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1.
Chem Biol Interact ; 311: 108795, 2019 Sep 25.
Artigo em Inglês | MEDLINE | ID: mdl-31419397

RESUMO

Citreoviridin (CIT), a mycotoxin and ATP synthase inhibitor, is regarded as one of aetiology factors of cardiac beriberi and Keshan disease. Thiamine (VB1) and selenium (Se) improve the recovery of these two diseases respectively. The underlying mechanisms of cardiotoxic effect of CIT and cardioprotective effect of VB1 and Se have not been fully elucidated. In this study, we found that ectopic ATP synthase was more sensitive to CIT treatment than mitochondrial ATP synthase in H9c2 cardiomyocytes. CIT inhibited the transcriptional activity of peroxisome proliferator activated receptor gamma (PPAR-γ) in mice hearts and H9c2 cells. PPAR-γ agonist attenuated the inhibitory effect of CIT on mechanistic target of rapamycin complex 2 (mTORC2) and stimulatory effect of CIT on autophagy in cardiomyocytes. CIT induced apoptosis through lysosomal-mitochondrial axis in cardiomyocytes. PPAR-γ agonist and autophagy inhibitor alleviated CIT-induced apoptosis and accelerated cardiac biomarker. VB1 and Se accelerated the basal transcriptional activity of PPAR-γ in mice hearts and H9c2 cells. Furthermore, VB1 and Se reversed the effect of CIT on PPAR-γ, autophagy and apoptosis. Our findings defined PPAR-γ-mTORC2-autophagy pathway as the key link between CIT cardiotoxicity and cardioprotective effect of VB1 and Se. The present study would shed new light on the pathogenesis of cardiomyopathy and the cardioprotective mechanism of micronutrients.


Assuntos
Apoptose/efeitos dos fármacos , Aurovertinas/farmacologia , Autofagia/efeitos dos fármacos , Substâncias Protetoras/farmacologia , Selênio/farmacologia , Tiamina/farmacologia , Animais , Aquaporinas/genética , Aquaporinas/metabolismo , Peso Corporal/efeitos dos fármacos , Linhagem Celular , Masculino , Alvo Mecanístico do Complexo 2 de Rapamicina/genética , Alvo Mecanístico do Complexo 2 de Rapamicina/metabolismo , Camundongos , Camundongos Endogâmicos ICR , Miocárdio/metabolismo , Miocárdio/patologia , PPAR gama/agonistas , PPAR gama/genética , PPAR gama/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Ratos , Proteína X Associada a bcl-2/metabolismo
2.
BMC Plant Biol ; 19(1): 345, 2019 Aug 07.
Artigo em Inglês | MEDLINE | ID: mdl-31390991

RESUMO

BACKGROUND: Aquaporin (AQP) proteins comprise a group of membrane intrinsic proteins (MIPs) that are responsible for transporting water and other small molecules, which is crucial for plant survival under stress conditions including salt stress. Despite the vital role of AQPs, little is known about them in cucumber (Cucumis sativus L.). RESULTS: In this study, we identified 39 aquaporin-encoding genes in cucumber that were separated by phylogenetic analysis into five sub-families (PIP, TIP, NIP, SIP, and XIP). Their substrate specificity was then assessed based on key amino acid residues such as the aromatic/Arginine (ar/R) selectivity filter, Froger's positions, and specificity-determining positions. The putative cis-regulatory motifs available in the promoter region of each AQP gene were analyzed and results revealed that their promoter regions contain many abiotic related cis-regulatory elements. Furthermore, analysis of previously released RNA-seq data revealed tissue- and treatment-specific expression patterns of cucumber AQP genes (CsAQPs). Three aquaporins (CsTIP1;1, CsPIP2;4, and CsPIP1;2) were the most transcript abundance genes, with CsTIP1;1 showing the highest expression levels among all aquaporins. Subcellular localization analysis in Nicotiana benthamiana epidermal cells revealed the diverse and broad array of sub-cellular localizations of CsAQPs. We then performed RNA-seq to identify the expression pattern of CsAQPs under salt stress and found a general decreased expression level of root CsAQPs. Moreover, qRT-PCR revealed rapid changes in the expression levels of CsAQPs in response to diverse abiotic stresses including salt, polyethylene glycol (PEG)-6000, heat, and chilling stresses. Additionally, transient expression of AQPs in N. benthamiana increased leaf water loss rate, suggesting their potential roles in the regulation of plant water status under stress conditions. CONCLUSIONS: Our results indicated that CsAQPs play important roles in response to salt stress. The genome-wide identification and primary function characterization of cucumber aquaporins provides insight to elucidate the complexity of the AQP gene family and their biological functions in cucumber.


Assuntos
Aquaporinas/fisiologia , Cucumis sativus/genética , Proteínas de Plantas/fisiologia , Aquaporinas/genética , Aquaporinas/metabolismo , Cucumis sativus/metabolismo , Expressão Gênica , Regulação da Expressão Gênica de Plantas , Genoma de Planta , Peróxido de Hidrogênio/metabolismo , Filogenia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Estresse Fisiológico/genética , Transcriptoma , Água/metabolismo
3.
Zhonghua Gan Zang Bing Za Zhi ; 27(6): 450-456, 2019 Jun 20.
Artigo em Chinês | MEDLINE | ID: mdl-31357762

RESUMO

Objective: To observe the effect of differentiated mature adipocytes on hepatic steatosis and aquaporin-9 (AQP9) expressions in HepG2 cells and further explore its possible mechanism of action. Methods: Human preadipocytes were cultured and differentiated to full maturity. HepG2 cells were co-cultured with non-differentiated adipocytes and differentiated mature adipocytes for 48 h, and then labeled as control group and experimental group. Oil red O staining and intracellular triglyceride content were performed on co-cultured HepG2 cells and simultaneous changes in phosphatidylinositol 3-kinase (PI3K) - serine/threonine kinase (Akt) signaling pathway, and AQP9 mRNA and protein levels were detected. The experimental group was co-cultured with recombinant human insulin-like growth factor-I (IGF-I), with the addition of 100ng/ml PI3K-Akt pathway agonist, labeled as experimental group + IGF-I group. The activation of PI3K-Akt pathway was verified by Western blotting (WB). The expression of AQP9 was detected by RT-q PCR and WB. The recombinant lentivirus LV-AQP9 or empty-loaded virus LV-PWPI was transfected with HepG2 cells by recombinant lentiviral transfection tecnique, and labeled as HepG2-AQP9 and HepG2-PWPI. The transfection efficiency was assessed by confocal laser scanning microscopy and RT-qPCR and WB detected the change of AQP9 expression level after virus transfection. Afterwards, the stable over-expressed HepG2-AQP9 cells and the empty-loaded HepG2-PWPI cells were co-cultured with differentiated mature adipocytes for 48h, and labeled as HepG2-AQP9 co-culture group, and then intracellular triglyceride content were detected with Oil red O staining. Finally, IGF-I was added to the HepG2-AQP9 co-culture group, which was recorded as HepG2-AQP9 co-culture + IGF-I group. Intracellular triglyceride content was detected with Oil red O staining, and WB verified PI3K-Akt signaling pathway activation and changes in AQP9 mRNA and protein levels. A t-test was used to compare the two independent samples. Results: The intracellular lipid droplets and triglyceride content (0.052 ± 0.005) in the experimental group was increased significantly than the control group (0.033 ± 0.003) (t= 5.225,P= 0.006), suggesting that adipocyte co-culture had induced steatosis in HepG2 cells. RT-qPCR and WB results indicated that the expression levels of AQP9 mRNA (3.615 ± 0.330) and protein levels (0.072 ± 0.005) in the experimental group were significantly higher than the control group (t= 13.708, 11.225,P= 0.005, < 0.001). WB results showed that the expression level of phosphorylated Akt (p-Akt) protein (0.116±0.003) in the experimental group was significantly lower than the control group (0.202 ± 0.003) (t= 27.136,P< 0.001). The total Akt protein was constant, and the p-Akt/total Akt (0.182 ± 0.017)was significantly lower than the control group (0.327 ± 0.019) (t= 2.431,P= 0.001), suggesting that adipocyte co-culture had inhibited PI3K- Akt signaling pathway in HepG2 cells and up-regulated the expression level of AQP9. WB results indicated that the expression level of p-Akt protein (0.194 ± 0.021) in the experimental group + IGF-I group was significantly higher than the experimental group (0.132 ± 0.003) (t= 5.082,P= 0.007). The total Akt protein was constant, and the p-Akt/total Akt (0.281 ± 0.009) was significantly higher than the control group (0.184 ± 0.132) (t= 10.311,P< 0.001). Simultaneously, RT-qPCR and WB results indicated that the expression levels of AQP9 mRNA (0.327 ± 0.347) and protein levels (0.042 ± 0.004) in the experimental group + IGF-I group were significantly lower than the experimental group (t= 33.573, 5.598,P< 0.001, 0.005), suggesting that adipocyte co-culture had possibility to regulate the expression level of AQP9 through the PI3K-Akt pathway. Confocal laser microscopy analysis showed that the transfection efficiency was more than 90%. RT-q PCR and WB results indicated that the expression levels of AQP9 mRNA and protein levels (0.373 ± 0.221) in HepG2-AQP9 group were significantly higher than HepG2-PWPI group (t=14.953, 28.931,P= 0.002 and 0.000), suggesting that the stable overexpression of AQP9 cell line was successfully constructed. The intracellular lipid droplets and triglyceride content in HepG2-AQP9 co-culture group was significantly increased (t= 5.478, 5.369,P= 0.005) than HepG2-PWPI co-culture group and HepG2-AQP9 co-culture+ IGF-I group, suggesting that the increased expression of AQP9 had promoted HepG2 steatosis in co-cultured adipocytes. WB results showed the expression levels of p-Akt protein (0.168 ± 0.006) and p-Akt/total Akt (0.265±0.009) in HepG2-AQP9 co-culture + IGF-1 group was significantly increased (t= 16.311, 8.769,P< 0.001) than HepG2-AQP9 co-culture group, while the expression levels of AQP9 mRNA (0.327 ± 0.034) and protein (0.375 ± 0.025) was significantly decreased (t= 33.573, 9.146,P< 0.001 and 0.001). Conclusion: Adipocytes co-culture can induce steatosis in HepG2 cells, and may participate in inhibiting PI3K-Akt signaling pathway to upregulate the expression of AQP9 in steatotic HepG2 cells.


Assuntos
Adipócitos , Aquaporinas , Regulação da Expressão Gênica no Desenvolvimento , Adipócitos/citologia , Adipócitos/metabolismo , Aquaporinas/genética , Técnicas de Cocultura , Células Hep G2 , Humanos , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo
4.
BMC Genomics ; 20(1): 380, 2019 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-31092186

RESUMO

BACKGROUND: Aquaporins (AQPs) are a class of integral membrane proteins that facilitate the passive transport of water and other small solutes across biological membranes. Despite their importance, little information is available in cassava (Manihot esculenta), a perennial shrub of the Euphorbiaceae family that serves the sixth major staple crop in the world. RESULTS: This study presents a genome-wide analysis of the AQP gene family in cassava. The family of 42 members in this species could be divided into five subfamilies based on phylogenetic analysis, i.e., 14 plasma membrane intrinsic proteins (PIPs), 13 tonoplast intrinsic proteins (TIPs), nine NOD26-like intrinsic proteins (NIPs), four X intrinsic proteins (XIPs), and two small basic intrinsic proteins (SIPs). Best-reciprocal-hit-based sequence comparison and synteny analysis revealed 34 orthologous groups (OGs) present in the Euphorbiaceae ancestor, and nearly one-to-one or two-to-one orthologous relationships were observed between cassava with rubber/physic nut, reflecting the occurrence of one so-called ρ recent whole-genome duplication (WGD) in the last common ancestor of cassava and rubber. In contrast to a predominant role of the ρ WGD on family expansion in rubber, cassava AQP duplicates were derived from the WGD as well as local duplication. Species-specific gene loss was also observed in cassava, which includes the entire NIP4 group and/or six OGs. Comparison of conserved motifs and gene expression profiles revealed divergence of paralogs in cassava as observed in rubber. CONCLUSIONS: Our findings will not only improve our knowledge on family evolution in Euphorbiaceae, but also provide valuable information for further functional analysis of AQP genes in cassava and rubber.


Assuntos
Aquaporinas/genética , Duplicação Gênica , Genoma de Planta , Manihot/genética , Proteínas de Plantas/genética , Borracha/metabolismo , Sequenciamento Completo do Genoma/métodos , Regulação da Expressão Gênica de Plantas , Família Multigênica , Filogenia
5.
Int J Mol Sci ; 20(10)2019 May 14.
Artigo em Inglês | MEDLINE | ID: mdl-31091755

RESUMO

In this study, we generated transgenic durum wheat cv. Maali overexpressing the wheat plasma membrane aquaporin TdPIP2;1 gene under the control of PrTdPIP2;1 promoter or under the constitutive PrCaMV35S promoter. Histochemical analysis of the fusion PrTdPIP2;1::TdPIP2;1::GusA in wheat plants showed that the ß-glucuronidase (GUS) activity was detected in the leaves, stems and roots of stably transformed wheat T3 plants. Our results showed that transgenic wheat lines overexpressing the TdPIP2;1 gene exhibited improved germination rates and biomass production and retained low Na+ and high K+ concentrations in their shoots under high salt and osmotic stress conditions. In a long-term study under greenhouse conditions on salt or drought stress, transgenic TdPIP2;1 lines produced filled grains, whereas wild-type (WT) plants either died at the vegetative stage under salt stress or showed drastically reduced grain filling under drought stress. Performing real time RT-PCR experiments on wheat plants transformed with the fusion PrTdPIP2;1::GusA, we showed an increase in the accumulation of GusA transcripts in the roots of plants challenged with salt and drought stress. Study of the antioxidant defence system in transgenic wheat TdPIP2;1 lines showed that these lines induced the antioxidative enzymes Catalase (CAT) and Superoxide dismutase (SOD) activities more efficiently than the WT plants, which is associated with lower malondialdehyde and hydrogen peroxide contents. Taken together, these results indicate the high potential of the TdPIP2;1 gene for reducing water evaporation from leaves (water loss) in response to water deficit through the lowering of transpiration per unit leaf area (stomatal conductance) and engineering effective drought and salt tolerance in transgenic TdPIP2;1 lines.


Assuntos
Aquaporinas/genética , Secas , Proteínas de Plantas/genética , Tolerância ao Sal , Triticum/genética , Aquaporinas/metabolismo , Catalase/genética , Catalase/metabolismo , Germinação , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/metabolismo , Superóxido Dismutase/genética , Superóxido Dismutase/metabolismo , Triticum/metabolismo , Triticum/fisiologia , Regulação para Cima
6.
Life Sci ; 229: 104-115, 2019 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-31100324

RESUMO

AIM: Blockage of the urinary tract is often connected with renal function impediment, including reductions in glomerular filtration rate (GFR) and the power to control sodium as well as water elimination through urination. Melatonin, known to be the primary product of the pineal gland, prevents renal damage caused by ischemic reperfusion. However, the effects of melatonin on urinary obstruction, as well as release of obstruction induced kidney injury are still largely unknown. The aim of present study was to investigate the effect of melatonin on mediating protection against renal injury triggered from either bilateral ureteral obstruction (BUO) or BUO release (BUO-R). MAIN METHODS: Adult male Sprague-Dawley rats (n = 60) were clustered into six treatment groups: sham treated-1; BUO-non-treated (24 h BUO only); BUO + melatonin; sham treated-2; BUO-48hR (24 h of BUO and then release for 2 days); and BUO-48hR + melatonin. Kidney tissues, blood and urine samples were obtained for further assessment. KEY FINDINGS: It was found that melatonin treatment remarkably promoted the recovery of the handling capacity of urinary excretion of water as well as sodium in BUO and BUO-48hR models. Melatonin treatment partially inhibited inflammatory cytokine expression and the downregulation of aquaporin (AQPs, AQP-1, -2 and -3) expression in these two models. Moreover, the cytoarchitecture of BUO rats exposed to melatonin was well preserved. SIGNIFICANCE: Melatonin treatment potently prevents BUO or BUO-R induced renal injury, which may be partially attributed to restoring the expression of AQPs and inhibition of inflammatory response, as well as preserving renal ultrastructural integrity.


Assuntos
Antioxidantes/farmacologia , Nefropatias/prevenção & controle , Melatonina/farmacologia , Substâncias Protetoras/farmacologia , Obstrução Ureteral/complicações , Animais , Aquaporinas/genética , Aquaporinas/metabolismo , Taxa de Filtração Glomerular , Nefropatias/etiologia , Nefropatias/patologia , Masculino , Ratos , Ratos Sprague-Dawley , Obstrução Ureteral/fisiopatologia
7.
Artif Cells Nanomed Biotechnol ; 47(1): 1782-1787, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31062612

RESUMO

Hepatic steatosis is one of the most important features of the pathogenesis for non-alcoholic fatty liver disease. Fat deposition in liver cells can influence hepatic lipogenesis along with other metabolic pathways and further lead to the irreversible liver cirrhosis and injury. However, the underlying mechanism of steatosis remains largely unexplored. Our previous study revealed that AQP7 played an important role in liver steatosis. In this study, we determined that the transcriptional level of AQP7 was up-regulated by estrogen receptor alpha (ERα) upon 17ß-estradiol (E2) and oleic acids treated HepG2 cells. Furthermore, we identified long non-coding RNA nuclear enriched abundant transcript 1 (NEAT1) as a potential hallmark which was down-regulated in ERα silencing HepG2 cells by RNA-Seq. Finally, we validated that the 3' terminal nucleotides of NEAT1 were contributed for the interaction with ERα to facilitate AQP7 transcription to suppress liver steatosis. Overall, our study gave evidence that NEAT1 played an important role in the activation of ERα to regulate AQP7-mediated hepatic steatosis.


Assuntos
Aquaporinas/genética , Receptor alfa de Estrogênio/metabolismo , Fígado Gorduroso/genética , Regulação da Expressão Gênica/genética , RNA Longo não Codificante/genética , Fígado Gorduroso/metabolismo , Fígado Gorduroso/patologia , Células Hep G2 , Humanos , RNA Longo não Codificante/metabolismo , Transcrição Genética/genética
8.
Virol J ; 16(1): 58, 2019 05 02.
Artigo em Inglês | MEDLINE | ID: mdl-31046802

RESUMO

BACKGROUND: MicroRNAs (miRNAs) have gained much attention as cellular factors regulating hepatitis C virus (HCV) infection. miR-27b has been shown to regulate HCV infection in the hepatocytes via various mechanisms that have not been fully elucidated. In this study, therefore, we examined the mechanisms of miR-27b-mediated regulation of HCV infection. METHODS: In silico screening analysis, transfection with miR-27b mimic, and a cell-based reporter assay were performed to identify miR-27b target genes. Cell cultured-derived HCV (HCVcc) was added to Huh7.5.1 cells knocked down for aquaporin (AQP) 11 (AQP11) and overexpressing AQP11. HCV replication levels were evaluated by real-time RT-PCR analysis of HCVcc genome. RESULTS: Infection of Huh7.5.1 cells with HCVcc resulted in significant elevation in miR-27b expression levels. In silico analysis revealed that AQP11, which is an AQP family member and is mainly localized in the endoplasmic reticulum (ER), was a candidate for a target gene of miR-27b. Transfection of a miR-27b mimic significantly reduced AQP11 expression, but a cell-based reporter assay demonstrated that miR-27b did not suppress the expression of a reporter gene containing the 3'-untranslated region of the AQP11 gene, suggesting that miR-27b indirectly suppressed AQP11 expression. AQP11 expression levels were significantly reduced by infection with HCVcc in Huh7.5.1 cells. Knockdown and over-expression of AQP11 significantly reduced and increased HCVcc genome levels in the cells following infection, respectively, however, AQP11 knockdown did not show significant effects on the HCVcc titers in the culture supernatants. CONCLUSIONS: These results indicated that HCV infection induced a miR-27b-mediated reduction in AQP11 expression, leading to a modest reduction in HCV genome levels in the cells, not HCV titers in the culture supernatants.


Assuntos
Aquaporinas/genética , Hepacivirus/genética , Hepatócitos/virologia , MicroRNAs/genética , RNA Viral/análise , Linhagem Celular , Regulação da Expressão Gênica , Técnicas de Silenciamento de Genes , Genoma Viral , Humanos , RNA Viral/genética , Transfecção , Carga Viral
9.
Gene Expr Patterns ; 32: 38-43, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30951885

RESUMO

The aquaporins are integral membrane proteins from a larger family of major intrinsic protein (MIP) that form pores in the membrane of cells. These proteins selectively transport water and other small uncharged solutes across cell plasma membranes. The organization of water within cells and tissues is fundamental to life, and the aquaporins play an important role in serving as the plumbing system for cells. As many as thirteen mammalian AQPs have been characterized, which have been shown to be vital for the regulation of water homeostasis in most tissues, such as renal water balance and brain-fluid homeostasis. However, complete expression patterns of most of the aquaporins in lower vertebrate at embryo stages has not been elucidated. Currently, we systematically described the temporal-spatial expression pattern of nine zebrafish aquaporins, using whole amount in situ hybridization. The results of whole mount in situ hybridization revealed that members of aquaporins family displayed diverse expression pattern, each of aquaporins has its unique distribution in different cell types and tissues, suggesting that they might play distinct roles in the embryonic development. Overall, current study will provide new insight into the expression of vertebrate quaporins and an important basis for the functional analysis of aquaporins in zebrafish development.


Assuntos
Aquaporinas/genética , Aquaporinas/metabolismo , Sequência de Aminoácidos , Animais , Desenvolvimento Embrionário/genética , Homeostase , Transcriptoma/genética , Peixe-Zebra/embriologia , Proteínas de Peixe-Zebra/metabolismo
10.
Plant Mol Biol ; 100(3): 335-349, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30963359

RESUMO

KEY MESSAGE: The ER membrane localized aquaporin SIP2;1 is involved in adaptation to ER stresses during pollen tube elongation. Aquaporins play multifaceted roles through selective transport of water and small neutral substrates. Here, we focused on the physiological roles of Arabidopsis thaliana aquaporins, namely SIP1;1, SIP1;2 and SIP2;1, which are localized to the endoplasmic reticulum (ER). While their loss-of-function mutants displayed normal vegetative growth. We identified defects in pollen of sip2;1. Whereas the germination rate of sip2;1 pollen was ~ 60% that of the wild type (WT), in vitro germinated sip2;1 pollen tube length was reduced up to 82% compared to the WT. Importantly, most pollen tubes on pistils from sip2;1 stopped elongation in the mid-region of pistils, and the bottom region of sip2;1 siliques lacked seeds. Consistently, silique of sip2;1 were short, whereby the average seed number per silique was nearly the half of the WT. The above phenotypes recovered in SIP2;1 complementation lines. We detected mRNA of SIP2;1 and protein in pollen, and further revealed that the GFP-linked SIP2;1 localization in the ER of growing pollen tubes. The basal mRNA level of BINDING PROTEIN 3 (BiP3), a key gene induced by ER stress, in pollen was markedly higher than that in roots, suggesting that the pollen underwent ER stress under normal growth conditions. BiP3 mRNA was dramatically increased in sip2;1 pollen. Altogether, our findings suggest that the aquaporin SIP2;1 is probably involved in the alleviation of ER stress and that the lack of SIP2;1 reduces both pollen germination and pollen tube elongation.


Assuntos
Aquaporinas/metabolismo , Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Retículo Endoplasmático/metabolismo , Germinação , Tubo Polínico/metabolismo , Pólen/metabolismo , Aquaporinas/genética , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Sobrevivência Celular , Fertilidade , Flores/metabolismo , Regulação da Expressão Gênica de Plantas , Chaperonas Moleculares , Mutação , Fenótipo , Proteínas de Plantas/metabolismo , Raízes de Plantas , Pólen/crescimento & desenvolvimento , Tubo Polínico/crescimento & desenvolvimento , RNA Mensageiro
11.
Planta ; 250(1): 319-332, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31030328

RESUMO

MAIN CONCLUSION: Depending on the N source and plant ontogenetic state, the epiphytic tank-forming bromeliad Vriesea gigantea can modulate aquaporin expression to maximize the absorption of the most available nitrogen source. Epiphytic bromeliads frequently present a structure formed by the overlapping of leaf bases where water and nutrients can be accumulated and absorbed, called tank. However, this structure is not present during the juvenile ontogenetic phase, leading to differences in nutrient acquisition strategies. Recent studies have shown a high capacity of the bromeliad Vriesea gigantea, an epiphytic tank-forming bromeliad, to absorb urea by their leaves. Since plant aquaporins can facilitate the diffusion of urea through the membranes, we cloned three foliar aquaporin genes, VgPIP1;1, VgPIP1;2 and VgTIP2;1 from V. gigantea plants. Through functional studies, we observed that besides water, VgTIP2;1 was capable of transporting urea while VgPIP1;2 may facilitate ammonium/ammonia diffusion. Moreover, aiming at identifying urea and ammonium-induced changes in aquaporin expression in leaves of juvenile and adult-tank plants, we showed that VgPIP1;1 and VgPIP1;2 transcripts were up-regulated in response to either urea or ammonium only in juvenile plants, while VgTIP2;1 was up-regulated in response to urea only in adult-tank plants. Thereby, an ontogenetic shift from juvenile to adult-tank-forming-plant appears to occur with metabolic changes regarding nitrogen metabolism regulation. Investigating urea metabolism in wild species that naturally cope with organic N sources, such as V. gigantea, may provide the knowledge to modify nitrogen use efficiency of crop plants.


Assuntos
Aquaporinas/metabolismo , Bromeliaceae/metabolismo , Nitrogênio/metabolismo , Ureia/metabolismo , Aquaporinas/genética , Bromeliaceae/genética , Folhas de Planta/genética , Folhas de Planta/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Água/metabolismo
12.
BMC Genomics ; 20(1): 222, 2019 Mar 18.
Artigo em Inglês | MEDLINE | ID: mdl-30885116

RESUMO

BACKGROUND: Aquaporins (AQPs) facilitate transport of water and small solutes across cell membranes and play an important role in different physiological processes in plants. Despite their importance, limited data is available about AQP distribution and function in the economically important oilseed crop peanut, Arachis hypogea (AABB). The present study reports the identification and structural and expression analysis of the AQPs found in the diploid progenitor genomes of A. hypogea i.e. Arachis duranensis (AA) and Arachis ipaensis (BB). RESULTS: Genome-wide analysis revealed the presence of 32 and 36 AQPs in A. duranensis and A. ipaensis, respectively. Phylogenetic analysis showed similar numbers of AQPs clustered in five distinct subfamilies including the plasma membrane intrinsic proteins (PIPs), the tonoplast intrinsic proteins (TIPs), the nodulin 26-like intrinsic proteins (NIPs), the small basic intrinsic proteins (SIPs), and the uncharacterized intrinsic proteins (XIPs). A notable exception was the XIP subfamily where XIP1 group was observed only in A. ipaensis genome. Protein structure evaluation showed a hydrophilic aromatic/arginine (ar/R) selectivity filter (SF) in PIPs whereas other subfamilies mostly contained a hydrophobic ar/R SF. Both genomes contained one NIP2 with a GSGR SF indicating a conserved ability within the genus to uptake silicon. Analysis of RNA-seq data from A. hypogea revealed a similar expression pattern for the different AQP paralogs of AA and BB genomes. The TIP3s showed seed-specific expression while the NIP1s' expression was confined to roots and root nodules. CONCLUSIONS: The identification and the phylogenetic analysis of AQPs in both Arachis species revealed the presence of all five sub-families of AQPs. Within the NIP subfamily, the presence of a NIP2 in both genomes supports a conserved ability to absorb Si within plants of the genus. The global expression profile of AQPs in A. hypogea revealed a similar pattern of AQP expression regardless of the subfamilies or the genomes. The tissue-specific expression of AQPs suggests an important role in the development and function of the respective organs. The AQPs identified in the present study will serve as a resource for further characterization and possible exploitation of AQPs to understand their physiological role in A. hypogea.


Assuntos
Aquaporinas/genética , Arachis/classificação , Arachis/genética , Regulação da Expressão Gênica de Plantas , Genoma de Planta , Proteínas de Plantas/genética , Aquaporinas/metabolismo , Arachis/metabolismo , Diploide , Evolução Molecular , Perfilação da Expressão Gênica , Família Multigênica , Filogenia , Proteínas de Plantas/metabolismo
13.
Anim Reprod Sci ; 204: 22-30, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30862405

RESUMO

Fluid regulation within the male gonad is an important process for promoting sperm differentiation and maturation. Aquaporins (AQPs) are a family of thirteen integral membrane proteins involved in these processes. The expression of several genes of AQPs occurs in the male reproductive tract of humans and other animal species, although there are few studies on domestic animals. In this study, the localization of AQP7, AQP8, and AQP9 as well as the abundances of protein and mRNA transcripts were examined in normal and cryptorchid dog testes. There was immunohistochemical localization of AQP7, AQP8, and AQP9 in both the tubular and interstitial compartments of the normal and retained testes and crytorchid dogs, albeit there was an obvious difference in cellular localization with the testes from the cryptorchid dogs. These results were supported by western blotting and real-time RT-PCR analyses, there was a lesser AQP7 and greater AQP9 abundance of protein and mRNA transcripts in the cryptorchid testis. These findings indicate combined testicular functions of AQPs in cell volume regulation. In addition, with the cryptorchid condition characterized there was a different cellular distribution of AQPs supporting the thought that early detection is important for controlling possible side effects of cyptorchidism, such as pre-neoplastic and carcinogenic outcomes.


Assuntos
Aquaporinas/metabolismo , Criptorquidismo/veterinária , Doenças do Cão/patologia , Testículo/citologia , Animais , Aquaporinas/genética , Criptorquidismo/patologia , Cães , Regulação da Expressão Gênica/fisiologia , Imuno-Histoquímica , Masculino , RNA , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Testículo/patologia
14.
Invest Ophthalmol Vis Sci ; 60(4): 858-867, 2019 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-30821811

RESUMO

Purpose: Investigate the effects of the absence of 17 amino acids at the C-terminal end of Aquaporin 0 (AQP0) on lens transparency, focusing property, and homeostasis. Methods: A knockin (KI) mouse model (AQP0ΔC/ΔC) was developed to express AQP0 only as the end-cleaved form in the lens. For this, AQP0 was genetically engineered as C-terminally end-cleaved with amino acids 1 to 246, instead of the full length 1 to 263 of the wild type (WT). After verifying the KI integration into the genome and its expression, the mouse model was bred for several generations. AQP0 KI homozygous (AQP0ΔC/ΔC) and heterozygous (AQP0+/ΔC) lenses were imaged and analyzed at different developmental stages for transparency. Correspondingly, aberrations in the lens were characterized using the standard metal grid focusing method. Data were compared with age-matched WT, AQP0 knockout (AQP0-/-), and AQP0 heterozygous (AQP0+/-) lenses. Results: AQP0ΔC/ΔC lenses were transparent throughout the embryonic development and until postnatal day 15 (P15) in contrast to age-matched AQP0-/- lenses, which developed cataract at embryonic stage itself. However, there was distortion aberration in AQP0ΔC/ΔC lens at P5; after P15, cataract began to develop and progressed faster surpassing that of age-matched AQP0-/- lenses. AQP0+/ΔC lenses were transparent even at the age of 1 year in contrast to AQP0+/- lenses; however, there was distortion aberration starting at P15. Conclusions: A specific distribution profile of intact and end-cleaved AQP0 from the outer cortex to the inner nucleus is required in the lens for establishing refractive index gradient to enable proper focusing without aberrations and for maintaining transparency.


Assuntos
Sequência de Aminoácidos/genética , Aquaporinas/genética , Catarata/genética , Proteínas do Olho/genética , Cristalino/patologia , Erros de Refração/genética , Deleção de Sequência/genética , Animais , Western Blotting , Catarata/embriologia , Catarata/fisiopatologia , Células Cultivadas , Modelos Animais de Doenças , Técnicas de Introdução de Genes , Imuno-Histoquímica , Camundongos , Camundongos Endogâmicos C57BL , Erros de Refração/embriologia , Erros de Refração/fisiopatologia , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Transfecção
15.
Nat Plants ; 5(3): 290-299, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30833710

RESUMO

Plants have evolved sophisticated systems in response to environmental changes, and growth arrest is a common strategy used to enhance stress tolerance. Despite the growth-survival trade-off being essential to the shaping of plant productivity, the mechanisms balancing growth and survival remain largely unknown. Aquaporins play a crucial role in growth and stress responses by controlling water transport across membranes. Here, we show that RhPIP2;1, an aquaporin from rose (Rosa sp.), interacts with a membrane-tethered MYB protein, RhPTM. Water deficiency triggers nuclear translocation of the RhPTM C terminus. Silencing of RhPTM causes continuous growth under drought stress and a consequent decrease in survival rate. RNA sequencing (RNA-seq) indicated that RhPTM influences the expression of genes related to carbohydrate metabolism. Water deficiency induces phosphorylation of RhPIP2;1 at Ser 273, which is sufficient to promote nuclear translocation of the RhPTM C terminus. These results indicate that the RhPIP2;1-RhPTM module serves as a key player in orchestrating the trade-off between growth and stress survival in Rosa.


Assuntos
Aquaporinas/metabolismo , Proteínas de Plantas/metabolismo , Rosa/fisiologia , Fatores de Transcrição/metabolismo , Aquaporinas/genética , Arabidopsis/genética , Metabolismo dos Carboidratos , Núcleo Celular/metabolismo , Citocininas/metabolismo , Desidratação , Secas , Regulação da Expressão Gênica de Plantas , Ácidos Indolacéticos/metabolismo , Fosforilação , Proteínas de Plantas/genética , Plantas Geneticamente Modificadas , Mapeamento de Interação de Proteínas , Rosa/crescimento & desenvolvimento , Serina/metabolismo , Transdução de Sinais , Fatores de Transcrição/genética
16.
J Med Food ; 22(3): 294-304, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30724689

RESUMO

Constipation is an acute or chronic illness attributed to various causes, ranging from lifestyle habits to side effects of a disease. To improve the laxative effects of some traditional medicines, herbal mixtures of Liriope platyphylla, Glycyrrhiza uralensis, and Cinnamomum cassia (LGC) were evaluated for their mechanism of action and therapeutic effects in loperamide (Lop)-induced constipated Sprague Dawley rats by examining alterations in excretion parameters, histological structure, mucin secretion, and related protein levels. Food intake and water consumption were constant for all animals. We observed that the Lop+LGC-treated group had significantly greater excretion of stool and urine than was observed in the Lop+Vehicle-treated group. Administration of LGC in the constipation model restored the intestinal transit ratio to normal levels, and increased the number of goblet cells, mucosal layer, and muscle thickness. Mucin secretion was greater in the Lop+LGC-treated group than in the Lop+Vehicle-treated group, and the expression of MUC2 and AQP8 genes were also increased. In addition, reverse transcription polymerase chain reaction and Western blot revealed an increase in the muscarinic acetylcholine receptors (mAChRs) in the Lop+LGC-treated group compared to the Lop+Vehicle-treated group. Furthermore, compared with the Lop+Vehicle-treated group, treatment with LGC reduced the phosphorylation of PKC and PI3K, and expression of Gα protein, but increased levels of IP3. Our results suggest that the traditional herbal mixture of LGC induces a potent laxative effect in Lop-induced constipation through mucosal tissue changes and mucin production. We also demonstrated that the laxative effect of LGC is closely related to the expression of mAChR and its downstream signals, suggesting the possibility of developing a constipation-laxative agent using LGC.


Assuntos
Cinnamomum aromaticum/química , Constipação Intestinal/tratamento farmacológico , Glycyrrhiza uralensis/química , Laxantes/administração & dosagem , Liriope (Planta)/química , Extratos Vegetais/administração & dosagem , Animais , Aquaporinas/genética , Aquaporinas/metabolismo , Constipação Intestinal/induzido quimicamente , Constipação Intestinal/genética , Constipação Intestinal/metabolismo , Sinergismo Farmacológico , Loperamida/efeitos adversos , Masculino , Mucina-2/genética , Mucina-2/metabolismo , Ratos , Ratos Sprague-Dawley
17.
PLoS One ; 14(2): e0212059, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30730995

RESUMO

Formation of adventitious roots in plants is a common response to hypoxia caused by flooding. In tobacco, after one week of root hypoxia treatment, plants produced twice as many adventitious roots as the aerated plants, but their maximum length was reduced. Hypoxia severely reduced net photosynthesis, transpiration rates, and photosynthetic light responses. Relative transcript abundance of the examined aquaporins in lateral roots was reduced by hypoxia, but in adventitious roots it remained unchanged. This apparent lack of an effect of root hypoxia on the aquaporin expression likely contributed to maintenance of high hydraulic conductance in adventitious roots. Lateral roots had lower porosity compared with adventitious roots and the expression of the ACS (1-aminocyclopropane-1-carboxylate synthase) gene was induced in hypoxic lateral roots, but not in adventitious roots, providing additional evidence that lateral roots were more affected by hypoxia compared with adventitious roots. ATP concentrations were markedly lower in both hypoxic lateral and adventitious roots compared with aerated roots, while the expression of fermentation-related genes, ADH1 (alcohol dehydrogenase 1) and PDC1 (pyruvate decarboxylase 1), was higher in lateral roots compared with adventitious roots. Since root porosity was greater in adventitious compared with lateral roots, the results suggest that the improved O2 delivery and stable root aquaporin expression in adventitious roots were likely the key factors helping flooded tobacco plants maintain high rates of root hydraulic conductance and, consequently, shoot gas exchange.


Assuntos
Aquaporinas/metabolismo , Hipóxia , Proteínas de Plantas/metabolismo , Tabaco/metabolismo , Trifosfato de Adenosina/metabolismo , Álcool Desidrogenase/genética , Álcool Desidrogenase/metabolismo , Aquaporinas/genética , Liases/genética , Liases/metabolismo , Proteínas de Plantas/genética , Raízes de Plantas/metabolismo , Piruvato Descarboxilase/genética , Piruvato Descarboxilase/metabolismo , Água/metabolismo
18.
Int J Mol Sci ; 20(4)2019 Feb 20.
Artigo em Inglês | MEDLINE | ID: mdl-30791644

RESUMO

Integral membrane proteins of the aquaporin family facilitate rapid water flux across cellular membranes in all domains of life. Although the water-conducting pore is clearly defined in an aquaporin monomer, all aquaporins assemble into stable tetramers. In order to investigate the role of protomer⁻protomer interactions, we analyzed the activity of heterotetramers containing increasing fractions of mutated monomers, which have an impaired oligomerization propensity and activity. In order to enforce interaction between the protomers, we designed and analyzed a genetically fused homotetramer of GlpF, the aquaglyceroporin of the bacterium Escherichia coli (E. coli). However, increasing fractions of the oligomerization-impaired mutant GlpF E43A affected the activity of the GlpF heterotetramer in a nearly linear manner, indicating that the reduced protein activity, caused by the introduced mutations, cannot be fully compensated by simply covalently linking the monomers. Taken together, the results underline the importance of exactly positioned monomer⁻monomer contacts in an assembled GlpF tetramer.


Assuntos
Aquaporinas/química , Aquaporinas/metabolismo , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/metabolismo , Multimerização Proteica , Aquaporinas/genética , Proteínas da Membrana Bacteriana Externa/química , Proteínas da Membrana Bacteriana Externa/genética , Proteínas da Membrana Bacteriana Externa/metabolismo , Transporte Biológico , Clonagem Molecular , Proteínas de Escherichia coli/genética , Expressão Gênica , Mutação , Proteínas Recombinantes de Fusão , Relação Estrutura-Atividade
19.
DNA Cell Biol ; 38(4): 374-384, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30807211

RESUMO

Aquaporins (AQPs) are transmembrane channels that are essential for the movement of water and other small molecules between biofilms in various physiological processes in plants. In this study, based on transcriptome-wide data, we identified and described a total of 21 AQP genes in common vetch (Vicia sativa subsp. sativa), which is an economically important pasture legume worldwide. Based on phylogenetic analyses, the VsAQPs were sorted into four subfamilies, including four plasma membrane intrinsic proteins (PIPs), six tonoplast intrinsic proteins (TIPs), seven NOD26-like intrinsic proteins, and four small basic intrinsic proteins. Furthermore, chemical and physical properties of these VsAQPs, including the isoelectric point and theoretical molecular weight, were analyzed. Analyses of the AQP signature sequences and key residues indicated the substrate specificity of each VsAQP. A set of VsAQPs was selected for gene expression analysis in a number of tissues and after drought stress treatments using real-time quantitative reverse transcription/polymerase chain reaction assays. Most of the PIPs and TIPs were proposed to have critical roles in regulating the flow of water during drought stress. Heterologous expression experiments in yeast indicated that VsPIP1;2 and VsPIP2;2 are key candidate genes for improving drought stress tolerance. The results reported in this study could be a crucial resource for further practical analyses and for genetic improvement of drought stress tolerance in common vetch.


Assuntos
Aquaporinas/genética , Secas , Perfilação da Expressão Gênica , Estresse Fisiológico/genética , Vicia sativa/genética , Vicia sativa/fisiologia , Sequência de Aminoácidos , Aquaporinas/química , Aquaporinas/metabolismo , Sequência Conservada , Filogenia
20.
Int J Mol Sci ; 20(3)2019 Jan 28.
Artigo em Inglês | MEDLINE | ID: mdl-30696013

RESUMO

Seed priming is a pre-sowing method successfully used to improve seed germination. Since water plays a crucial role in germination, the aim of this study was to investigate the relationship between better germination performances of osmoprimed Brassica napus seeds and seed water status during germination. To achieve this goal, a combination of different kinds of approaches was used, including nuclear magnetic resonance (NMR) spectroscopy, TEM, and SEM as well as semi-quantitative PCR (semi-qPCR). The results of this study showed that osmopriming enhanced the kinetics of water uptake and the total amount of absorbed water during both the early imbibition stage and in the later phases of seed germination. The spin⁻spin relaxation time (T2) measurement suggests that osmopriming causes faster water penetration into the seed and more efficient tissue hydration. Moreover, factors potentially affecting water relations in germinating primed seeds were also identified. It was shown that osmopriming (i) changes the microstructural features of the seed coat, e.g., leads to the formation of microcracks, (ii) alters the internal structure of the seed by the induction of additional void spaces in the seed, (iii) increases cotyledons cells vacuolization, and (iv) modifies the expression pattern of aquaporin genes.


Assuntos
Brassica napus/crescimento & desenvolvimento , Germinação , Sementes/crescimento & desenvolvimento , Água/fisiologia , Aquaporinas/genética , Aquaporinas/metabolismo , Brassica napus/ultraestrutura , Cotilédone/citologia , Cotilédone/ultraestrutura , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Cinética , Sementes/ultraestrutura , Vacúolos/metabolismo
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