducted to express, extract, solubilize, purify, and study the functional adaptations of the aquaporin Z family from Pseudomonas sp. AMS3 via molecular approaches. In this study, AqpZ1 AMS3 was successfully subcloned and expressed in E. coli BL21 (DE3) as a recombinant protein. The AqpZ1 AMS3 gene was expressed under optimized conditions and the best optimized condition for the AQP was in 0.5 mM IPTG incubated at 25°C for 20 h induction time. A zwitterionic mild detergent [(3-cholamidopropyl) dimethylammonio]-1-propanesulfonate was the suitable surfactant for the protein solubilization. The protein was then purified via affinity chromatography. Liposome and proteoliposome was reconstituted to determine the particle size using dynamic light scattering. This information obtained from this psychrophilic AQP identified provides new insights into the structural adaptation of this protein at low temperatures and could be useful for low temperature application and molecular engineering purposes in the future.

Asunto(s)

Acuaporinas , Proteínas Bacterianas , Clonación Molecular , Escherichia coli , Pseudomonas , Proteínas Recombinantes , Pseudomonas/metabolismo , Pseudomonas/genética , Pseudomonas/química , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/aislamiento & purificación , Escherichia coli/genética , Escherichia coli/metabolismo , Acuaporinas/química , Acuaporinas/genética , Acuaporinas/metabolismo , Proteínas Bacterianas/química , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Expresión Génica , Proteolípidos/metabolismo , Proteolípidos/química , Regiones Antárticas , Liposomas/metabolismo , Liposomas/química , Agua/química , Agua/metabolismo , Solubilidad , Secuencia de Aminoácidos

RESUMEN

The increasing incidence of male infertility in humans and animals creates the need to search for new factors that significantly affect the course of reproductive processes. Therefore, the aim of this study was to determine the temporospatial expression of aquaglyceroporins (AQP3, AQP7 and AQP9) in the bovine (Bos taurus) reproductive system using immunohistochemistry and Western blotting. The study also included morphological analysis and identification of GATA-4. In brief, in immature individuals, AQP3 and AQP7 were found in gonocytes. In reproductive bulls, AQP3 was observed in spermatocytes and spermatogonia, while AQP7 was visible in all germ cells and the Sertoli cells. AQP7 and AQP9 were detected in the Leydig cells. Along the entire epididymis of reproductive bulls, aquaglyceroporins were visible, among others, in basal cells (AQP3 and AQP7), in epididymal sperm (AQP7) and in the stereocilia of the principal cells (AQP9). In males of all ages, aquaglyceroporins were identified in the principal and basal cells of the vas deferens. An increase in the expression of AQP3 in the testis and cauda epididymis and a decrease in the abundance of AQP7 in the vas deferens with age were found. In conclusion, age-related changes in the expression and/or distribution patterns of AQP3, AQP7 and AQP9 indicate the involvement of these proteins in the normal development and course of male reproductive processes in cattle.

Asunto(s)

Acuagliceroporinas , Acuaporinas , Humanos , Bovinos , Masculino , Animales , Acuaporina 3/genética , Acuaporina 3/metabolismo , Acuaporinas/metabolismo , Semen/metabolismo , Epidídimo/metabolismo , Acuagliceroporinas/metabolismo

RESUMEN

Aquaporins (AQPs) are ubiquitous channel proteins that play a critical role in the homeostasis of the cellular environment by allowing the transit of water, chemicals, and ions. They can be found in many different types of cells and organs, including the lungs, eyes, brain, glands, and blood vessels. By controlling the osmotic water flux in processes like cell growth, energy metabolism, migration, adhesion, and proliferation, AQPs are capable of exerting their regulatory influence over a wide range of cellular processes. Tumour cells of varying sources express AQPs significantly, especially in malignant tumours with a high propensity for metastasis. New insights into the roles of AQPs in cell migration and proliferation reinforce the notion that AQPs are crucial players in tumour biology. AQPs have recently been shown to be a powerful tool in the fight against pathogenic antibodies and metastatic cell migration, despite the fact that the molecular processes of aquaporins in pathology are not entirely established. In this review, we shall discuss the several ways in which AQPs are expressed in the body, the unique roles they play in tumorigenesis, and the novel therapeutic approaches that could be adopted to treat carcinoma.

Asunto(s)

Acuaporinas , Neoplasias , Humanos , Neoplasias/patología , Carcinogénesis , Transformación Celular Neoplásica , Agua/metabolismo , Acuaporinas/química , Acuaporinas/metabolismo

RESUMEN

Graphene oxide (GO) with its atomic thickness and abundant functional groups holds great potential in molecular-scale membrane separation. However, constructing high-speed and highly selective water transport channels within GO membranes remains a key challenge. Herein, sulfonato calix[n]arenes (SCn) molecules with a cavity structure, hydrophilic entrance, and hydrophobic wall were incorporated into GO interlayer channels through a layer-by-layer assembly approach to facilitate water permeation in a water/ethanol separation process. The hydrophilic entrance enables preferential access of water molecules to the cavity over ethanol molecules, while the high hydrophobicity of the cavity wall confers low resistance for water diffusion. After incorporating SCn molecules, the membrane shows a remarkable increase in the water/ethanol separation factor from 732 to 1260, while the permeate flux also increases by about 50%. In addition, the strong electrostatic interactions between the building blocks endow the membrane with excellent swelling resistance even under a high water content. This work provides an effective strategy of constructing high-efficiency water transport channels in membrane.

RESUMEN

Congenital cataracts are the leading cause of irreversible visual disability in children, and genetic factors play an important role in their development. In this study, targeted exome sequencing revealed a novel single-base deletional mutation of MIP (c.301delG; p.Ala101Profs*16) segregated with congenital punctate cataract in a Chinese family. The hydrophobic properties, and secondary and tertiary structures for truncated MIP were predicted to affect the function of protein by bioinformatics analysis. When MIP-WT and MIP-Ala101fs expression constructs were singly transfected into HeLa cells, it was found that the mRNA level showed no significant difference, while the protein level of the mutant was remarkably reduced compared to that of the wild-type MIP. Immunofluorescence images showed that the MIP-WT was principally localized to the plasma membrane, whereas the MIP-Ala101fs protein was aberrantly trapped in the cytoplasm. Furthermore, the cell-to-cell adhesion capability and the cell-to-cell communication property were both significantly reduced for MIP-Ala101fs compared to the MIP-WT (all *p < 0.05). This is the first report of the c.301delG mutation in the MIP gene associated with autosomal dominant congenital cataracts. We propose that the cataract is caused by the decreased protein expression and reduced cell-to-cell adhesion by the mutant MIP. The impaired trafficking or instability of the mutant protein, as well as compromised intercellular communication is probably a concurrent result of the mutation. The results expand the genetic and phenotypic spectra of MIP and help to better understand the molecular basis of congenital cataracts.

Asunto(s)

Catarata , Proteínas del Ojo , Niño , Humanos , Catarata/genética , Catarata/congénito , Adhesión Celular/genética , China , Proteínas del Ojo/genética , Células HeLa , Mutación

RESUMEN

Background Water channel aquaporin 1 (AQP1) protein expression is enhanced in the tunica vaginalis of patients with adult-onset non-communicating hydrocele testis and may contribute to the development of non-communicating hydrocele testis. We performed genetic and epigenetic analyses of the AQP1 gene in the tunica vaginalis of patients with adult-onset non-communicating hydrocele testis to elucidate the cause of enhanced AQP1 protein expression. Methodology The genotype was determined for Tag single-nucleotide polymorphisms (SNPs) representing the AQP1 gene and SNPs in the 5'-upstream region of the AQP1 gene. Then, by performing association analysis, the applicability of various genetic models was investigated for each SNP. Moreover, the methylation rate of CpG sites was examined for the CpG island related to the AQP1 gene. Results There was no significant association between each SNP and hydrocele testis for any of the genetic models. The average methylation rate of the 17 CpG sites evaluated was not significantly different between controls and hydrocele testis, but the methylation rate was lower in hydrocele testis than in controls at one CpG site. Conclusions There was a significant decrease in the methylation rate at one of the CpG sites in the CpG island associated with the AQP1 gene in the tunica vaginalis of patients with non-communicating hydrocele testis. This may increase AQP1 protein expression and contribute to the formation of hydrocele testis. SNPs related to the AQP1 gene were not associated with hydrocele testis.

RESUMEN

The thyroid-stimulating hormone receptor (TSHR) transmembrane domain (TMD) is found in the plasma membrane and consists of lipids and water molecules. To understand the role of TSHR-associated water molecules, we used molecular dynamic simulations of the TMD and identified a network of putative receptor-associated transmembrane water channels. This result was confirmed with extended simulations of the full-length TSHR with and without TSH ligand binding. While the transport time observed in the simulations via the TSHR protein was slower than via the lipid bilayer itself, we found that significantly more water traversed via the TSHR than via the lipid bilayer, which more than doubled with the binding of TSH. Using rat thyroid cells (FRTL-5) and a calcein fluorescence technique, we measured cell volumes after blockade of aquaporins 1 and 4, the major thyroid cell water transporters. TSH showed a dose-dependent ability to influence water transport, and similar effects were observed with stimulating TSHR autoantibodies. Small molecule TSHR agonists, which are allosteric activators of the TMD, also enhanced water transport, illustrating the role of the TMD in this phenomenon. Furthermore, the water channel pathway was also mapped across 2 activation motifs within the TSHR TMD, suggesting how water movement may influence activation of the receptor. In pathophysiological conditions such as hypothyroidism and hyperthyroidism where TSH concentrations are highly variable, this action of TSH may greatly influence water movement in thyroid cells and many other extrathyroidal sites where the TSHR is expressed, thus affecting normal cellular function.

RESUMEN

PURPOSE: Chronic pancreatitis (CP) is associated with serious complications and reduced quality of life. Kidney failure is a frequent complication of acute pancreatitis (AP), however limited information is available regarding the impact of CP on this condition. In the kidney, 9 aquaporins (AQPs) are expressed to maintain body water homeostasis and concentrate urine. The purpose of this study was to morphologically assess and analyze the location and expression of AQP2, AQP3 and AQP4 and determine whether CP affects renal structure and expression of AQPs in collecting duct (CD) principal cells. MATERIALS/METHODS: CP was induced in domestic pigs through intramuscular injections of cerulein (1 â€‹µg/kg â€‹bw/day for 6 days; n â€‹= â€‹5); pigs without CP (n â€‹= â€‹5) were used as a control group. Kidney samples were collected 6 weeks after the last injection and subjected to histological examination. Expression of AQPs was determined by immunohistochemistry and Western blot. RESULTS: The kidneys of animals with CP exhibited moderate changes, including glomerular enlargement, increased collagen percentage, numerous stromal erythrorrhages and inflammatory infiltrations compared to control group. Although the total abundance of AQP2 in the CD decreased in pigs after cerulein administration, the difference was not statistically significant. Expression of AQP3 and AQP4 was limited to the basolateral membrane of the CD cells. AQP4 abundance remained relatively stable in both groups, while AQP3 expression increased nearly three-fold in pigs with CP. CONCLUSION: This study identified morphological alterations and a statistically significant increase in the expression of renal AQP3 when pigs developed CP.

Asunto(s)

Acuaporina 2 , Pancreatitis Crónica , Animales , Porcinos , Acuaporina 2/metabolismo , Ceruletida/metabolismo , Enfermedad Aguda , Calidad de Vida , Acuaporina 3/metabolismo , Riñón/metabolismo

RESUMEN

Cellular systems must deal with mechanical forces to satisfy their physiological functions. In this context, proteins with mechanosensitive properties play a crucial role in sensing and responding to environmental changes. The discovery of aquaporins (AQPs) marked a significant breakthrough in the study of water transport. Their transport capacity and regulation features make them key players in cellular processes. To date, few AQPs have been reported to be mechanosensitive. Like mechanosensitive ion channels, AQPs respond to tension changes in the same range. However, unlike ion channels, the aquaporin's transport rate decreases as tension increases, and the molecular features of the mechanism are unknown. Nevertheless, some clues from mechanosensitive ion channels shed light on the AQP-membrane interaction. The GxxxG motif may play a critical role in the water permeation process associated with structural features in AQPs. Consequently, a possible gating mechanism triggered by membrane tension changes would involve a conformational change in the cytoplasmic extreme of the single file region of the water pathway, where glycine and histidine residues from loop B play a key role. In view of their transport capacity and their involvement in relevant processes related to mechanical forces, mechanosensitive AQPs are a fundamental piece of the puzzle for understanding cellular responses.

RESUMEN

Maintenance of optimal leaf tissue humidity is important for plant productivity and food security. Leaf humidity is influenced by soil and atmospheric water availability, by transpiration and by the coordination of water flux across cell membranes throughout the plant. Flux of water and solutes across plant cell membranes is influenced by the function of aquaporin proteins. Plants have numerous aquaporin proteins required for a multitude of physiological roles in various plant tissues and the membrane flux contribution of each aquaporin can be regulated by changes in protein abundance, gating, localisation, post-translational modifications, protein:protein interactions and aquaporin stoichiometry. Resolving which aquaporins are candidates for influencing leaf humidity and determining how their regulation impacts changes in leaf cell solute flux and leaf cavity humidity is challenging. This challenge involves resolving the dynamics of the cell membrane aquaporin abundance, aquaporin sub-cellular localisation and location-specific post-translational regulation of aquaporins in membranes of leaf cells during plant responses to changes in water availability and determining the influence of cell signalling on aquaporin permeability to a range of relevant solutes, as well as determining aquaporin influence on cell signalling. Here we review recent developments, current challenges and suggest open opportunities for assessing the role of aquaporins in leaf substomatal cavity humidity regulation.

RESUMEN

Water channels, which are small membrane proteins almost entirely buried in lipid membranes, are challenging research targets for single-particle cryo-electron microscopy (cryo-EM), a powerful technique routinely used to determine the structures of membrane proteins. Because the single-particle method enables structural analysis of a whole protein with flexible parts that interfere with crystallization, we have focused our efforts on analyzing water channel structures. Here, utilizing this system, we analyzed the structure of full-length aquaporin-2 (AQP2), a primary regulator of vasopressin-dependent reabsorption of water at the renal collecting ducts. The 2.9 Å resolution map revealed a cytoplasmic extension of the cryo-EM density that was presumed to be the highly flexible C-terminus at which the localization of AQP2 is regulated in the renal collecting duct cells. We also observed a continuous density along the common water pathway inside the channel pore and lipid-like molecules at the membrane interface. Observations of these constructions in the AQP2 structure analyzed without any fiducial markers (e.g., a rigidly bound antibody) indicate that single-particle cryo-EM will be useful for investigating water channels in native states as well as in complexes with chemical compounds.

Asunto(s)

Acuaporina 2 , Proteínas de la Membrana , Acuaporina 2/metabolismo , Microscopía por Crioelectrón/métodos , Proteínas de la Membrana/química , Agua , Lípidos

RESUMEN

The three-dimensional (3D) structure of the cake layer, which could be influenced by water quality factors, plays a significant role in the ultrafiltration (UF) efficiency of water purification. However, it remains challenging to precisely reveal the variation of cake layer 3D structures and water channel characteristics. Herein, we systematically report the variation in the cake layer 3D structure at the nanoscale induced by key water quality factors and reveal its influence on water transport, in particular the abundance of water channels within the cake layer. In comparison with pH and Na+, Ca2+ played more significant role in determining cake layer structures. The sandwich-like cake layer, which was induced by the asynchronous deposition of humic acids and sodium alginate (SA), shifted to an isotropic structure when Ca2+ was present due to the Ca2+ bridging. In comparison with the sandwich-like structure, the isotropic cake layer has higher fractions of free volume (voids) and more water channels, leading to a 147% improvement in the water transport coefficient, 60% reduction in the cake layer resistance, and 21% increase in the final membrane specific flux. Our work elucidates a structure-property relationship where improving the isotropy of the cake layer 3D structure is conducive to the optimization of water channels and water transport within cake layers. This could inspire tailored regulation strategies for cake layers to enhance the UF efficiency of water purification.

Asunto(s)

Ultrafiltración , Purificación del Agua , Ultrafiltración/métodos , Calidad del Agua , Membranas Artificiales , Purificación del Agua/métodos , Sustancias Húmicas/análisis