ABSTRACT
Emotional stability-Neuroticism is a complex construct influenced by genetics and environmental factors. Women tend to exhibit higher neuroticism scores than men, which may be associated with an increased risk of suffering from some common mental conditions. Some authors have pointed out the influence of sex hormones, since they induce sexual differentiation of the brain that can lead to sex-specific behaviors. 2D:4D digit ratio is commonly used as a marker of prenatal sex hormones. In this study we analyzed whether there was an association between 2D:4D and personality measured through the BFQ in a homogeneous sample of 101 young women college students. We found a positive association between 2D:4D and emotional stability, as well as with its subdimensions emotion control and impulse control. This association could be quadratic and nonlinear. However, no association was found with the other four dimensions. We also measured anxiety, depression and global life satisfaction, variables related to neuroticism. We observed that emotional stability is positively associated to social desirability and global life satisfaction, and negatively related to anxiety and depression. On the other hand, we did not find any association between 2D:4D and anxiety, depression, and global life satisfaction. These results can be linked to other aspects such as subjective well-being and psychopathological symptoms. This study may help to better understand how these constructs are related and could lead to future projects to elucidated how these variables influence personality.
Subject(s)
Anxiety/physiopathology , Emotions/physiology , Gonadal Steroid Hormones/metabolism , Mental Disorders/epidemiology , Adult , Anxiety/epidemiology , Anxiety/metabolism , Emotional Regulation/physiology , Female , Gonadal Steroid Hormones/genetics , Humans , Male , Mental Disorders/physiopathology , Neuroticism/physiology , Personal Satisfaction , Personality/genetics , Personality/physiology , Pregnancy , Sex Characteristics , Sex Differentiation/genetics , Sex Differentiation/physiology , Sexual Behavior/physiology , Social Desirability , Students/psychology , Young AdultABSTRACT
BACKGROUND: Neuroticism is associated with low emotional stability, and it is characterized by a tendency to perceive ordinary situations as threatening and difficult to manage. This personality trait has been associated with psychological distress and predicts some mental disorders. Previous studies have shown that women tend to be more neurotic than men and, in general, females have also a higher incidence of anxious and depressive disorders. METHODS: We analyzed in a sample of 99 female university students (from 18 to 26 years old) if emotional stability, measured using the Big Five Questionnaire, was linked to polymorphic variants in candidate genes related to dopaminergic and serotonergic systems, and other personality variables. RESULTS: We found that emotional stability and its subdimensions are genetically associated with MAOA-uVNTR polymorphism. Thus, women carriers of the 3-repeat allele (lower MAO-A expression) showed higher levels of emotional stability. No associations were found with other polymorphisms analyzed, including COMT Val158 Met, 5-HTTLPR, and DAT 3'UTR VNTR. Furthermore, our results showed a negative correlation between emotional stability and depression, state anxiety, and trait anxiety. In fact, MAOA-uVNTR and trait anxiety also explained emotional stability and its subdimensions. We also found that other genetic characteristic, phenylthiocarbamide tasting, explained impulsivity, specifically tasters controlled impulses better than nontasters. CONCLUSION: Our results indicate that neuroticism might be regulated by MAOA and could be a common factor between different phenotypes, such as aggressive behaviors or personality disorders, observed in women with higher activity genotype who had been exposed to negative environments during childhood. This study could lead to a better understanding of the basis of emotional stability and could lead to future projects for this purpose.
Subject(s)
Emotions , Monoamine Oxidase/genetics , Neuroticism , Polymorphism, Genetic/genetics , Promoter Regions, Genetic/genetics , Adolescent , Adult , Emotional Regulation , Female , Humans , Students/psychology , Students/statistics & numerical data , Young AdultABSTRACT
The neurotransmitters dopamine and serotonin participate in specific behavioral neuromuscular mechanisms in the nematode Caenorhabditis elegans. Dopamine is involved in the gentle touch response and serotonin in the pharyngeal pumping rate. In its genome, the worm presents genes encoding dopamine and serotonin receptors orthologous to those of human genes. Risperidone and aripiprazole are a class of drugs known as atypical antipsychotics commonly used to treat schizophrenia, bipolar disorder, and irritability associated with autism. Risperidone is an antagonist of the dopamine D2 and serotonin 5-HT2A receptors. Aripiprazole functions as a partial agonist of the dopamine D2 receptor and as a partial agonist and antagonist of 5-HT1A and 5-HT2A serotonin receptors, respectively. Our results show that risperidone and aripiprazole alter the touch response and pharyngeal pumping in wild-type worm animals. Furthermore, in the presence of the drugs, both behaviors change to varying degrees in dopamine (dop-1, dop-2, and dop-3), serotonin (ser-1), and tyramine (ser-2) receptor-deficient mutants. This variation in response reveals specific targets for these antipsychotics in the nematode. Interestingly, their effect on behavior persisted to some extent in successive generations, indicating that they might induce epigenetic changes throughout development. Sodium butyrate, a histone deacetylase inhibitor, eliminated the consecutive generation effect of both drugs. In addition, these transgenerational effects were also abolished after the dauer stage. These observations suggest that risperidone and aripiprazole, in addition to interacting with specific receptors impairing the function of the nervous system of the nematode, may lead to the deposition of long-lasting epigenetic marks.
ABSTRACT
Neurexins and neuroligins are neuronal membrane adhesion molecules that have been involved in neuropsychiatric and neurodevelopmental disorders. The nrx-1 and nlg-1 genes of Caenorhabditis elegans encode NRX-1 and NLG-1, orthologue proteins of human neurexins and neuroligins, respectively. Dopaminergic and serotoninergic signalling control the locomotory rate of the nematode. When well-fed animals are transferred to a plate with food (bacterial lawn), they reduce the locomotory rate. This behavior, which depends on dopamine, is known as basal slowing response (BSR). Alternatively, when food-deprived animals are moved to a plate with a bacterial lawn, further decrease their locomotory rate. This behavior, known as enhanced slowing response (ESR), is serotonin dependent. C. elegans nlg-1-deficient mutants are impaired in BSR and ESR. Here we report that nrx-1-deficient mutants were defective in ESR, but not in BSR. The nrx-1;nlg-1 double mutant was impaired in both behaviors. Interestingly, the nlg-1 mutants upregulate the expression of comt-4 which encodes an enzyme with putative catechol-O-methyltransferase activity involved in dopamine degradation. Our study also shows that comt-4(RNAi) in nlg-1-deficient mutants rescues the wild type phenotypes of BSR and ESR. On the other hand, comt-4(RNAi) in nlg-1-deficient mutants also recovers, at least partially, the gentle touch response and the pharyngeal pumping rate that were impaired in these mutants. These latter behaviors are dopamine and serotonin dependent, respectively. Based on these results we propose a model for the neuroligin function in modulating the dopamine-dependent locomotory behavior in the nematode.
Subject(s)
Caenorhabditis elegans Proteins/metabolism , Cell Adhesion Molecules, Neuronal/genetics , Cell Adhesion Molecules, Neuronal/metabolism , Animals , Caenorhabditis elegans/genetics , Caenorhabditis elegans/metabolism , Caenorhabditis elegans Proteins/genetics , Catechol O-Methyltransferase/genetics , Catechol O-Methyltransferase/metabolism , Catechol O-Methyltransferase/physiology , Dopamine/metabolism , Locomotion/genetics , Locomotion/physiology , RNA Interference , Serotonin/metabolismABSTRACT
Current research indicates that the causes of autism spectrum disorders (ASDs) are multifactorial and include both genetic and environmental factors. To date, several works have associated ASDs with mutations in genes that encode proteins involved in neuronal synapses; however other factors and the way they can interact with the development of the nervous system remain largely unknown. Some studies have established a direct relationship between risk for ASDs and the exposure of the fetus to high testosterone levels during the prenatal stage. In this work, in order to explain possible mechanisms by which this androgenic hormone may interact with the nervous system, C. elegans was used as an experimental model. We observed that testosterone was able to alter the behavioral pattern of the worm, including the gentle touch response and the pharyngeal pumping rate. This impairment of the behavior was abolished using specific RNAi against genes orthologous to the human androgen receptor gene. The effect of testosterone was eliminated in the nhr-69 (ok1926) deficient mutant, a putative ortholog of human AR gene, suggesting that this gene encodes a receptor able to interact with the hormone. On the other hand the testosterone effect remained in the gentle touch response during four generations in the absence of the hormone, indicating that some epigenetic mechanisms could be involved. Sodium butyrate, a histone deacetylase inhibitor, was able to abolish the effect of testosterone. In addition, the lasting effect of testosterone was eliminated after the dauer stage. These results suggest that testosterone may impair the nervous system function generating transgenerational epigenetic marks in the genome. This work may provide new paradigms for understanding biological mechanisms involved in ASDs traits.
ABSTRACT
Neuroligins are neuronal and neuromuscular transmembrane proteins that have been implicated in autism spectrum disorder and other cognitive diseases. The nlg-1 gene from Caenorhabditis elegans is orthologous to human neuroligin genes. In the nematode, the locomotory rate is mediated by dopaminergic and serotonergic pathways, which result in two different behavioral responses known as basal slowing response (BSR) and enhanced slowing response (ESR), respectively. We report that nlg-1-deficient mutants are defective in both the BSR and ESR behaviors. In addition, we demonstrate that methylphenidate (a dopamine reuptake inhibitor) and fluoxetine (a serotonin reuptake inhibitor), two drugs widely used for the treatment of behavioral disorders in humans, are able to restore the BSR and ESR wild type phenotypes, respectively, in nlg-1 defective mutant nematodes. The abnormal locomotory behavior patterns were rescued in nlg-1-deficient mutant by expressing a cDNA from the human NLGN1 gene under the C. elegans nlg-1 promoter. However, human NLGN1 (R453C) and NLGN1 (D432X) mutant alleles did not rescue any of the two mutant phenotypes. The results indicate that neuroligin is involved in modulating the action of dopamine and serotonin in the nematode and suggest that the functional mechanism underpinning both methylphenidate and fluoxetine in C. elegans might be comparable to that in humans. The neuroligin-deficient mutants may undergo inefficient synaptic transmissions which could affect different traits in the nervous system. In particular, neuroligin might be required for normal neurotransmitters release. The understanding of the mechanisms by which methylphenidate and fluoxetine are able to restore the behavior of these mutants could help to explain the etiology of some human neurological diseases.
Subject(s)
Cell Adhesion Molecules, Neuronal/physiology , Dopamine/metabolism , Locomotion/physiology , Serotonin/metabolism , Animals , Caenorhabditis elegans , Cell Adhesion Molecules, Neuronal/genetics , Dopamine Uptake Inhibitors/pharmacology , Fluoxetine/pharmacology , Locomotion/drug effects , Methylphenidate/pharmacology , Neural Pathways/physiology , Selective Serotonin Reuptake Inhibitors/pharmacologyABSTRACT
Neuroligins are cell adhesion proteins that interact with neurexins at the synapse. This interaction may contribute to differentiation, plasticity and specificity of synapses. In humans, single mutations in neuroligin encoding genes lead to autism spectrum disorder and/or mental retardation. Caenorhabditis elegans mutants deficient in nlg-1, an orthologue of human neuroligin genes, have defects in different behaviors. Here we show that the expression of human NLGN1 or rat Nlgn1 cDNAs in C. elegans nlg-1 mutants rescues the fructose osmotic strength avoidance and gentle touch response phenotypes. Two specific point mutations in NLGN3 and NLGN4 genes, involved in autistic spectrum disorder, were further characterized in this experimental system. The R451C allele described in NLGN3, was analyzed with both human NLGN1 (R453C) and worm NLG-1 (R437C) proteins, and both were not functional in rescuing the osmotic avoidance behavior and the gentle touch response phenotype. The D396X allele described in NLGN4, which produces a truncated protein, was studied with human NLGN1 (D432X) and they did not rescue any of the behavioral phenotypes analyzed. In addition, RNAi feeding experiments measuring gentle touch response in wild type strain and worms expressing SID-1 in neurons (which increases the response to dsRNA), both fed with bacteria expressing dsRNA for nlg-1, provided evidence for a postsynaptic in vivo function of neuroligins both in muscle cells and neurons, equivalent to that proposed in mammals. This finding was further confirmed generating transgenic nlg-1 deficient mutants expressing NLG-1 under pan-neuronal (nrx-1) or pan-muscular (myo-3) specific promoters. All these results suggest that the nematode could be used as an in vivo model for studying particular synaptic mechanisms with proteins orthologues of humans involved in pervasive developmental disorders.
Subject(s)
Caenorhabditis elegans/genetics , Caenorhabditis elegans/metabolism , Cell Adhesion Molecules, Neuronal/genetics , Cell Adhesion Molecules, Neuronal/metabolism , Mutation , Phenotype , Animals , Animals, Genetically Modified , Cell Adhesion Molecules, Neuronal/deficiency , DNA, Complementary , Fructose/metabolism , Gene Expression , Humans , Muscle Cells/metabolism , Neurons/metabolism , Osmosis , RNA Interference , Rats , TouchABSTRACT
The nematode Caenorhabditis elegans has a very well-defined and genetically tractable nervous system which offers an effective model to explore basic mechanistic pathways that might be underpin complex human neurological diseases. Here, the role C. elegans is playing in understanding two neurodegenerative conditions, Parkinson's and Alzheimer's disease (AD), and a complex neurological condition, autism, is used as an exemplar of the utility of this model system. C. elegans is an imperfect model of Parkinson's disease because it lacks orthologues of the human disease-related genes PARK1 and LRRK2 which are linked to the autosomal dominant form of this disease. Despite this fact, the nematode is a good model because it allows transgenic expression of these human genes and the study of the impact on dopaminergic neurons in several genetic backgrounds and environmental conditions. For AD, C. elegans has orthologues of the amyloid precursor protein and both human presenilins, PS1 and PS2. In addition, many of the neurotoxic properties linked with Aß amyloid and tau peptides can be studied in the nematode. Autism spectrum disorder is a complex neurodevelopmental disorder characterised by impairments in human social interaction, difficulties in communication, and restrictive and repetitive behaviours. Establishing C. elegans as a model for this complex behavioural disorder is difficult; however, abnormalities in neuronal synaptic communication are implicated in the aetiology of the disorder. Numerous studies have associated autism with mutations in several genes involved in excitatory and inhibitory synapses in the mammalian brain, including neuroligin, neurexin and shank, for which there are C. elegans orthologues. Thus, several molecular pathways and behavioural phenotypes in C. elegans have been related to autism. In general, the nematode offers a series of advantages that combined with knowledge from other animal models and human research, provides a powerful complementary experimental approach for understanding the molecular mechanisms and underlying aetiology of complex neurological diseases.
Subject(s)
Alzheimer Disease/genetics , Caenorhabditis elegans , Child Development Disorders, Pervasive/genetics , Disease Models, Animal , Parkinson Disease/genetics , Alzheimer Disease/metabolism , Amyloid beta-Protein Precursor/genetics , Amyloid beta-Protein Precursor/metabolism , Animals , Animals, Genetically Modified , Caenorhabditis elegans/genetics , Caenorhabditis elegans/metabolism , Caenorhabditis elegans Proteins/genetics , Caenorhabditis elegans Proteins/metabolism , Child , Child Development Disorders, Pervasive/metabolism , Dopaminergic Neurons/metabolism , Humans , Leucine-Rich Repeat Serine-Threonine Protein Kinase-2 , Nerve Degeneration/genetics , Nerve Degeneration/metabolism , Parkinson Disease/metabolism , Presenilins/genetics , Presenilins/metabolism , Protein Serine-Threonine Kinases/genetics , Protein Serine-Threonine Kinases/metabolism , Synapses/metabolism , alpha-Synuclein/genetics , alpha-Synuclein/metabolismABSTRACT
INTRODUCTION: Bipolar disorder is a severe mental disease of unknown etiology that affects about 1% of the population. It is characterized by mood changes, alternating episodes of mania with depression. The current experimental and epidemiological data suggest that bipolar disorder represents a group of disorders with similar symptoms caused by the combination of genetic and environmental factors. Genetic analysis has identified several genes whose dysfunction might predispose to the disorder, although most of the results have not been confirmed in other studies. DEVELOPMENT: The aim of this review is to analyze the origin of bipolar disorder from a genetic perspective, with emphasis on miRNAs encoding genes. The response of patients to drugs such as lithium and valproate, and a series of data from different experimental approaches show that in some cases of bipolar disorder, genes encoding miRNAs might be involved. These non coding RNAs elements regulate gene expression and could participate in the development of diseases such as cancer, immune system disorders, heart disease and different mental and neurological disorders. CONCLUSIONS: Because the administration of mood stabilizers has been found to modify the expression of some miRNAs, this paper suggests that knowledge of the functions of these regulators could help to diagnose some cases of bipolar disorder identifying molecular markers in plasma. This methodology would also allow applying new strategy founded on effective therapeutic targets for personalized treatment.
Subject(s)
Bipolar Disorder/diagnosis , Bipolar Disorder/genetics , Bipolar Disorder/therapy , MicroRNAs/therapeutic use , Animals , Biomarkers/blood , Central Nervous System Agents/therapeutic use , Epigenesis, Genetic , Genetic Predisposition to Disease , Genome-Wide Association Study , Humans , Precision MedicineABSTRACT
Introducción. El trastorno bipolar es una grave enfermedad mental de etiología desconocida que afecta aproximadamente al 1% de la población. Se caracteriza por cambios del estado de ánimo, alternándose episodios de manía con estados depresivos. Los datos epidemiológicos y experimentales actuales sugieren que es un conjunto de trastornos con sintomatologías similares que se originan por la combinación de factores genéticos y ambientales. Los análisis genéticos han identificado varios genes cuya disfunción podrían predisponer a padecer el trastorno, aunque la mayoría de las veces los resultados no han podido ser confirmados en otros estudios. Desarrollo. El objetivo de esta revisión es analizar las causas que originan el trastorno bipolar desde una perspectiva genética, haciendo hincapié en los genes que codifican microARN. La respuesta de los pacientes a fármacos como el litio y el ácido valproico, y una serie de datos procedentes de diferentes metodologías experimentales, señalan que, en ciertos casos de trastorno bipolar, podrían estar implicados estos pequeños ácidos ribonucleicos que no se traducen a proteínas. Los microARN regulan la expresión génica y pueden participar en el desarrollo de enfermedades como el cáncer, del sistema inmune, enfermedades coronarias, y diferentes trastornos mentales y neurológicos. Conclusiones. Partiendo del hecho de que la administración de estabilizadores del ánimo modifica la expresión de determinados microARN, este trabajo sugiere que el conocimiento de las funciones de estos elementos reguladores podría facilitar el diagnóstico de algunos casos de trastorno bipolar, cuantificando estos marcadores moleculares en plasma. Esta metodología permitiría, además, tratamientos más eficaces basados en el diseño de nuevas estrategias terapéuticas personalizadas en función de dianas específicas de microARN (AU)
Introduction. Bipolar disorder is a severe mental disease of unknown etiology that affects about 1% of the population. It is characterized by mood changes, alternating episodes of mania with depression. The current experimental and epidemiological data suggest that bipolar disorder represents a group of disorders with similar symptoms caused by the combination of genetic and environmental factors. Genetic analysis has identified several genes whose dysfunction might predispose to the disorder, although most of the results have not been confirmed in other studies. Development. The aim of this review is to analyze the origin of bipolar disorder from a genetic perspective, with emphasison miRNAs encoding genes. The response of patients to drugs such as lithium and valproate, and a series of data from different experimental approaches show that in some cases of bipolar disorder, genes encoding miRNAs might be involved. These non coding RNAs elements regulate gene expression and could participate in the development of diseases such as cancer, immune system disorders, heart disease and different mental and neurological disorders. Conclusions. Because the administration of mood stabilizers has been found to modify the expression of some miRNAs, this paper suggests that knowledge of the functions of these regulators could help to diagnose some cases of bipolar disorder identifying molecular markers in plasma. This methodology would also allow applying new strategy founded on effective therapeutic targets for personalized treatment (AU)
Subject(s)
Humans , MicroRNAs/analysis , Bipolar Disorder/genetics , Valproic Acid/therapeutic use , Lithium/therapeutic use , Genetic MarkersABSTRACT
Neurexins and neuroligins are cell adhesion molecules present in excitatory and inhibitory synapses, and they are required for correct neuron network function. These proteins are found at the presynaptic and postsynaptic membranes. Studies in mice indicate that neurexins and neurologins have an essential role in synaptic transmission. Recent reports have shown that altered neuronal connections during the development of the human nervous system could constitute the basis of the etiology of numerous cases of autism spectrum disorders. Caenorhabditis elegans could be used as an experimental tool to facilitate the study of the functioning of synaptic components, because of its simplicity for laboratory experimentation, and given that its nervous system and synaptic wiring has been fully characterized. In C. elegans nrx-1 and nlg-1 genes are orthologous to human NRXN1 and NLGN1 genes which encode alpha-neurexin-1 and neuroligin-1 proteins, respectively. In humans and nematodes, the organization of neurexins and neuroligins is similar in respect to functional domains. The head of the nematode contains the amphid, a sensory organ of the nematode, which mediates responses to different stimuli, including osmotic strength. The amphid is made of 12 sensory bipolar neurons with ciliated dendrites and one presynaptic terminal axon. Two of these neurons, named ASHR and ASHL are particularly important in osmotic sensory function, detecting water-soluble repellents with high osmotic strength. The dendrites of these two neurons lengthen to the tip of the mouth and the axons extend to the nerve ring, where they make synaptic connections with other neurons determining the behavioral response. To evaluate the implications of neurexin and neuroligin in high osmotic strength avoidance, we show the different response of C. elegans mutants defective in nrx-1 and nlg-1 genes, using a method based on a 4M fructose ring. The behavioral phenotypes were confirmed using specific RNAi clones. In C. elegans, the dsRNA required to trigger RNAi can be administered by feeding. The delivery of dsRNA through food induces the RNAi interference of the gene of interest thus allowing the identification of genetic components and network pathways.
Subject(s)
Autistic Disorder/genetics , Caenorhabditis elegans/physiology , Neural Cell Adhesion Molecules/genetics , Synapses/physiology , Animals , Caenorhabditis elegans/genetics , Humans , Osmotic PressureABSTRACT
DNA damages can be removed by different repair processes, but lesions sometimes remain and block DNA replication. Specialized polymerases are needed to overcome this difficulty. In Arabidopsis, AtPOLH and AtREV1 genes code for two polymerases that are involved in replication of damaged DNA. Alternative splicing was detected in both genes. Complementation analysis of the alternative splicing forms in Saccharomycescerevisiae showed that the C-terminal extreme of AtPOLH protein is essential for recovering wild type UV viability in Rad30 deficient strain. None of the alternative AtREV1 forms recovered the yeast wild type phenotype of Rev1 deficient yeast strains after UV light irradiation or methyl methane sulphonate exposition, suggesting that AtREV1 may not be able to interact with other yeast specific proteins needed for DNA translesion synthesis.
ABSTRACT
Plants are continually exposed to external and internal DNA-damaging agents. Although lesions can be removed by different repair processes, damages often remain in the DNA during replication. Synthesis of template damages requires the replacement of replicative enzymes by translesion synthesis polymerases, which are able to perform DNA synthesis opposite specific lesions. These proteins, in contrast to replicative polymerases, operate at low processivity and fidelity. DNA polymerase eta and Rev 1 are two proteins found in eukaryotes that are involved in translesion DNA synthesis. In Arabidopsis, DNA polymerase eta and Rev 1 are encoded by AtPOLH and AtREV1 genes, respectively. Transgenic plants over-expressing AtPOLH showed increased resistance to ultraviolet light. Only plants with moderate AtREV1 over-expression were obtained, indicating that this enzyme could be toxic at high levels. Transgenic plants that over-expressed or disrupted AtREV1 showed reduced germination percentage, but the former exhibited a higher stem growth rate than the wild type during development.
Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/enzymology , Arabidopsis/genetics , DNA-Directed DNA Polymerase/metabolism , Gene Expression Regulation, Plant/physiology , Nucleotidyltransferases/metabolism , Ultraviolet Rays , Arabidopsis/radiation effects , Arabidopsis Proteins/genetics , DNA-Directed DNA Polymerase/genetics , Gene Expression Regulation, Developmental/physiology , Germination , Nucleotidyltransferases/genetics , Plant Stems/growth & development , Plants, Genetically ModifiedABSTRACT
Cellular DNA is continually exposed to a large variety of external and internal DNA-damaging agents. Although lesions can be removed by different repair processes, damages often remain in the DNA during replication, and specialized DNA polymerases are needed to perform translesion synthesis past damaged sites. These enzymes, in contrast to replicative polymerases, operate at low processivity and fidelity. DNA polymerase eta and Rev 1 are two proteins found in eukaryotes that are involved in translesion replication past specific DNA damages. In Arabidopsis, DNA polymerase eta and Rev 1 are encoded by AtPOLH and AtREV1 genes, respectively. The beta-glucuronidase gene product under the control of AtPOLH and AtREV1 gene promoters was used to determine their expression in different tissues. The GUS assay showed a ubiquitous expression of the reporter gene in all tissues and during the complete life cycle. In addition, quantitative real-time RT-PCR confirmed the ubiquitous expression of AtPOLH and AtREV1, and showed that the average expression of AtREV1 was approximately five times higher than AtPOLH. Transcription of both genes did not increase in the presence of visible light or after UV irradiation.
Subject(s)
Arabidopsis/enzymology , Arabidopsis/genetics , DNA-Directed DNA Polymerase/genetics , Gene Expression Regulation, Plant , Arabidopsis Proteins/genetics , DNA Primers , Genome, Plant , Glucuronidase/genetics , Nucleotidyltransferases/genetics , Plants, Genetically Modified/enzymology , Plants, Genetically Modified/genetics , Reverse Transcriptase Polymerase Chain Reaction , Site-Specific DNA-Methyltransferase (Adenine-Specific)/geneticsABSTRACT
DNA polymerase eta belongs to the Y-family of DNA polymerases, enzymes that are able to synthesize past template lesions that block replication fork progression. This polymerase accurately bypasses UV-associated cis-syn cyclobutane thymine dimers in vitro and therefore may contributes to resistance against sunlight in vivo, both ameliorating survival and decreasing the level of mutagenesis. We cloned and sequenced a cDNA from Arabidopsis thaliana which encodes a protein containing several sequence motifs characteristics of Pol eta homologues, including a highly conserved sequence reported to be present in the active site of the Y-family DNA polymerases. The gene, named AtPOLH, contains 14 exons and 13 introns and is expressed in different plant tissues. A strain from Saccharomyces cerevisiae, deficient in Pol eta activity, was transformed with a yeast expression plasmid containing the AtPOLH cDNA. The rate of survival to UV irradiation in the transformed mutant increased to similar values of the wild type yeast strain, showing that AtPOLH encodes a functional protein. In addition, when AtPOLH is expressed in Escherichia coli, a change in the mutational spectra is detected when bacteria are irradiated with UV light. This observation might indicate that AtPOLH could compete with DNA polymerase V and then bypass cyclobutane pyrimidine dimers incorporating two adenylates.
Subject(s)
Arabidopsis/genetics , DNA-Directed DNA Polymerase/genetics , Escherichia coli/genetics , Mutation/radiation effects , Ultraviolet Rays , Amino Acid Sequence , Animals , Arabidopsis/enzymology , Bacteriophage T4/growth & development , DNA, Complementary/genetics , Dose-Response Relationship, Radiation , Escherichia coli/radiation effects , Escherichia coli/virology , Genes, Suppressor/radiation effects , Genetic Complementation Test/methods , Humans , Molecular Sequence Data , Phylogeny , Plant Proteins/genetics , Pyrimidine Dimers/radiation effects , RNA, Transfer, Gln/genetics , Saccharomyces cerevisiae/genetics , Saccharomyces cerevisiae/growth & development , Saccharomyces cerevisiae/radiation effects , Sequence Homology, Amino AcidABSTRACT
Survival of irradiated spores from Fusarium oxysporum with ultraviolet radiation (UV) was increased following exposition to visible light, indicating that this phytopathogenic fungus has a mechanism of photoreactivation able to counteract the lethal effects of UV. A genomic sequence containing the complete photolyase gene (phr1) from F. oxysporum was isolated by heterologous hybridisation with the Neurospora crassa photolyase gene. The F. oxysporum phr1 cDNA was isolated and expressed in a photolyase deficient Escherichia coli strain. The complementation of the photoreactivation deficiency of this E. coli mutant by phr1 cDNA demonstrated that the photolyase gene from F. oxysporum encodes a functional protein. The F. oxysporum PHR1 protein has a domain characteristic of photolyases from fungi (Trichoderma harziaium, N. crassa, Magnaporthe grisea, Saccharomyces cerevisiae) to bacteria (E. coli), and clusters in the photolyases phylogenetic tree with fungal photolyases. The F. oxysporum phr1 gene was inducible by visible light. The phr1 expression was also detected in presence of alpha-tomatine, a glycoalkaloid from tomato damaging cell membranes, suggesting that phr1 is induced by this cellular stress.