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In JoVE (1)
Other Publications (6)
Articles by Manuel Ruiz-Rubio in JoVE
JoVE General
Osmotic Avoidance in Caenorhabditis elegans: Synaptic Function of Two Genes, Orthologues of Human NRXN1 and NLGN1, as Candidates for Autism
1Departamento de Genética, Facultad de Ciencias, Universidad de Córdoba, 2Instituto Maimónides de Investigación Biomédica de Córdoba (IMIBIC)
Neurexins and neuroligins are membrane-neuron adhesion proteins which perform essential roles in synaptic differentiation and transmission. Neuroligin deficient mutants of C. elegans are defective in detecting osmotic strength, but when they also contain a mutation in the gene coding neurexin, they recover the wild type phenotype.
Other articles by Manuel Ruiz-Rubio on PubMed
The Photolyase Gene from the Plant Pathogen Fusarium Oxysporum F. Sp. Lycopersici is Induced by Visible Light and Alpha-tomatine from Tomato Plant
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.
Analysis of UV-induced Mutation Spectra in Escherichia Coli by DNA Polymerase Eta from Arabidopsis Thaliana
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.
Ubiquitous Expression of Two Translesion Synthesis DNA Polymerase Genes in Arabidopsis
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.
Two Translesion Synthesis DNA Polymerase Genes, AtPOLH and AtREV1, Are Involved in Development and UV Light Resistance in Arabidopsis
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.
[MicroRNAs in Bipolar Disorder: Diagnostic and Therapeutic Applications]
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.
Caenorhabditis Elegans As an Experimental Tool for the Study of Complex Neurological Diseases: Parkinson's Disease, Alzheimer's Disease and Autism Spectrum Disorder
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 Aβ 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.
