RESUMO
The Elongator complex plays a pivotal role in the wobble uridine modification of the tRNA anticodon. Comprising two sets of six distinct subunits, namely, Elongator proteins (ELP1-ELP6) and associated proteins, the holo-Elongator complex demonstrates remarkable functional and structural conservation across eukaryotes. However, the precise details of the evolutionary conservation of the holo-Elongator complex and its individual sub-complexes (i.e., ELP123; ELP456) in plants remain limited. In this study, we conducted an in vivo analysis of protein-protein interactions among Arabidopsis ELP4, ELP5, and ELP6 proteins. Additionally, we predicted their structural configurations and performed a comparative analysis with the structure of the yeast Elp456 sub-complex. Protein-protein interaction analysis revealed that AtELP4 interacts with AtELP6 but not directly with AtELP5. Furthermore, we found that the Arabidopsis Elongator-associated protein, Deformed Roots and Leaves 1 (DRL1), did not directly bind to AtELP proteins. The structural comparison of the ELP456 sub-complex between Arabidopsis and yeast demonstrated high similarity, encompassing the RecA-ATPase fold and the positions of hydrogen bonds, despite their relatively low sequence homology. Our findings suggest that Arabidopsis ELP4, ELP5, and ELP6 proteins form a heterotrimer, with ELP6 serving as a bridge, indicating high structural conservation between the ELP456 sub-complexes from Arabidopsis and yeast.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Evolução Molecular , Ligação Proteica , Saccharomyces cerevisiae , Arabidopsis/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/genética , Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/genética , Modelos MolecularesRESUMO
The pharmacological potential of industrial hemp (Cannabis sativa) has been widely studied. However, the majority of studies have focused on cannabidiol, isolated from the inflorescence and leaf of the plant. In the present study, we evaluated the anti-diabetic potential of hemp root water (HWE) and ethanol extracts (HEE) in streptozotocin (STZ)-induced insulin-deficient diabetic mice. The administration of HWE and HEE ameliorated hyperglycemia and improved glucose homeostasis and islet function in STZ-treated mice (p < 0.05). HWE and HEE suppressed ß-cell apoptosis and cytokine-induced inflammatory signaling in the pancreas (p < 0.05). Moreover, HWE and HEE normalized insulin-signaling defects in skeletal muscles and apoptotic response in the liver and kidney induced by STZ (p < 0.05). Gas chromatography-mass spectrometry analysis of HWE and HEE showed possible active compounds which might be responsible for the observed anti-diabetic potential. These findings indicate the possible mechanisms by which hemp root extracts protect mice against insulin-deficient diabetes, and support the need for further studies geared towards the application of hemp root as a novel bioactive material.
Assuntos
Cannabis , Diabetes Mellitus Experimental , Camundongos , Animais , Cannabis/química , Insulina/farmacologia , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Experimental/induzido quimicamente , Extratos Vegetais/uso terapêutico , Pâncreas , Estreptozocina/farmacologiaRESUMO
Bacterial biofilm formation is a major cause of drug resistance and bacterial persistence; thus, controlling pathogenic biofilms is an important component of strategies targeting infectious bacterial diseases. Cinnamaldehyde (CNMA) has broad-spectrum antimicrobial and antibiofilm activities. In this study, we investigated the antibiofilm effects of ten CNMA derivatives and trans-CNMA against Gram-negative uropathogenic Escherichia coli (UPEC) and Gram-positive Staphylococcus aureus. Among the CNMA analogs tested, 4-nitrocinnamaldehyde (4-nitroCNMA) showed antibacterial and antibiofilm activities against UPEC and S. aureus with minimum inhibitory concentrations (MICs) for cell growth of 100 µg/mL, which were much more active than those of trans-CNMA. 4-NitroCNMA inhibited UPEC swimming motility, and both trans-CNMA and 4-nitroCNMA reduced extracellular polymeric substance production by UPEC. Furthermore, 4-nitroCNMA inhibited the formation of mixed UPEC/S. aureus biofilms. Collectively, our observations indicate that trans-CNMA and 4-nitroCNMA potently inhibit biofilm formation by UPEC and S. aureus. We suggest efforts be made to determine the therapeutic scope of CNMA analogs, as our results suggest CNMA derivatives have potential therapeutic use for biofilm-associated diseases.
Assuntos
Escherichia coli Uropatogênica , Acroleína/análogos & derivados , Antibacterianos/farmacologia , Biofilmes , Matriz Extracelular de Substâncias Poliméricas , Testes de Sensibilidade Microbiana , Staphylococcus aureusRESUMO
Phorbol 12-myristate 13-acetate (PMA) is a potent tumor promoter and highly inflammatory in nature. Here, we investigated the toxic effects of PMA on different model system. PMA (10 µg) caused chromosomal aberrations on the Allium cepa root tip and induced mitotic dysfunction. Similarly, PMA caused embryonic and larval deformities and a plummeted survivability rate on zebrafish embryo in a dose-dependent manner. Persistently, PMA treatment on immortalized human keratinocyte human keratinocyte (HaCaT) cells caused massive inflammatory rush at 4 h and a drop in cell survivability at 24 h. Concomitantly, we replicated a cutaneous inflammation similar to human psoriasis induced by PMA. Herein, we used tangeretin (TAN), as an antagonist to counteract the inflammatory response. Results from an in vivo experiment indicated that TAN (10 and 30 mg/kg) significantly inhibited PMA stimulated epidermal hyperplasia and intra-epidermal neutrophilic abscesses. In addition, its treatment effectively neutralized PMA induced elevated reactive oxygen species (ROS) generation on in vitro and in vivo systems, promoting antioxidant response. The association of hypoxia-inducible factor 1-alpha (HIF-1α)-nuclear factor kappa-light-chain-enhancer of activated b cells (NF-κB) crosstalk triggered by PMA enhanced PKCα-ERK1/2-NF-κB pathway; its activation was also significantly counteracted after TAN treatment. Conclusively, we demonstrated TAN inhibited the nuclear translocation of HIF-1α and NF-κB p65. Collectively, TAN treatment ameliorated PMA incited malignant inflammatory response by remodeling the cutaneous microenvironment.
Assuntos
Flavonas/farmacologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , NF-kappa B/metabolismo , Transdução de Sinais/efeitos dos fármacos , Acetato de Tetradecanoilforbol/efeitos adversos , Animais , Antioxidantes , Biomarcadores , Linhagem Celular Transformada , Anormalidades Congênitas , Desenvolvimento Embrionário/genética , Epiderme , Humanos , Inflamação/etiologia , Inflamação/metabolismo , Queratinócitos/metabolismo , Peroxidação de Lipídeos , Cebolas/efeitos dos fármacos , Cebolas/genética , Cebolas/metabolismo , Estresse Oxidativo , Espécies Reativas de Oxigênio/metabolismo , Peixe-ZebraRESUMO
BACKGROUND: Ginger leaf (GL) has long been used as a vegetable, tea and herbal medicine. However, its pharmacological properties are still poorly understood. Thus, we performed in vitro studies to evaluate anti-cancer properties of ginger leaf and then elucidate the potential mechanisms involved. METHODS: Cell viability was measured by MTT assay. ATF3 expression level was evaluated by Western blot or RT-PCR and ATF3 transcriptional activity was determined using a dual-luciferase assay kit after the transfection of ATF3 promoter constructs. In addition, ATF3-dependent apoptosis was evaluated by Western blot after ATF3 knockdown using ATF3 siRNA. RESULTS: Exposure of GL to human colorectal cancer cells (HCT116, SW480 and LoVo cells) reduced the cell viability and induced apoptosis in a dose-dependent manner. In addition, GL reduced cell viability in MCF-7, MDA-MB-231 and HepG-2 cells. ATF3 knockdown attenuated GL-mediated apoptosis. GL increased activating transcription factor 3 (ATF3) expressions in both protein and mRNA level and activated ATF3 promoter activity, indicating transcriptional activation of ATF3 gene by GL. In addition, our data showed that GL-responsible sites might be between -318 and -85 region of the ATF3 promoter. We also observed that ERK1/2 inhibition by PD98059 attenuated GL-mediated ATF3 expression but not p38 inhibition by SB203580, indicating ERK1/2 pathway implicated in GL-induced ATF3 activation. CONCLUSIONS: These findings suggest that the reduction of cell viability and apoptosis by GL may be a result of ATF3 promoter activation and subsequent increase of ATF3 expression through ERK1/2 activation in human colorectal cancer cells.
Assuntos
Fator 3 Ativador da Transcrição/genética , Antineoplásicos/farmacologia , Neoplasias Colorretais/genética , Extratos Vegetais/farmacologia , Folhas de Planta/química , Zingiber officinale/química , Fator 3 Ativador da Transcrição/metabolismo , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/fisiopatologia , Humanos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Regiões Promotoras Genéticas , Transdução de Sinais/efeitos dos fármacos , Ativação Transcricional/efeitos dos fármacosRESUMO
Cannabis belongs to the family Cannabaceae, and phytocannabinoids are produced by the Cannabis sativa L. plant. A long-standing debate regarding the plant is whether it contains one or more species. Phytocannabinoids are bioactive natural products found in flowers, seeds, and fruits. They can be beneficial for treating human diseases (such as multiple sclerosis, neurodegenerative diseases, epilepsy, and pain), the cellular metabolic process, and regulating biological function systems. In addition, several phytocannabinoids are used in various therapeutic and pharmaceutical applications. This study provides an overview of the different sources of phytocannabinoids; further, the biosynthesis of bioactive compounds involving various pathways is elucidated. The structural classification of phytocannabinoids is based on their decorated resorcinol core and the bioactivities of naturally occurring cannabinoids. Furthermore, phytocannabinoids have been studied in terms of their role in animal models and antimicrobial activity against bacteria and fungi; further, they show potential for therapeutic applications and are used in treating various human diseases. Overall, this review can help deepen the current understanding of the role of biotechnological approaches and the importance of phytocannabinoids in different industrial applications.
RESUMO
CtBP/BARS is a unique protein family in having quite diversified cellular functions, intercellular localizations, and developmental roles. ANGUSTIFOLIA (AN) is the sole homolog of CtBP/BARS from Arabidopsis thaliana, although it has plant AN-specific motifs and a long C-terminus. Previous studies suggested that AN would function in the nucleus as a transcriptional co-repressor, as CtBPs function in animals; however, precise verification has been lacking. In this paper, we isolated a homologous gene (MAN) of AN from liverwort, Marchantia polymorpha. Transformation of the Arabidopsis an-1 mutant with 35S-driven MAN completely complemented the an-1 phenotype, although it lacks the putative nuclear localization signal (NLS) that exists in AN proteins isolated from other plant species. We constructed several plasmids for expressing modified ANs with amino acid substitutions in known motifs. The results clearly indicated that modified AN with mutations in the putative NLS-like domain could complement the an-1 phenotype. Therefore, we re-examined localization of AN using several techniques. Our results demonstrated that AN localizes on punctuate structures around the Golgi, partially overlapping with a trans-Golgi network resident, which highlighted an unexpected link between leaf development and membrane trafficking. We should reconsider the roles and evolutionary traits of AN based on these findings.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Núcleo Celular/metabolismo , Marchantia/genética , Proteínas Repressoras/metabolismo , Motivos de Aminoácidos , Sequência de Aminoácidos , Substituição de Aminoácidos , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Núcleo Celular/genética , Genes de Plantas , Genes Reporter , Teste de Complementação Genética , Vetores Genéticos/genética , Vetores Genéticos/metabolismo , Marchantia/metabolismo , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Meristema/metabolismo , Meristema/ultraestrutura , Microscopia Eletrônica de Transmissão , Dados de Sequência Molecular , Mutação , Sinais de Localização Nuclear/metabolismo , Fenótipo , Células Vegetais/metabolismo , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/metabolismo , Plasmídeos/genética , Plasmídeos/metabolismo , RNA de Plantas/genética , RNA de Plantas/metabolismo , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Proteínas Repressoras/genética , Especificidade da Espécie , Transformação Genética , Rede trans-Golgi/metabolismo , Rede trans-Golgi/ultraestruturaRESUMO
The Elongator complex in eukaryotes has conserved tRNA modification functions and contributes to various physiological processes such as transcriptional control, DNA replication and repair, and chromatin accessibility. ARABIDOPSIS ELONGATOR PROTEIN 4 (AtELP4) is one of the six subunits (AtELP1-AtELP6) in Arabidopsis Elongator. In addition, there is an Elongator-associated protein, DEFORMED ROOTS AND LEAVES 1 (DRL1), whose homolog in yeast (Kti12) binds tRNAs. In this study, we explored the functions of AtELP4 in plant-specific aspects such as leaf morphogenesis and evolutionarily conserved ones between yeast and Arabidopsis. ELP4 comparison between yeast and Arabidopsis revealed that plant ELP4 possesses not only a highly conserved P-loop ATPase domain but also unknown plant-specific motifs. ELP4 function is partially conserved between Arabidopsis and yeast in the growth sensitivity toward caffeine and elevated cultivation temperature. Either single Atelp4 or drl1-102 mutants and double Atelp4 drl1-102 mutants exhibited a reduction in cell proliferation and changed the adaxial-abaxial polarity of leaves. In addition, the single Atelp4 and double Atelp4 drl1-102 mutants showed remarkable downward curling at the whole part of leaf blades in contrast to wild-type leaf blades. Furthermore, our genetic study revealed that AtELP4 might epistatically act on DRL1 in the regulation of cell proliferation and dorsoventral polarity in leaves. Taken together, we suggest that AtELP4 as part of the plant Elongator complex may act upstream of a regulatory pathway for adaxial-abaxial polarity and cell proliferation during leaf development.
RESUMO
Effective treatment choices to the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) are limited because of the absence of effective target-based therapeutics. The main object of the current research was to estimate the antiviral activity of cannabinoids (CBDs) against the human coronavirus SARS-CoV-2. In the presented research work, we performed in silico and in vitro experiments to aid the sighting of lead CBDs for treating the viral infections of SARS-CoV-2. Virtual screening was carried out for interactions between 32 CBDs and the SARS-CoV-2 Mpro enzyme. Afterward, in vitro antiviral activity was carried out of five CBDs molecules against SARS-CoV-2. Interestingly, among them, two CBDs molecules namely Δ9 -tetrahydrocannabinol (IC50 = 10.25 µM) and cannabidiol (IC50 = 7.91 µM) were observed to be more potent antiviral molecules against SARS-CoV-2 compared to the reference drugs lopinavir, chloroquine, and remdesivir (IC50 ranges of 8.16-13.15 µM). These molecules were found to have stable conformations with the active binding pocket of the SARS-CoV-2 Mpro by molecular dynamic simulation and density functional theory. Our findings suggest cannabidiol and Δ9 -tetrahydrocannabinol are possible drugs against human coronavirus that might be used in combination or with other drug molecules to treat COVID-19 patients.
Assuntos
Antivirais/farmacologia , Tratamento Farmacológico da COVID-19 , COVID-19/virologia , Canabinoides/farmacologia , SARS-CoV-2/efeitos dos fármacos , Antivirais/química , Antivirais/farmacocinética , Canabidiol/química , Canabidiol/farmacocinética , Canabidiol/farmacologia , Canabinoides/química , Canabinoides/farmacocinética , Simulação por Computador , Proteases 3C de Coronavírus/antagonistas & inibidores , Proteases 3C de Coronavírus/química , Proteases 3C de Coronavírus/efeitos dos fármacos , Dronabinol/química , Dronabinol/farmacocinética , Dronabinol/farmacologia , Avaliação Pré-Clínica de Medicamentos , Humanos , Técnicas In Vitro , Ligantes , Modelos Biológicos , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Pandemias , Inibidores de Proteases/química , Inibidores de Proteases/farmacologia , SARS-CoV-2/químicaRESUMO
Decursinol angelate (DA) is a pyranocoumarin purified from the roots of Angelica gigas. Here, we synthesized DA and determined its anti-inflammatory potential on TPA-induced mice ear inflammation. First, we evaluated the non-toxic behaviour of DA on HaCaT cells. Additionally, we observed the free radical scavenging potential of DA at 60⯵M to be 50%. This finding was further supported by nitric oxide assay, malondialdehyde assay, H2DCFDA staining and western blotting analysis of antioxidant enzymes. DA also suppressed the activation and polarization of macrophage phagocytic activity on RAW 264.7â¯cells. We further evaluated the expression of ICAM-1, MCP-1, MIP-2 and MIP-1ß on in-vivo model system. Consequently, DA significantly reduced the production of NF-κB and COX-2 induced proinflammatory cytokine levels on TPA induced ear edema. Inhibition of MAPK and transcriptional factor NF-κB was also validated by western blotting analysis of p-ERK, p-p38, IKKα, IKKγ, IκBα, NF-κB-p65. Immunohistochemistry and immunofluorescence staining of NFκB-p65, TNF-α and IL-1ß were also performed to support the findings. Conclusively, these results suggest that topical administration of DA significantly inhibited the expression of pro-inflammatory cytokines by blocking the canonical NF-κB and MAPK pathway. Therefore, we suggest DA as a potent therapeutic compound against skin inflammation related diseases.
Assuntos
Benzopiranos/farmacologia , Butiratos/farmacologia , Citocinas/biossíntese , Mediadores da Inflamação/metabolismo , NF-kappa B/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Acetato de Tetradecanoilforbol/farmacologia , Animais , Antioxidantes/farmacologia , Linhagem Celular , Orelha , Humanos , Ativação de Macrófagos/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Células RAW 264.7 , Pele/efeitos dos fármacos , Pele/metabolismoRESUMO
The seminal discovery of browning of white adipose tissue (WAT) holds great promise for the treatment of obesity and metabolic syndrome. DJ-1 is evolutionarily conserved across species, and mutations in DJ-1 have been identified in Parkinson's disease. Higher levels of DJ-1 are associated with obesity, but the underlying mechanism is less understood. Here, we report the previously unappreciated role of DJ-1 in white adipocyte biology in mature models of obesity. We used DJ-1 knockout (KO) mouse models and wild-type littermates maintained on a normal diet or high-fat diet as well as in vitro cell models to show the direct effects of DJ-1 depletion on adipocyte phenotype, thermogenic capacity, fat metabolism, and microenvironment profile. Global DJ-1 KO mice show increased sympathetic input to WAT and ß3-adrenergic receptor intracellular signaling, leading to a previously unrecognized compensatory mechanism through browning of WAT with associated characteristics, including high mitochondrial contents, reduced lipid accumulation, adequate vascularization and attenuated autophagy. DJ-1 KO mice had normal body weight, energy balance, and adiposity, which were associated with protective effects on healthy WAT expansion by hyperplasia. Our findings revealed that browning of inguinal WAT occurred in DJ-1 KO mice that do not show increased predisposition to obesity and suggest that such potential mechanism may overcome the adverse metabolic consequences of obesity independent of an effect on body weight. Here, we provide the first direct evidence that targeting DJ-1 in adipocyte metabolic health may offer a unique therapeutic strategy for the treatment of obesity.
Assuntos
Adipócitos Marrons/patologia , Tecido Adiposo Branco/patologia , Obesidade/etiologia , Proteína Desglicase DJ-1/genética , Células 3T3-L1 , Adipócitos Marrons/fisiologia , Adipócitos Brancos/citologia , Animais , Autofagia , Peso Corporal/genética , Dieta Hiperlipídica/efeitos adversos , Metabolismo Energético/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Obesidade/genética , Obesidade/patologia , Proteína Desglicase DJ-1/metabolismoRESUMO
Patterning of the polar axis during the early leaf developmental stage is established by cell-to-cell communication between the shoot apical meristem (SAM) and the leaf primordia. In a previous study, we showed that the DRL1 gene, which encodes a homolog of the Elongator-associated protein KTI12 of yeast, acts as a positive regulator of adaxial leaf patterning and shoot meristem activity. To determine the evolutionally conserved functions of DRL1, we performed a comparison of the deduced amino acid sequence of DRL1 and its yeast homolog, KTI12, and found that while overall homology was low, well-conserved domains were presented. DRL1 contained two conserved plant-specific domains. Expression of the DRL1 gene in a yeast KTI12-deficient yeast mutant suppressed the growth retardation phenotype, but did not rescue the caffeine sensitivity, indicating that the role of Arabidopsis Elongator-associated protein is partially conserved with yeast KTI12, but may have changed between yeast and plants in response to caffeine during the course of evolution. In addition, elevated expression of DRL1 gene triggered zymocin sensitivity, while overexpression of KTI12 maintained zymocin resistance, indicating that the function of Arabidopsis DRL1 may not overlap with yeast KTI12 with regards to toxin sensitivity. In this study, expression analysis showed that class-I KNOX genes were downregulated in the shoot apex, and that YAB and KAN were upregulated in leaves of the Arabidopsis drl1-101 mutant. Our results provide insight into the communication network between the SAM and leaf primordia required for the establishment of leaf polarity by mediating histone acetylation or through other mechanisms.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/fisiologia , Proteínas de Arabidopsis/fisiologia , Arabidopsis/genética , Proteínas de Ligação ao GTP/fisiologia , Proteínas de Saccharomyces cerevisiae/fisiologia , Saccharomyces cerevisiae/genética , Proteínas Adaptadoras de Transdução de Sinal/química , Sequência de Aminoácidos , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Proteínas de Arabidopsis/química , Cafeína/farmacologia , Sequência Conservada , Proteínas de Ligação ao GTP/química , Teste de Complementação Genética , Dados de Sequência Molecular , Saccharomyces cerevisiae/efeitos dos fármacos , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/química , Estresse FisiológicoRESUMO
ANGUSTIFOLIA (AN) controls leaf morphology in the plant Arabidopsis thaliana. Previous studies on sequence similarity demonstrated that the closest proteins to AN are members of animal C-terminal-binding proteins (CtBPs) found in nematodes, arthropods, and vertebrates. Drosophila CtBP (dCtBP) functions as a transcriptional corepressor for deoxyribonucleic acid (DNA)-binding repressors containing the short amino acid motif, PXDLS, to regulate tissue specification and segmentation during early embryogenesis. It has previously been shown that AN was thought to repress transcription similar to the function of CtBPs; however, AN lacks some of the structural features that are conserved in animal CtBPs. In this paper, we examined whether AN is functionally related to dCtBP. Firstly, we re-examined sequence similarity among AN and various CtBPs from several representative species in the plant and animal kingdoms. Secondly, yeast two-hybrid assays demonstrated that AN failed to interact with an authentic CtBP-interacting factor, adenovirus E1A oncoprotein bearing the PXDLS motif. Thirdly, AN tethered to DNA was unable to repress the expression of reporter genes in transgenic Drosophila embryos. Fourthly, overexpression assays suggested that dCtBP and AN function differently in Drosophila tissues. Finally, AN failed to rescue the zygotic lethality caused by dCtBP loss-of-function. These data, taken together, suggest that AN is functionally distinct from dCtBP. Likely, ancestral CtBPs acquired corepressor function (capability of both repression and binding to repressors containing the PXDLS motif) after the animal-plant divergence but before the protostome-deuterostome split. We therefore propose to categorize AN as a subfamily member within the CtBP/BARS/RIBEYE/AN superfamily.
Assuntos
Oxirredutases do Álcool/química , Oxirredutases do Álcool/metabolismo , Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/metabolismo , Drosophila melanogaster/metabolismo , Proteínas Repressoras/química , Proteínas Repressoras/metabolismo , Homologia Estrutural de Proteína , Motivos de Aminoácidos , Sequência de Aminoácidos , Animais , DNA/metabolismo , Drosophila melanogaster/embriologia , Embrião não Mamífero/metabolismo , Evolução Molecular , Teste de Complementação Genética , Dados de Sequência Molecular , Fenótipo , Ligação Proteica , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Transgenes , Zigoto/metabolismoRESUMO
ANGUSTIFOLIA (AN) is the first C-terminal binding protein (CtBP) gene from plants and controls leaf width and pattern of trichome branching in Arabidopsis thaliana (L.) Heynh. We characterized an ortholog of AN from Ipomoea nil (L.) Roth (Japanese morning glory) and designated it Ipomoea nil's AN (IAN). IAN is a single-copy gene in the genome and is expressed ubiquitously in various organs of I. nil. IAN contains not only a D2-HDH motif, which is highly conserved within the CtBP family, but also LXCXE, NLS and PEST motifs, which are specific to the AN subfamily. The expression of IAN cDNA driven by the cauliflower mosaic virus 35S promoter restored a defect in leaf expansion in the leaf width direction in the angustifolia-1 (an-1) mutant of Arabidopsis, suggesting that IAN retains a common function with AN. In contrast, the complementation by IAN of a defect in the trichome branching pattern on the leaf surface of the an-1 mutant was less effective than that observed for leaf shape. These results suggest that the mechanisms by which AN regulates leaf width and trichome branching are separable.
Assuntos
Genes de Plantas , Ipomoea/genética , Sequência de Aminoácidos , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Sequência de Bases , DNA Complementar/genética , DNA de Plantas/genética , Ipomoea/crescimento & desenvolvimento , Ipomoea/metabolismo , Dados de Sequência Molecular , Mutação , Plantas Geneticamente Modificadas , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA de Plantas/genética , RNA de Plantas/metabolismo , Proteínas Repressoras/genética , Homologia de Sequência de Aminoácidos , Especificidade da EspécieRESUMO
We previously showed that the ANGUSTIFOLIA (AN) gene regulates the width of leaves of Arabidopsis thaliana, by controlling the polar elongation of leaf cells. In the present study, we found that the abnormal arrangement of cortical microtubules (MTs) in an leaf cells appeared to account entirely for the abnormal shape of the cells. It suggested that the AN gene might regulate the polarity of cell growth by controlling the arrangement of cortical MTs. We cloned the AN gene using a map-based strategy and identified it as the first member of the CtBP family to be found in plants. Wild-type AN cDNA reversed the narrow-leaved phenotype and the abnormal arrangement of cortical MTs of the an-1 mutation. In the animal kingdom, CtBPs self-associate and act as co-repressors of transcription. The AN protein can also self-associate in the yeast two-hybrid system. Furthermore, microarray analysis suggested that the AN gene might regulate the expression of certain genes, e.g. the gene involved in formation of cell walls, MERI5. A discussion of the molecular mechanisms involved in the leaf shape regulation is presented based on our observations.