Plasma Membrane Aquaporins Research Articles

Actin cytoskeleton and plasma membrane aquaporins are involved in different drought response of Arabidopsis rhd2 and der1 root hair mutants

Actin cytoskeleton and reactive oxygen species are principal determinants of root hair polarity and tip growth. Loss of function in RESPIRATORY BURST OXIDASE HOMOLOG C/ROOT HAIR DEFECTIVE 2 (AtRBOHC/RHD2), an NADPH oxidase emitting superoxide to the apoplast, and in ACTIN 2, a vegetative actin isovariant, in rhd2-1 and der1-3 mutants, respectively, lead to similar defects in root hair formation and elongation Since early endosome-mediated polar localization of AtRBOHC/RHD2 depends on actin cytoskeleton, comparing the proteome-wide consequences of both mutations might be of eminent interest. Therefore, we employed a differential proteomic analysis of Arabidopsis rhd2-1 and der1-3 mutants. Both mutants exhibited substantial alterations in abundances of stress-related proteins. Notably, plasma membrane (PM)-localized PIP aquaporins showed contrasting abundance patterns in the mutants compared to wild-types. Drought-responsive proteins were mostly downregulated in rhd2-1 but upregulated in der1-3. Proteomic data suggest that opposite to der1-3, altered vesicular transport in rhd2-1 mutant likely contributes to the deregulation of PM-localized proteins, including PIPs. Moreover, lattice light sheet microscopy revealed reduced actin dynamics in rhd2-1 roots, a finding contrasting with previous reports on der1-3 mutant. Phenotypic experiments demonstrated a drought stress susceptibility in rhd2-1 and resistance in der1-3. Thus, mutations in AtRBOHC/RHD2 and ACTIN2 cause similar root hair defects, but they differently affect the actin cytoskeleton and vesicular transport. Reduced actin dynamics in rhd2-1 mutant is accompanied by alteration of vesicular transport proteins abundance, likely leading to altered protein delivery to PM, including aquaporins, thereby significantly affecting drought stress responses.

Tissue-specific accumulation of PIP aquaporins of a particular heteromeric composition is part of the maize response to mycorrhiza and drought

The systemic coordination of accumulation of plasma membrane aquaporins (PIP) was investigated in this study in relation to mycorrhized maize response to a rapid development of severe drought followed by rewatering. In non-mycorrhizal roots, drought led to a drop in PIP abundance, followed by a transient increase under rewatering, whereas leaves showed an opposite pattern. In contrast, mycorrhiza contributed to maintenance of high and stable levels of PIPs in both plant organs after an initial increase, prolonged over the irrigation period. Isoelectric focusing electrophoresis resolved up to 13 aquaporin complexes with highly reproducible pl positions across leaf and root samples, symbiotic and non-symbiotic, stressed or not. Mass spectrometry recognized in leaves and roots a different ratio of PIP1 and PIP2 subunits within 2D spots that accumulated the most. Regardless of symbiotic status, drought regulation of aquaporins in roots was manifested as the prevalence of complexes that comprise almost exclusively PIP2 monomers. In contrast, the leaf response involved enrichment in PIP1s. PIP1s are thought to enhance water transport, facilitate CO2 diffusion but also affect stomatal movements. These features, together with elevated aquaporin levels, might explain a stress tolerance mechanism observed in mycorrhizal plants, resulting in faster recovery of stomatal water conductance and CO2 assimilation rate after drought.

Plasma membrane aquaporins function in moisture regulation during seed germination and leaf hydration in eelgrass

Eelgrass (Zostera marina) is a marine angiosperm distributed in shallow seas that has a unique mechanism for regulating water content. Water transfer across the plasma membrane is facilitated by aquaporins, which are membrane proteins. In this study, we investigated the physiological functions of plasma membrane intrinsic proteins (PIPs) in eelgrass. ZoPIP1;1 and ZoPIP2;1 in eelgrass leaves were amplified by RT-PCR. Both ZoPIPs localized to the plasma membrane, and only ZoPIP2;1 showed water permeability. ZoPIP1;1 and ZoPIP2;1 were expressed in all tissues except male flowers. ZoPIPs were expressed during seed and fruit development, and their expression levels were low in mature seeds and elevated in germinating seeds. Moisture content decreased in the later phases of seed development and increased during seed germination. These findings imply that ZoPIPs participate in water uptake during seed germination. In leaves, ZoPIP expression was upregulated by dehydration but downregulated by rehydration. Water reabsorption of leaves was inhibited by HgCl2, an inhibitor of aquaporins. These findings indicate that ZoPIPs are upregulated by dehydration during extremely low tides and promote water uptake when the tide turns. Therefore, eelgrass PIPs function in moisture regulation during growth, seed germination, and the flood-ebb tidal cycle in seawater.

Integrated proteomic analysis reveals interactions between phosphorylation and ubiquitination in rose response to Botrytis infection.

As two of the most abundant post-translational modifications, phosphorylation and ubiquitination play a significant role in modulating plant-pathogen interactions and increasing evidence indicates their crosstalk in plant immunity. Rose (Rosa sp.) is one of the most important ornamental plants and can be seriously infected by Botrytis cinerea. Here, integrated proteomics analysis was performed to detect global proteome, phosphorylation, and ubiquitination changes in rose upon B. cinerea infection and investigate the possible phosphorylation and ubiquitination crosstalk. A total of 6165 proteins, 11 774 phosphorylation and 10 582 ubiquitination sites, and 77 phosphorylation and 13 ubiquitination motifs were identified. Botrytis cinerea infection resulted in 169 up-regulated and 122 down-regulated proteins, 291 up-regulated and 404 down-regulated phosphorylation sites, and 250 up-regulated and 634 down-regulated ubiquitination sites. There were 12 up-regulated PR10 proteins and half of them also showed reduced ubiquitination. A lot of kinases probably involved in plant pattern-triggered immunity signaling were up-regulated phosphoproteins. Noticeably, numerous kinases and ubiquitination-related proteins also showed a significant change in ubiquitination and phosphorylation, respectively. A cross-comparison of phosphoproteome and ubiquitylome indicated that both of two post-translational modifications of 104 proteins were dynamically regulated, and many putative pattern-triggered immunity signaling components in the plant plasma membrane were co-regulated. Moreover, five selected proteins, including four PR10 proteins and a plasma membrane aquaporin, were proven to be involved in rose resistance to B. cinerea. Our study provides insights into the molecular mechanisms underlying rose resistance to B. cinerea and also increases the database of phosphorylation and ubiquitination sites in plants.

Root system architecture and genomic plasticity to salinity provide insights into salt-tolerant traits in tall fescue

Salinity is detrimental to soil health, plant growth, and crop productivity. Understanding salt tolerance mechanisms offers the potential to introduce superior crops, especially in coastal regions. Root system architecture (RSA) plasticity is vital for plant salt stress adaptation. Tall fescue is a promising forage grass in saline regions with scarce RSA studies. Here, we used the computer-integrated and -automated programs EZ-Rhizo II and ROOT-Vis II to analyze and identify natural RSA variations and adaptability to high salt stress at physiological and genetic levels in 17 global tall fescue accessions. Total root length rather than the number of lateral roots contribute more to water uptake and could be used to separate salt-tolerant (LS-11) and -sensitive accessions (PI531230). Comparative evaluation of LS-11 and PI531230 demonstrated that the lateral root length rather than the main root contributed more towards the total root length in LS-11. Also, high water uptake was associated with a larger lateral root vector and position while low water intake was associated with an insignificant correlation between root length, vector, and position. To examine candidate gene expression, we performed transcriptome and transcription analyses using high-throughput RNA sequencing and real-time quantitative PCR, respectively of the lateral and main roots. The main root displayed more differentially expressed genes than the lateral root. A Poisson comparison of LS-11 vs PI531230 demonstrated significant upregulation of PLASMA MEMBRANE AQUAPORIN 1 and AUXIN RESPONSE FACTOR 22 in both the main and lateral root, which are associated with transmembrane water transport and the auxin-activated signaling system, respectively. There is also an upregulation of BASIC HELIX-LOOP-HELIX 5 in the main root and a downregulation in the lateral root, which is ascribed to sodium ion transmembrane transport, as well as an upregulation of THE MEDIATOR COMPLEX 1 assigned to water transport in the lateral root and a downregulation in the main root. Gene-protein interaction analysis found that more genes interacting with aquaporins proteins were upregulated in the lateral root than in the main root. We inferred that deeper main roots with longer lateral roots emanating from the bottom of the main root were ideal for tall fescue water uptake and salt tolerance, rather than many shallow roots, and that, while both main lateral roots may play similar roles in salt sensing and water uptake, there are intrinsic genomic differences.

Regulation of drought tolerance in Arabidopsis involves the PLATZ4-mediated transcriptional repression of plasma membrane aquaporin PIP2;8.

Plant A/T-rich protein and zinc-binding protein (PLATZ) transcription factors play important roles in plant growth, development and abiotic stress responses. However, how PLATZ influences plant drought tolerance remains poorly understood. The present study showed that PLATZ4 increased drought tolerance in Arabidopsis thaliana by causing stomatal closure. Transcriptional profiling analysis revealed that PLATZ4 affected the expression of a set of genes involved in water and ion transport, antioxidant metabolism, small peptides and abscisic acid (ABA) signaling. Among these genes, the direct binding of PLATZ4 to the A/T-rich sequences in the plasma membrane intrinsic protein 2;8 (PIP2;8) promoter was identified. PIP2;8 consistently reduced drought tolerance in Arabidopsis through inhibiting stomatal closure. PIP2;8 was localized in the plasma membrane, exhibited water channel activity in Xenopus laevis oocytes and acted epistatically to PLATZ4 in regulating the drought stress response in Arabidopsis. PLATZ4 increased ABA sensitivity through upregulating the expression of ABSCISIC ACID INSENSITIVE 3 (ABI3), ABI4 and ABI5. The transcripts of PLATZ4 were induced to high levels in vegetative seedlings under drought and ABA treatments within 6 and 3 h, respectively. Collectively, these findings reveal that PLATZ4 positively influences plant drought tolerance through regulating the expression of PIP2;8 and genes involved in ABA signaling.

Physiological changes of sweet pepper under low irrigation regimes applied in three phenological stages of vegetative growth, reproductive growth, and fruit set

ABSTRACT A factorial experiment based on a completely randomised design (CRD) with 10 replicates was planned to evaluate if commercially low irrigation reduced output and how physiologically low irrigation influenced yield at different plant growth stages. In three phonological phases of the vegetative stage (VG), reproductive stage (RP), and fruit set (FS), two irrigation regimes of 100% of field capacity (FC) and 80% field capacity (low irrigation = LI) were employed. The results demonstrated that the shoot's fresh weight did not vary between the VG and RP phases, but rose during the FC stage. In FS, both photosynthesis and transpiration were reduced. VG had the highest amounts of photosynthesis and transpiration. Nevertheless, fruit yield rose during the second and third weeks of FS. As starch dropped and glucose, sucrose, and fructose increased, plasma membrane aquaporins (PIP) gene expression and abscisic acid (ABA) levels climbed. In conclusion, it was proved that lowering water consumption to 80% FC did not impact yield at any development stage, including FS, due to the larger sink of fruit rather than maintaining ideal photosynthetic conditions. ABA and PIP expression increased, which may serve as a signal for the plant to overcome the moderate stress generated by LI.

Association studies of genes in a Pb response-associated network in maize (Zea mays L.) reveal that ZmPIP2;5 is involved in Pb tolerance

Lead (Pb) in the soil affects the growth and development of plants and causes damages to the human body through the food chain. Here, we identified and cloned a Pb-tolerance gene ZmPIP2;5 based on a weighted gene co-expression network analysis and gene-based association studies. We showed that ZmPIP2;5 encodes a plasma membrane aquaporin and positively regulated Pb tolerance and accumulation in Arabidopsis and yeast. Overexpression of ZmPIP2;5 increased root length and fresh weight of Arabidopsis seedlings under Pb stress. Heterologous expression of ZmPIP2;5 in yeast caused the enhanced growth speed under Pb treatment and Pb accumulation in yeast cells. A (T/A) SNP in the ZmPIP2;5 promoter affected the expression abundance of ZmPIP2;5 and thereby led to the difference in Pb tolerance among different maize lines. Our study helps to understand the mechanism underlying plant tolerance to Pb stress and provides new ideas for breeding Pb-tolerance maize varieties via molecular marker-assisted selection.

Evolutionary Overview of Aquaporin Superfamily.

Aquaporins (AQPs) are present not only in three domains of life, bacteria, eukaryotes, and archaea, but also in viruses. With the accumulating arrays of AQP superfamily, the evolutional relationship has attracted much attention with multiple publications on "the genome-wide identification and phylogenetic analysis" of AQP superfamily. A pair of NPA boxes forming a pore is highly conserved throughout the evolution and renders key residues for the classification of AQP superfamily into four groups: AQP1-like, AQP3-like, AQP8-like, and AQP11-like. The complexity of AQP family has mostly been achieved in nematodes and subsequent evolution has been directed toward increasing the number of AQPs through whole-genome duplications (WGDs) to extend the tissue specific expression and regulation. The discovery of the intracellular AQP (iAQP: AQP8-like and AQP11-like) and substrate transports by the plasma membrane AQP (pAQP: AQP1-like and AQP3-like) have accelerated the AQP research much more toward the transport of substrates with complex profiles. This evolutionary overview based on a simple classification of AQPs into four subfamilies will provide putative structural, functional, and localization information and insights into the role of AQP as well as clues to understand the complex diversity of AQP superfamily.

Expression patterns of maize PIP aquaporins in middle or upper leaves correlate with their different physiological responses to drought and mycorrhiza.

Here we report the effect of Rhizophagus irregularis on maize leaf expression of six plasma membrane aquaporin isoforms from PIP1 and PIP2 subfamilies under severe drought development and recovery. The novelty of our study is the finding that leaf-specific mycorrhizal regulation of aquaporins is dependent on the position of the leaf on the shoot and changes in parallel with the rate of photosynthesis and the stomatal response to drought. The transcripts were isolated from the upper third (L3) or ear (L5) leaf, which differed greatly in physiological response to stress within each symbiotic variant. Aquaporins expression in upper L3 leaves appeared to be largely not sensitive to drought, regardless of symbiotic status. In contrast, L5 leaf of non-mycorrhizal plants, showed strong down-regulation of all PIPs. Mycorrhiza, however, protected L5 leaf from such limitation, which under maximal stress was manifested by 6-fold and circa 4-fold higher transcripts level for PIP1s and PIP2s, respectively. Distinct expression patterns of L3 and L5 leaves corresponded to differences in key parameters of leaf homeostasis - stomatal conductance, photosynthetic rates, and accumulation of ABA and SA as phytohormonal indicators of drought stress. In result symbiotic plants showed faster restoration of photosynthetic capability, regardless of leaf position, which we recognize as the hallmark of better stress tolerance. In summary, arbuscular mycorrhiza alleviates short-term drought effects on maize by preventing the down-regulation of plasma membrane aquaporins within middle leaves, thereby affecting stomatal conductance.

Plasma membrane aquaporins of the PIP1 and PIP2 subfamilies facilitate hydrogen peroxide diffusion into plant roots

BackgroundThe permeability of plasma membrane aquaporins (PIPs) to small solutes other than water greatly diversifies their potential functions in plant development and metabolic processes. One such process is stress signalling in which hydrogen peroxide (H2O2) plays a major role. Based on transport assays carried out in yeast, there are differences in the degree to which PIPs of Arabidopsis thaliana, are permeable to H2O2 and thus they may differentially facilitate transmembrane diffusion. Here, we test whether specific PIPs aid in the transmembrane diffusion of H2O2 to such an extent that knocking-out PIPs affects plant phenotype. We examined changes in growth and morphology, including biomass accumulation, root system architecture and relative water content, as well as gas exchange, across two H2O2 treatments in knockout mutants of A. thaliana.ResultsWe could infer that PIP-type aquaporins are permeable to H2O2in planta and that this permeability is physiologically relevant in a plant’s response to oxidative stress. In particular, the lack of functional PIP2;3 confers resistance to exogenously applied H2O2 indicating that it facilitates H2O2 entry into root cells. Additionally, PIP1;1 and PIP2;6 were found to facilitate H2O2 diffusion, while PIP2;2 is required for proper root growth under controlled conditions.Main findingsWe conclude that PIPs are physiologically relevant conduits for H2O2 diffusion in the A. thaliana roots and participate in the regulation of stress responses.

Ice plant root plasma membrane aquaporins are regulated by clathrin-coated vesicles in response to salt stress.

The regulation of root Plasma membrane (PM) Intrinsic Protein (PIP)-type aquaporins (AQPs) is potentially important for salinity tolerance. However, the molecular and cellular details underlying this process in halophytes remain unclear. Using free-flow electrophoresis and label-free proteomics, we report that the increased abundance of PIPs at the PM of the halophyte ice plant (Mesembryanthemum crystallinum L.) roots under salinity conditions is regulated by clathrin-coated vesicles (CCV). To understand this regulation, we analyzed several components of the M. crystallinum CCV complexes: clathrin light chain (McCLC) and subunits μ1 and μ2 of the adaptor protein (AP) complex (McAP1μ and McAP2μ). Co-localization analyses revealed the association between McPIP1;4 and McAP2μ and between McPIP2;1 and McAP1μ, observations corroborated by mbSUS assays, suggesting that AQP abundance at the PM is under the control of CCV. The ability of McPIP1;4 and McPIP2;1 to form homo- and hetero-oligomers was tested and confirmed, as well as their activity as water channels. Also, we found increased phosphorylation of McPIP2;1 only at the PM in response to salt stress. Our results indicate root PIPs from halophytes might be regulated through CCV trafficking and phosphorylation, impacting their localization, transport activity, and abundance under salinity conditions.

Hypoxia-Induced Aquaporins and Regulation of Redox Homeostasis by a Trans-Plasma Membrane Electron Transport System in Maize Roots

In plants, flooding-induced oxygen deficiency causes severe stress, leading to growth reduction and yield loss. It is therefore important to understand the molecular mechanisms for adaptation to hypoxia. Aquaporins at the plasma membrane play a crucial role in water uptake. However, their role during hypoxia and membrane redox changes is still not fully understood. The influence of 24 h hypoxia induction on hydroponically grown maize (Zea mays L.) was investigated using an oil-based setup. Analyses of physiological parameters revealed typical flooding symptoms such as increased ethylene and H2O2 levels, an increased alcohol dehydrogenase activity, and an increased redox activity at the plasma membrane along with decreased oxygen of the medium. Transcriptomic analysis and shotgun proteomics of plasma membranes and soluble fractions were performed to determine alterations in maize roots. RNA-sequencing data confirmed the upregulation of genes involved in anaerobic metabolism, biosynthesis of the phytohormone ethylene, and its receptors. Transcripts of several antioxidative systems and other oxidoreductases were regulated. Mass spectrometry analysis of the plasma membrane proteome revealed alterations in redox systems and an increased abundance of aquaporins. Here, we discuss the importance of plasma membrane aquaporins and redox systems in hypoxia stress response, including the regulation of plant growth and redox homeostasis.

A plasma membrane aquaporin is potentially involved in moderating water stress and photosynthetic depression by sustaining water transport and CO2 uptake under high VPD stress conditions in Solanum lycopersicum (tomato)

The molecular mechanism underlying the plant response to high vapor pressure deficit (VPD) stress remains unclear. In the present study, the roles of the plasma membrane aquaporin SlPIP1;2 in moderating plant water stress and photosynthetic depression in response to a series of VPD stress conditions are investigated. Plant water stress and photosynthetic depression are gradually exacerbated as VPD increases. The effects of SlPIP1;2 in mediating plant water status and photosynthesis are gradually enhanced as VPD increases. SlPIP1;2 exerts no significant effect on plant water status and photosynthesis under low VPD conditions. Overexpression of SlPIP1;2 moderates plant water stress by enhancing root water uptake under high VPD stress conditions, sustaining stomatal and mesophyll conductance for photosynthetic CO2 uptake and transport. Consequently, the relatively high CO2 availability inside chloroplasts in SlPIP1;2-overexpressing plants efficiently moderates photosynthetic depression under high VPD stress conditions. The present study demonstrates that SlPIP1;2 is potentially involved in moderating water stress and photosynthetic depression by sustaining water transport and CO2 uptake under high VPD stress conditions.

EuPIP1;2, a Plasma Membrane Aquaporin Gene from Eucommia ulmoides, Enhances Drought and Salt Tolerance in Transgenic Tobacco

Aquaporins are a specific type of membrane channel proteins that efficiently transport water molecules and other small molecular substrates in plants. In this study, we isolated the plasma membrane aquaporin gene EuPIP1;2 from Eucommia ulmoides, an important medicinal plant in China. The EuPIP1;2 protein was localized on the plasma membrane. Quantitative RT-PCR analysis revealed that EuPIP1;2 was constitutively expressed in all analyzed tissues, with the highest expression levels detected in the fruit and root. Overexpression of EuPIP1;2 in transgenic tobacco enhanced plant tolerance of drought and salinity. Under drought and salt stress, the transgenic lines exhibited higher percentage germination, longer roots, and enhanced percentage survival compared with wild-type plants. The contents of malonaldehyde and proline suggested that EuPIP1;2 improved drought and salt tolerance in transgenic lines by reducing damage to membranes and improving osmotic adjustment. We predict that EuPIP1;2 could be applied to improve drought and salt tolerance in transgenic plants.

The plasma membrane aquaporin ZmPIP2;5 enhances the sensitivity of stomatal closure to water deficit.

Increasing stomatal movement is beneficial to improve plant water use efficiency and drought resilience. Contradictory results indicate that aquaporins might regulate stomatal movement. Here, we tested whether the maize plasma membrane PIP2;5 aquaporin affects stomatal closure under water deficit, abscisic acid (ABA)or vapour pressure deficit (VPD) treatment in intact plants, detached leavesor peeled epidermis. Transpiration, stomatal conductance (gs ) and apertureand reactive oxygen species (ROS) in stomatal complexes were studied in maize lines with increased or knocked down (KD) PIP2;5 gene expression. In well-watered conditions, the PIP2;5 overexpressing (OE) plants transpired more than wild types (WTs), while no significant difference in transpiration was observed between pip2;5 KD and WT. Upon mild water deficit or low ABA concentration treatments, transpiration and gs decreased more in PIP2;5 OElines and less in pip2;5 KD lines, in comparison with WTs. In the detached epidermis, ABA treatment induced faster stomatal closing in PIP2;5 OE lines compared to WTs, while pip2;5 KD stomata were ABA insensitive. These phenotypes were associated with guard cell ROS accumulation. Additionally, PIP2;5 is involved in the transpiration decrease observed under high VPD. These data indicate that maize PIP2;5 is a key actor increasing the sensitivity of stomatal closure to water deficit.

The contribution of PIP2-type aquaporins to photosynthetic response to increased vapour pressure deficit.

The roles of different plasma membrane aquaporins (PIPs) in leaf-level gas exchange of Arabidopsis thaliana were examined using knockout mutants. Since multiple Arabidopsis PIPs are implicated in CO2 transport across cell membranes, we focused on identifying the effects of the knockout mutations on photosynthesis, and whether they are mediated through the control of stomatal conductance of water vapour (gs), mesophyll conductance of CO2 (gm), or both. We grew Arabidopsis plants in low and high humidity environments and found that the contribution of PIPs to gs was larger under low air humidity when the evaporative demand was high, whereas any effect of a lack of PIP function was minimal under higher humidity. The pip2;4 knockout mutant had 44% higher gs than wild-type plants under low humidity, which in turn resulted in an increased net photosynthetic rate (Anet). We also observed a 23% increase in whole-plant transpiration (E) for this knockout mutant. The lack of functional plasma membrane aquaporin AtPIP2;5 did not affect gs or E, but resulted in homeostasis of gm despite changes in humidity, indicating a possible role in regulating CO2 membrane permeability. CO2 transport measurements in yeast expressing AtPIP2;5 confirmed that this aquaporin is indeed permeable to CO2.

The potential implications of a plasma membrane aquaporin in improving CO2 transport capacity, photosynthetic potential and water use efficiency under contrasting CO2 source in Solanum lycopersicum (tomato)

Mesophyll conductance has been increasingly recognized as a limiting-factor for carbon transport and fixation during photosynthetic process. Despite mesophyll conductance performs a significant role in determining photosynthetic and yield potential, its molecular mechanisms are uncertain and controversy. In the present study, the function of a plasma membrane aquaporin SlPIP1;2 in facilitating CO2 transport across the plasma membrane under contrasting CO2 concentrations was evaluated, according to the photosynthetic performance of transgenic lines of overexpression and knock-out. The relative importance of PIP-channel in determining membrane permeability and mesophyll conductance is largely depend on the ambient and intercellular CO2 concentration. Under the current ambient CO2 concentration, SlPIP1;2 had minor effect on photosynthetic performance. Overexpression of the target gene substantially enhanced CO2 membrane permeability under CO2 enrichment, according to the photosynthetic performance: overexpression SlPIP1;2 significantly improved mesophyll conductance and photosynthesis rate, which translated into improvement in crop productivity and water use efficiency. The present results suggested that SlPIP1;2 is potentially involved in facilitating CO2 transport across the plasma membrane under CO2 enrichment, which subsequently affect crop productivity and water use efficiency.

Genome-wide identification of plasma membrane aquaporin gene family in Populus and functional identification of PIP1;1 involved in osmotic stress

Aquaporins are the membrane channels that facilitate the transportation of water and small neutral molecules in plants. As such, they play an important role in stress tolerance. This function may be more pronounced in woody plants, which must efficiently absorb water and transport it over long distances. Therefore, a functional analysis of plasma membrane aquaporins (PIP) under drought stress can provide insights into the roles of these proteins in woody plants. In this work, we obtained 15 members of the PtPIP gene family in poplar. They formed two groups in a phylogenetic analysis. Gene structure, including intron phases, was found to be remarkably well conserved among members of this gene family. Five paralogous gene pairs were identified in the PtPIP family, and four of those pairs arose via whole genome duplication. A selection pressure analysis showed that PtPIP genes have undergone positive selection. The transcripts of six PtPIP genes including PtPIP1;1 were predominantly detected in the root. Transgenic poplar overexpressing PtoPIP1;1 was sensitive to osmotic stress, which phenocopied PagMYB216-overexpressing poplar plants. Transactivation and yeast one-hybrid assays showed that PagMYB216, whose homologs in other plants are induced by abiotic stresses, is able to activate PtoPIP1;1 expression through binding to the MBS motif in its promoter. Our results provide new information about PtoPIPs and their roles in response to abiotic stresses, which will be helpful for better understanding their functions in Populus.

Plasma membrane aquaporins interact with the endoplasmic reticulum resident VAP27 proteins at ER-PM contact sites and endocytic structures.

Summary Plasma membrane (PM) intrinsic proteins (PIPs) are aquaporins facilitating the diffusion of water and small solutes. The functional importance of the PM organisation of PIPs in the interaction with other cellular structures is not completely understood.We performed a pull‐down assay using maize (Zea mays) suspension cells expressing YFP‐ZmPIP2;5 and validated the protein interactions by yeast split‐ubiquitin and bimolecular fluorescence complementation assays. We expressed interacting proteins tagged with fluorescent proteins in Nicotiana benthamiana leaves and performed water transport assays in oocytes. Finally, a phylogenetic analysis was conducted.The PM‐located ZmPIP2;5 physically interacts with the endoplasmic reticulum (ER) resident ZmVAP27‐1. This interaction requires the ZmVAP27‐1 cytoplasmic major sperm domain. ZmPIP2;5 and ZmVAP27‐1 localise in close vicinity in ER–PM contact sites (EPCSs) and endocytic structures upon exposure to salt stress conditions. This interaction enhances PM water permeability in oocytes. Similarly, the Arabidopsis ZmVAP27‐1 paralogue, AtVAP27‐1, interacts with the AtPIP2;7 aquaporin.Together, these data indicate that the PIP2–VAP27 interaction in EPCSs is evolutionarily conserved, and suggest that VAP27 might stabilise the aquaporins and guide their endocytosis in response to salt stress.