RESUMEN
Several diseases are characterized by the presence of point mutations, which are amenable to molecular detection using a number of methods such as PCR. However, certain mutations are particularly difficult to detect due to factors such as low abundance and the presence of special (e.g., oligonucleotide repeat) sequences. The mutation 7A in the oligoA sequence of exon 7 of the gene encoding the La autoantigen is difficult to detect at the DNA level, and even at the RNA level, due to both its estimated low abundance and its differentiation from the wild-type 8A sequence. This article describes a technique in which amplification of the excess wild-type 8A La sequence is suppressed by a peptide nucleic acid (PNA) during a nested PCR step. Detection of the amplified 7A mutant form was then performed by simple electrophoresis following a final primer extension step with an infrared dye-labeled primer. This technique allowed us to detect the mutation in 3 of 7 individuals harboring serum immunoglobulin G (IgG) antibodies reactive with a neo-B cell epitope in the 7A mutant protein product. We propose that this method is a viable screening test for mutations in regions containing simple polynucleotide repeats.
Asunto(s)
Autoantígenos/genética , Epítopos de Linfocito B/genética , Mutación , Oligodesoxirribonucleótidos/genética , Secuencias Repetitivas de Ácidos Nucleicos/genética , Ribonucleoproteínas/genética , Análisis de Secuencia de ADN , Autoanticuerpos/genética , Autoanticuerpos/inmunología , Autoantígenos/inmunología , Secuencia de Bases , Epítopos de Linfocito B/inmunología , Femenino , Humanos , Inmunoglobulina G/genética , Inmunoglobulina G/inmunología , Masculino , Datos de Secuencia Molecular , Mutación/inmunología , Oligodesoxirribonucleótidos/inmunología , Ácidos Nucleicos de Péptidos , Reacción en Cadena de la Polimerasa , Secuencias Repetitivas de Ácidos Nucleicos/inmunología , Ribonucleoproteínas/inmunología , Síndrome de Sjögren/genética , Síndrome de Sjögren/inmunología , Antígeno SS-BRESUMEN
Every aging theory intended to find the ultimate cause of aging, however, so far all have failed to present the clue as to why aging occurs. Senescence is a complex process and the aging hypotheses concentrated on only one or very few elements of this process. The goal of the Unifying Theory of Aging is to organize these theories into an organic complex. Despite all our hopes aging is inevitable, the present information level of the human system allows only a temporary existence of an individual in time. During aging the system gradually drifts away from the developmentally differentiated state, which results in loosing step-by step the capability to cope with the deteriorating entropy force of the environment. Nevertheless, it is still possible to increase the mean life span of the human population but without altering the present information level of the human system aging still remains an incurable disease.
Asunto(s)
Envejecimiento , Evolución Biológica , División Celular , Humanos , Modelos BiológicosRESUMEN
In previous studies, we detected a frame shift mutation in the gene encoding the autoantigen La of a patient with systemic lupus erythematosus. The mutant La mRNA contains a premature termination codon. mRNAs that prematurely terminate translation should be eliminated by RNA quality control mechanisms. As we find Abs specific for the mutant La form in approximately 30% of sera from anti-La-positive patients, we expected that mutant La mRNAs circumvent RNA control and the expression of mutant La protein could become harmful. Indeed, real-time PCR, immunostaining, and immunoblotting data of mice transgenic for the mutant La form show that mutant La mRNAs are not repressed in these animals and are translated to mutant La protein. In addition to the mutant La protein, we detected a minor portion of native human La in the mutant La-transgenic mice. Therefore, ribosomal frame shifting may allow the mutant La mRNA to escape from RNA control. Interestingly, expression of the mutant La mRNA results in a lupus-like disease in the experimental mice. Consequently, escape of mutant La mRNA from RNA control can have two effects: it 1) results in the expression of an immunogenic (neo)epitope, and 2) predisposes to autoimmunity.
Asunto(s)
Autoanticuerpos/biosíntesis , Autoantígenos/genética , Epítopos/genética , Estabilidad del ARN/inmunología , ARN Mensajero/metabolismo , Ribonucleoproteínas/genética , Células 3T3 , Adulto , Secuencia de Aminoácidos , Animales , Autoanticuerpos/sangre , Autoantígenos/biosíntesis , Autoantígenos/inmunología , Codón sin Sentido , Epítopos/sangre , Epítopos/inmunología , Femenino , Humanos , Líquido Intracelular/inmunología , Líquido Intracelular/metabolismo , Lupus Eritematoso Sistémico/genética , Lupus Eritematoso Sistémico/inmunología , Masculino , Ratones , Ratones Transgénicos , Persona de Mediana Edad , Datos de Secuencia Molecular , Biosíntesis de Proteínas , Estabilidad del ARN/genética , ARN Mensajero/biosíntesis , ARN Mensajero/genética , Ribonucleoproteínas/biosíntesis , Ribonucleoproteínas/inmunología , Síndrome de Sjögren/genética , Síndrome de Sjögren/inmunología , Antígeno SS-BRESUMEN
Several types of diseases, among others autoimmune illnesses, could be coupled with the general processes of aging. The two-edged sword of immune defense is directed on one side against environmental attacks and on the other against the body itself. However, one has to make a difference between normal (physiological) clearance and autoimmune diseases, although both sides of autoimmunity are influenced by the general processes of senescence. Aging of the thymus seems to be one of the key elements in the etiology of autoimmunity, although other cell types and their aging also play a substantial role in this process. Spontaneous genetic instability, acquired genetic mutations due to aging and the age-related alterations in the information level of the body may together be important elements in the pathomechanism of both physiological autoimmunity and autoimmune diseases. Nevertheless, physiological autoimmunity seems to be directed mostly by natural factors (such as aging and apoptosis) but primary autoimmune diseases may be caused by genetic instability that is enhanced by aging as well.
Asunto(s)
Envejecimiento , Enfermedades Autoinmunes/etiología , Autoinmunidad , Envejecimiento/genética , Envejecimiento/fisiología , Enfermedades Autoinmunes/genética , Autoinmunidad/genética , Autoinmunidad/fisiología , HumanosRESUMEN
INTRODUCTION: The Epstein-Barr virus is a member of herpesvirus family. It plays an important role in the etiopathogenesis of Burkitt lymphoma, other B-cell non-Hodgkin lymphomas, nasopharyngeal carcinoma, X chromosome-linked lymphoproliferative disease, Hodgkin lymphomas and a part of T-cell lymphomas. It has been suggested that association of Epstein-Barr virus with lymphomas and its pathological significance in disease varies in different geographical areas. The aim of our study was to reveal the role of Epstein-Barr virus in B-cell non-Hodgkin lymphomas diagnosed in Hungary. The authors asked: A) What is the frequency of the presence of virus genome in the biopsy specimen from B-cell non-Hodgkin patients? B) Which types of virus latency can be observed? PATIENTS AND METHODS: Genomic DNA and EBER-specific RNA of Epstein-Barr virus in paraffin-embedded specimens were detected by polymerase chain reaction and in situ hybridisation, respectively. RESULTS: Out of 36 B-cell non-Hodgkin lymphoma cases, 16 (45%) were found to be positive for Epstein-Barr virus by the above methods. Expression of latency genes, nuclear antigene-2 and latent membrane protein-1 was studied by immunohistochemical technique. The 16 virus-positive non-Hodgkin lymphoma cases showed Epstein-Barr virus latency types II. (37%) or III. (63%). CONCLUSIONS: The authors data suggest that Epstein-Barr virus may be associated with the development of B-cell non-Hodgkin lymphomas in Hungarian patients.
