Plasma Membrane Intrinsic Protein Genes Research Articles

A Plasma Membrane Intrinsic Protein Gene OfPIP2 Involved in Promoting Petal Expansion and Drought Resistance in Osmanthus fragrans

Osmanthus fragrans, a native to China, is renowned as a highly popular gardening plant. However, this plant faces significant challenges from drought stress, which can adversely affect its flowering. In this study, we found that the plasma membrane-localized gene OfPIP2 exhibited a substantial upregulation during the flowering stages and in response to drought stress. GUS staining has illustrated that the OfPIP2 promoter can drive GUS activity under drought conditions. The overexpression of OfPIP2 was found to enhance petal size by modulating epidermal cell dimensions in Petunia and tobacco. Moreover, this overexpression also bolstered drought tolerance, as evidenced by a reduction in stomatal aperture in both species. Furthermore, yeast one-hybrid (Y1H) and dual-luciferase (Dual-LUC) assays have indicated that the transcription factor OfMYB28 directly binds to the OfPIP2 promoter, thereby regulating its expression. Together, we speculated that a module of OfMYB28-OfPIP2 was not only involved in the enhancement of petal size but also conferred the improvement of drought tolerance in O. fragrans. These results contribute valuable insights into the molecular function of the OfPIP2 gene and lay a foundation for molecular breeding strategies in O. fragrans.

Superior osmotic stress tolerance in oilseed rape transformed with wild-type Rhizobium rhizogenes

Key messageNatural transformation with R. rhizogenes enhances osmotic stress tolerance in oilseed rape through increasing osmoregulation capacity, enhancing maintenance of hydraulic integrity and total antioxidant capacity.Transformation of plants using wild strains of agrobacteria is termed natural transformation and is not covered by GMO legislation in, e.g., European Union and Japan. In this study, offspring lines of Rhizobium rhizogenes naturally transformed oilseed rape (Brassica napus), i.e., A11 and B3 (termed root-inducing (Ri) lines), were investigated for osmotic stress resilience. Under polyethylene glycol 6000 (PEG) 10% (w/v)-induced osmotic stress, the Ri lines, particularly A11, had less severe leaf wilting, higher stomatal conductance (8.2 times more than WT), and a stable leaf transpiration rate (about 2.9 mmol m−2 s−1). Although the leaf relative water content and leaf water potential responded similarly to PEG treatment between the Ri lines and WT, a significant reduction of the turgid weight to dry weight ratio in A11 and B3 indicated a greater capacity of osmoregulation in the Ri lines. Moreover, the upregulation of plasma membrane intrinsic proteins genes (PIPs) in roots and downregulation of these genes in leaves of the Ri lines implied a better maintenance of hydraulic integrity in relation to the WT. Furthermore, the Ri lines had greater total antioxidant capacity (TAC) than the WT under PEG stress. Collectively, the enhanced tolerance of the Ri lines to PEG-induced osmotic stress could be attributed to the greater osmoregulation capacity, better maintenance of hydraulic integrity, and greater TAC than the WT. In addition, Ri-genes (particularly rolA and rolD) play roles in response to osmotic stress in Ri oilseed rape. This study reveals the potential of R. rhizogenes transformation for application in plant drought resilience.

Variations in Sweet Pepper Growth, Physiology, and Gene Expression in Shock and Gradual Salt Stress

ABSTRACT Salinity is a major abiotic stressor that can significantly impact plant growth and productivity. To evaluate the physiological responses of sweet pepper (Capsicum annuum L. Ps301) to gradual salinity stress (SLS) and shock salinity stress (STS), an experiment was conducted using a factorially based completely randomized design (CRD). The sodium chloride (NaCl) induced salinity treatments were administered in the nutrient solution in two forms: STS and SLS. The SLS was applied for 8 days, with stress levels increasing every 2 days; STS was administered for 48 h. The signaling response was activated more rapidly and abruptly in response to STS; however, after the various adaptive responses to salinity, these signals were more activated in response to SLS. The STS led to plant growth a greater decrease than SLS. The electrolyte leakage (EL) increased under both SLS and STS by 60% and 61% compared to the control, respectively. Other stress indices, such as phenol and abscisic acid (ABA) content, increased by 26% and 71% under SLS compared to STS. Salinity increased the leaf water potential (LWP) in two salt regimes, and the root water potential (RWP) in STS conditions only. Salinity stress decreased plasma membrane intrinsic protein gene expression in roots, especially in STS compared to SLS. The STS increased plasma membrane intrinsic protein (PIP) expression in leaves more than SLS. It was concluded that salinity impairs the growth and physiological characteristics of pepper; however, the signaling and stress indices respond differently to STS and SLS application.

PIP family-based association studies uncover ZmPIP1;6 involved in Pb accumulation and water absorption in maize roots

Excessive lead (Pb) in the soil affects crop growth and development, thus threatening human beings via food chains. Plasma membrane intrinsic proteins (PIPs) facilitate the transport of substrates across cell membranes. Herein, we characterized maize PIPs and identified eight Pb accumulation-associated PIP genes using association studies. Among these, ZmPIP1;6 was simultaneously correlated with root Pb concentrations under various Pb treatment stages. Significant correlations were observed between the ZmPIP1;6 expression abundance and Pb accumulation in maize roots. Ectopic expression in yeast showed that ZmPIP1;6 conferred Pb accumulation in the cells and affected Pb tolerance in yeast. Overexpression in maize demonstrated that ZmPIP1;6 altered the Pb concentration performance and root moisture content under Pb stress. Meanwhile, protein interaction analyses suggested that ZmPIP1; 6 and three PIP2 members formed isoforms and facilitate water uptake in maize roots. However, ZmPIP1; 6 improved Pb absorption in maize roots probably by interacting with CASP-like protein 2C3 and/or another metal transporter. Moreover, the significant variants in the ZmPIP1;6 promoter caused the variations in ZmPIP1;6 expression level and Pb accumulation among various maize germplasms. Our study will contribute to understanding of PIP family-mediated Pb accumulation in crops and bioremediation of Pb-polluted soils.

Genetic variability of aquaporin expression in maize: From eQTLs to a MITE insertion regulating PIP2;5 expression.

Plant aquaporins are involved in numerous physiological processes, such as cellular homeostasis, tissue hydraulics, transpiration, and nutrient supply, and are key players of the response to environmental cues. While varying expression patterns of aquaporin genes have been described across organs, developmental stages, and stress conditions, the underlying regulation mechanisms remain elusive. Hence, this work aimed to shed light on the expression variability of 4 plasma membrane intrinsic protein (PIP) genes in maize (Zea mays) leaves, and its genetic causes, through expression quantitative trait locus (eQTL) mapping across a 252-hybrid diversity panel. Significant genetic variability in PIP transcript abundance was observed to different extents depending on the isoforms. The genome-wide association study mapped numerous eQTLs, both local and distant, thus emphasizing the existing natural diversity of PIP gene expression across the studied panel and the potential to reveal regulatory actors and mechanisms. One eQTL associated with PIP2;5 expression variation was characterized. Genomic sequence comparison and in vivo reporter assay attributed, at least partly, the local eQTL to a transposon-containing polymorphism in the PIP2;5 promoter. This work paves the way to the molecular understanding of PIP gene regulation and its possible integration into larger networks regulating physiological and stress adaptation processes.

Root pruning improves maize water-use efficiency by root water absorption.

Root systems are an important component of plants that impact crop water-use efficiency (WUE) and yield. This study examined the effects of root pruning on maize yield, WUE, and water uptake under pot and hydroponic conditions. The pot experiment showed that root pruning significantly decreased root/shoot ratio. Both small root pruning (cut off about 1/5 of the root system, RP1) and large root pruning (cut off about 1/3 of the root system, RP2) improved WUE and root hydraulic conductivity (Lpr) in the residual root system. Compared with that in the un-cut control, at the jointing stage, RP1 and RP2 increased Lpr by 43.9% and 31.5% under well-watered conditions and 27.4% and 19.8% under drought stress, respectively. RP1 increased grain yield by 12.9% compared with that in the control under well-watered conditions, whereas both pruning treatments did not exhibit a significant effect on yield under drought stress. The hydroponic experiment demonstrated that root pruning did not reduce leaf water potential but increased residual root hydraulic conductivity by 26.2% at 48 h after root pruning under well-watered conditions. The foregoing responses may be explained by the upregulation of plasma membrane intrinsic protein gene and increases in abscisic acid and jasmonic acid in roots. Increased auxin and salicylic acid contributed to the compensated lateral root growth. In conclusion, root pruning improved WUE in maize by root water uptake.

Herbaceous Peony Polyphenols Extend the Vase Life of Cut Flowers

Herbaceous peony is a potential material for cut flowers, but its short vase life seriously affects the development of cut herbaceous peony flowers industry. In this study, herbaceous peony polyphenols were applied to extend the vase life of cut flowers, and the results indicated that 8% mass concentration of herbaceous peony polyphenols increased the superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) activities; increased the soluble protein content of the cut flowers; and effectively reduced the malondialdehyde (MDA) content. Meanwhile, herbaceous peony polyphenols increased the water balance value of cut flowers. In addition, the observation of microstructures indicated that herbaceous peony polyphenols reduced the blockage mainly caused by Aspergillus spp. at the stem ends and inhibited the growth of Aspergillus spp. Additionally, aquaporin genes (AQPs), including three plasma membrane intrinsic protein genes (PlPIP1;2, PlPIP2;1, and PlPIP2;2) and one intrinsic protein gene (PlNIP), were isolated. PlPIP1;2, PlPIP2;1, and PlPIP2;2, which were induced by polyphenol treatment, had common effects on maintaining the water balance of cut flowers. Therefore, herbaceous peony polyphenols can significantly extend the vase life of cut flowers; these results provide for the application of the theoretical reference of herbaceous peony polyphenols in extending the vase life of cut flowers.

Mycorrhizal Effects on Growth and Expressions of Stress-Responsive Genes (aquaporins and SOSs) of Tomato under Salt Stress.

Environmentally friendly arbuscular mycorrhizal fungi (AMF) in the soil can alleviate host damage from abiotic stresses, but the underlying mechanisms are unclear. The objective of this study was to analyze the effects of an arbuscular mycorrhizal fungus, Paraglomus occultum, on plant growth, nitrogen balance index, and expressions of salt overly sensitive genes (SOSs), plasma membrane intrinsic protein genes (PIPs), and tonoplast intrinsic protein genes (TIPs) in leaves of tomato (Solanum lycopersicum L. var. Huapiqiu) seedlings grown in 0 and 150 mM NaCl stress. NaCl stress severely inhibited plant growth, but P. occultum inoculation significantly improved plant growth. NaCl stress also suppressed the chlorophyll index, accompanied by an increase in the flavonoid index, whereas inoculation with AMF significantly promoted the chlorophyll index as well as reduced the flavonoid index under NaCl conditions, thus leading to an increase in the nitrogen balance index in inoculated plants. NaCl stress regulated the expression of SlPIP1 and SlPIP2 genes in leaves, and five SlPIPs genes were up-regulated after P. occultum colonization under NaCl stress, along with the down-regulation of only SlPIP1;2. Both NaCl stress and P. occultum inoculation induced diverse expression patterns in SlTIPs, coupled with a greater number of up-regulated TIPs in inoculated versus uninoculated plants under NaCl stress. NaCl stress up-regulated SlSOS2 expressions of mycorrhizal and non-mycorrhizal plants, while P. occultum significantly up-regulated SlSOS1 expressions by 1.13- and 0.45-fold under non-NaCl and NaCl conditions, respectively. It was concluded that P. occultum inoculation enhanced the salt tolerance of the tomato, associated with the nutrient status and stress-responsive gene (aquaporins and SOS1) expressions.

Overexpression of CfPIP1-1, CfPIP1-2, and CfPIP1-4 genes of Catalpa fargesii in transgenic Arabidopsis thaliana under drought stress

Catalpa fargesii is an important economic tree species used for furniture and timber production because of its high density and hardness. Its survival and growth are severely affected and primarily limited by drought stress. Thus, to better understand the mechanism of drought resistance in C. fargesii, we used qRT-PCR to reveal significantly different expression of three plasma membrane intrinsic protein genes: CfPIP1-1, CfPIP1-2 and CfPIP1-4. We then cloned their full-length cDNA sequences and characterized the encoded proteins. We analyzed the genes phylogenetically and predicted conserved motifs, domains, and secondary and tertiary structures. To verify the function of the CfPIP1 genes further, we ectopically expressed CfPIP1 transgenes in Arabidopsis thaliana. The results showed that CfPIP1-1, CfPIP1-2 and CfPIP1-4 had several characteristics of aquaporins. The transgenic plants grew better than the WT plants did under drought stress, and overexpression of the CfPIP1 genes increased the plant water content and resistance to drought. Thus, CfPIP1-1, CfPIP1-2 and CfPIP1-4 of C. fargesii play key roles in regulating the intracellular and extracellular water balance and in mediating the plant response to drought.

Effects of partial root-zone irrigation and rational close planting on yield and water productivity of cotton in arid area.

The objective of this study was to evaluate the effects and underlying physiological mecha-nisms of partial root zone irrigation (PRI) and rational close planting, as well as their interaction on yield and water productivity (WP) of cotton and to explore new alternatives of water-saving irrigation in dry land areas. A factorial field experiment with irrigation mode (normal irrigation, partial root-zone irrigation and deficient irrigation) and plant population density (135000, 180000 and 225000 plants·hm-2) was conducted in the west of Inner Mongolia to examine their effects on cotton growth, yield, water productivity and related physiological characters. The results showed that the irrigation mode and plant density as well as their interaction significantly affected the biomass, yield, yield components and harvest index. Under normal irrigation, the biomass and the number of bolls per unit area increased with the increasing of plant density, but the harvest index and boll weight significantly reduced. The yield of high plant density was comparable to that of medium plant density, both of which were increased significantly compared with that of low plant density. The content of abscisic acid (ABA) significantly increased and that of auxin (IAA) significantly reduced in cotton leaves under partial root-zone irrigation, which significantly increased the harvest index by improving the partitioning of assimilates to reproductive organs under partial root-zone irrigation. The number of bolls per unit area increased and boll mass remained unchanged with the increasing of density under partial root-zone irrigation. The yield of high density increased by 6.7% and 11.5% compared with that of medium and low density under partial root zone irrigation. The pre-frost seed cotton increased by 22.5%, the amount of irrigation reduced by 30%, and water productivity increased by 49.3% under partial root zone irrigation compared with that under normal irrigation at high plant density. Plant density did not affect photosynthetic rate (Pn) of functional leaves, but irrigation mode significantly affected Pn. Deficient irrigation significantly reduced the Pn of the main-stem functional leaves, but the Pn under partial root-zone irrigation was comparable to that of normal irrigation. The jasmonate (JA) content and the expression level of plasma membrane intrinsic protein (PIP) gene were significantly increased in the hydrated root under partial root-zone irrigation compared with those under normal irrigation. The results suggested that the increased JA content, as a signal molecule, up-regulated the expression level of PIP gene in dehydrated root and increased water uptake capacity of roots and guaranteed water balance of leaves, and then contributed to a relatively high Pn. Partial root-zone irrigation combined with relatively high plant density (225000 plants·hm-2) is an important agronomic alternative for water saving in cotton plantation in the dry land areas.

Molecular cloning and induced expression of the plasma membrane intrinsic protein gene and promoter from mulberry (Morus multicaulis)

To isolate the plasma membrane intrinsic protein 2 (PIP2) gene and its promoter from mulberry (Morus multicaulis), and to analyse its expression under stress conditions using quantitative real-time polymerase chain reaction (qRT-PCR) analysis, the mulberry MmPIP2 gene was cloned and sequenced. The putative MmPIP2 protein was 282 amino acids long and contained Asn–Pro–Ala signature motifs. Phylogenetic analysis showed that MmPIP2 is highly conserved, exhibiting strong homology with other plant PIP2s. The 5′ flanking sequence was cloned by genome walking and numerous transcription factor binding sites were identified. qRT-PCR revealed that the expression levels of MmPIP2 differed with changes in abiotic stress, indicating that PIP2 is a multifunctional molecule in mulberry. Studying the molecular mechanisms behind adaptations to stress and strengthening stress tolerance in plants are of fundamental importance to mulberry production.

Nano-silver modifies the vase life of cut herbaceous peony (Paeonia lactiflora Pall.) flowers.

Herbaceous peony (Paeonia lactiflora Pall.) is a popular high-grade cut flower because of higher ornamental value. However, its short flowering time severely restricts the production and application of cut P. lactiflora flowers. In this study, nano-silver (NS) was applied to prolong the vase life of cut P. lactiflora flowers. Under the NS treatment, related physiological indices including relative electrical conductivity (REC), malondialdehyde (MDA), superoxide anion free radical (O2·-), hydrogen peroxide (H2O2) and free proline contents, and protective enzyme activities including superoxide dismutase (SOD), peroxidase (POD) and ascorbic acid peroxidase (APX) all increased in cut P. lactiflora flowers except soluble protein. Meanwhile, NS treatment increased relative water uptake (RWU) and Ag+ distribution. Moreover, the observation of microstructures indicated that the stem-ends without NS treatment were blocked by microbes which were identified as Alternaria sp. and Phoma sp., and NS effectively inhibited their growth by antibacterial efficacy observation. Additionally, three aquaporin genes (AQPs) including two plasma membrane intrinsic protein genes (PlPIP1;2, PlPIP2;1) and one NOD26-like intrinsic protein gene (PlNIP) were isolated, PlPIP1;2, and PlPIP2;1 that were induced by NS treatment took common effects on maintaining the water balance of cut P. lactiflora flowers. Consequently, the vase life of cut P. lactiflora flowers was prolonged and flower fresh weight together with flower diameter was well kept because of these above factors. These results would provide a theoretical basis for prolonging the vase life and improving the ornamental quality of cut P. lactiflora flowers with NS application.

Carbon dioxide and water transport through plant aquaporins.

Aquaporins are channel proteins that function to increase the permeability of biological membranes. In plants, aquaporins are encoded by multigene families that have undergone substantial diversification in land plants. The plasma membrane intrinsic proteins (PIPs) subfamily of aquaporins is of particular interest given their potential to improve plant water relations and photosynthesis. Flowering plants have between 7 and 28 PIP genes. Their expression varies with tissue and cell type, through development and in response to a variety of factors, contributing to the dynamic and tissue specific control of permeability. There are a growing number of PIPs shown to act as water channels, but those altering membrane permeability to CO2 are more limited. The structural basis for selective substrate specificities has not yet been resolved, although a few key amino acid positions have been identified. Several regions important for dimerization, gating and trafficking are also known. PIP aquaporins assemble as tetramers and their properties depend on the monomeric composition. PIPs control water flux into and out of veins and stomatal guard cells and also increase membrane permeability to CO2 in mesophyll and stomatal guard cells. The latter increases the effectiveness of Rubisco and can potentially influence transpiration efficiency.

Plasma Membrane Intrinsic Proteins SlPIP2;1, SlPIP2;7 and SlPIP2;5 Conferring Enhanced Drought Stress Tolerance in Tomato.

The function of aquaporin (AQP) protein in transporting water is crucial for plants to survive in drought stress. With 47 homologues in tomato (Solanum lycopersicum) were reported, but the individual and integrated functions of aquaporins involved in drought response remains unclear. Here, three plasma membrane intrinsic protein genes, SlPIP2;1, SlPIP2;7 and SlPIP2;5, were identified as candidate aquaporins genes because of highly expressed in tomato roots. Assay on expression in Xenopus oocytes demonstrated that SlPIP2s protein displayed water channel activity and facilitated water transport into the cells. With real-time PCR and in situ hybridization analysis, SlPIP2s were considered to be involved in response to drought treatment. To test its function, transgenic Arabidopsis and tomato lines overexpressing SlPIP2;1, SlPIP2;7 or SlPIP2;5 were generated. Compared with wild type, the over-expression of SlPIP2;1, SlPIP2;7 or SlPIP2;5 transgenic Arabidopsis and tomato plants all showed significantly higher hydraulic conductivity levels and survival rates under both normal and drought conditions. Taken together, this study concludes that aquaporins (SlPIP2;1, SlPIP2;7 and SlPIP2;5) contribute substantially to root water uptake in tomato plants through improving plant water content and maintaining osmotic balance.

Cloning PIP genes in drought-tolerant vetiver grass and responses of transgenic VzPIP2;1 soybean plants to water stress

Vetiver grass [Vetiveria zizanioides (L.) Nash] displays comprehensive abiotic stress tolerance closely related to fine maintenance of plant water relation mediated by plasma membrane intrinsic proteins (PIPs). Two open reading frame sequences of PIPs (867 and 873 bp) were cloned from vetiver grass and named as VzPIP1;1 and VzPIP2;1, respectively. Expression of green fluorescent protein revealed only subcellular localization of VzPIP2;1 in the plasma membrane. Agrobacterium tumefaciens mediated transgenic (VzPIP2;1) soybean plants had a higher water content in above-ground parts under sufficient water supply through enhancing transpiration as compared to the non-transgenic plants but displayed a more severe drought injury because of a lower photosynthesis and a higher transpiration rate. However, A. rhizogenes mediated transgenic soybean plants kept a higher water content in above-ground parts by improving root water transport and kept a more effective photosynthesis under normal and drought conditions.

Expression patterns of genes encoding plasma membrane aquaporins during fruit development in cucumber (Cucumis sativus L.)

Aquaporins are membrane channels precisely regulating water movement through cell membranes in most living organisms. Despite the advances in the physiology of fruit development, their participation during fruit development in cucumber still barely understood. In this paper, the expressions of 12 genes encoding plasma membrane intrinsic proteins (PIPs) were analyzed during cucumber fruit development in our work. Based on the homology search with known PIPs from rice, Arabidopsis and strawberry, 12 cucumber PIP genes subfamily members were identified. Cellular localization assays indicated that CsPIPs were localized in the plasma membrane. The qRT-PCR analysis of CsPIPs showed that 12 CsPIPs were differentially expressed during fruit development. These results suggest that 12 genes encoding plasma membrane intrinsic proteins (CsPIPs) play very important roles in cucumber life cycle and the data generated will be helpful in understanding their precise roles during fruit development in cucumber.

Ectopic overexpression of a novel Glycine soja stress-induced plasma membrane intrinsic protein increases sensitivity to salt and dehydration in transgenic Arabidopsis thaliana plants.

Plasma membrane intrinsic proteins (PIPs) belong to the aquaporin family and facilitate water movement across plasma membranes. Existing data indicate that PIP genes are associated with the abilities of plants to tolerate certain stress conditions. A review of our Glycine soja expressed sequence tag (EST) dataset revealed that abiotic stress stimulated expression of a PIP, herein designated as GsPIP2;1 (GenBank_Accn: FJ825766). To understand the roles of this PIP in stress tolerance, we generated a coding sequence for GsPIP2;1 by in silico elongation and cloned the cDNA by 5'-RACE. Semiquantitative RT-PCR showed that GsPIP2;1 expression was stimulated in G. soja leaves by cold, salt, or dehydration stress, whereas the same stresses suppressed GsPIP2;1 expression in the roots. Transgenic Arabidopsis thaliana plants overexpressing GsPIP2;1 grew normally under unstressed and cold conditions, but exhibited depressed tolerance to salt and dehydration stresses. Moreover, greater changes in water potential were detected in the transgenic A. thaliana shoots, implying that GsPIP2;1 may negatively impact stress tolerance by regulating water potential. These results, deviating from those obtained in previous reports, provide new insights into the relationship between PIPs and abiotic stress tolerance in plants.

Aquaporins and Leaf Hydraulics: Poplar Sheds New Light

To help understand leaf hydraulic conductance (Kleaf) modulation under high irradiance, well-watered poplars (Populus trichocarpa Torr. & Gray ex Hook and Populus nigra L.) were studied diurnally at molecular and ecophysiological scales. Transcriptional and translational modulations of plasma membrane intrinsic protein (PIP) aquaporins were evaluated in leaf samples during diurnal time courses. Among the 15 poplar PIP genes, a subset of two PIP1s and seven PIP2s are precociously induced within the first hour of the photoperiod concomitantly with a Kleaf increase. Since expression patterns were cyclic and reproducible over several days, we hypothesized that endogenous signals could be involved in PIP transcriptional regulation. To address this question, plants were submitted to forced darkness during their subjective photoperiod and compared with their control counterparts, which showed that some PIP1s and PIP2s have circadian regulation while others did not. Promoter analysis revealed that a large number of hormone, light, stress response and circadian elements are present. Finally, involvement of aquaporins is supported by the reduction of Kleaf by HgCl2 treatment.

Isolation and expression analysis of LoPIP2, a lily (Lilium Oriental Hybrids) aquaporin gene involved in desiccation-induced anther dehiscence

The anthers of many flowers accumulate water before undergoing desiccation and finally dehiscence. Here we report that an internal water deficit and ambient humidity inhibited the dehiscence of lily (Lilium Oriental Hybrids) anthers. This implies that this process also occurs during the dehiscence of lily anthers. Based on speculation that plasma membrane intrinsic proteins (PIPs) may facilitate the dehiscence of lily anthers by promoting the passage of water through lily anther cells, we isolated two PIP genes from lily, LoPIP1 and LoPIP2, and characterized their potential roles in anther dehiscence. Both LoPIP1 and LoPIP2 were constitutively expressed during different phases of anther development; however, only LoPIP2 showed significantly reduced levels of expression in anthers of cut lilies subjected to water deficit for >24h. We silenced LoPIP2 in lily flowers by virus-induced gene silencing and found that this dramatically prevented anther dehiscence. Furthermore, whereas flowers subject to a water deficit had shrunken and indehiscent anthers, flowers in which LoPIP2 expression was silenced had plump and indehiscent anthers that resembled those of flowers exposed to high humidity. Thus the absence of LoPIP2 inhibited anther dehiscence mainly by blocking anther desiccation upon flower opening and not by preventing the earlier stage of water accumulation during anther development and maturation.

Transgenic banana plants overexpressing a native plasma membrane aquaporin MusaPIP1;2 display high tolerance levels to different abiotic stresses

Water transport across cellular membranes is regulated by a family of water channel proteins known as aquaporins (AQPs). As most abiotic stresses like suboptimal temperatures, drought or salinity result in cellular dehydration, it is imperative to study the cause-effect relationship between AQPs and the cellular consequences of abiotic stress stimuli. Although plant cells have a high isoform diversity of AQPs, the individual and integrated roles of individual AQPs in optimal and suboptimal physiological conditions remain unclear. Herein, we have identified a plasma membrane intrinsic protein gene (MusaPIP1;2) from banana and characterized it by overexpression in transgenic banana plants. Cellular localization assay performed using MusaPIP1;2::GFP fusion protein indicated that MusaPIP1;2 translocated to plasma membrane in transformed banana cells. Transgenic banana plants overexpressing MusaPIP1;2constitutively displayed better abiotic stress survival characteristics. The transgenic lines had lower malondialdehyde levels, elevated proline and relative water content and higher photosynthetic efficiency as compared to equivalent controls under different abiotic stress conditions. Greenhouse-maintained hardened transgenic plants showed faster recovery towards normal growth and development after cessation of abiotic stress stimuli, thereby underlining the importance of these plants in actual environmental conditions wherein the stress stimuli is often transient but severe. Further, transgenic plants where the overexpression of MusaPIP1;2 was made conditional by tagging it with a stress-inducible native dehydrin promoter also showed similar stress tolerance characteristics in in vitro and in vivo assays. Plants developed in this study could potentially enable banana cultivation in areas where adverse environmental conditions hitherto preclude commercial banana cultivation.