RESUMO
Patients with the autoimmune disease multiple sclerosis (MS) typically present with the clinically isolated syndromes (CIS) transverse myelitis (TM) or optic neuritis (ON). B-cell disturbances have been well documented in patients with MS and CIS patients with ON, but not in CIS patients with TM, despite the fact that these patients have the worst clinical outcome of all CIS types. The goal of this study was to characterize the B-cell populations and immunoglobulin genetics in TM patients. We found a unique expansion of CD27(high) plasmablasts in both the cerebrospinal fluid and periphery of TM patients that is not present in ON patients. Additionally, plasmablasts from TM patients show evidence for positive selection with increased somatic hypermutation accumulation in VH4(+) B cells and receptor editing that is not observed in ON patients. These characteristics unique to TM patients may impact disease severity and progression.
Assuntos
Linfócitos B/imunologia , Mielite Transversa/imunologia , Plasmócitos/imunologia , Hipermutação Somática de Imunoglobulina/imunologia , Membro 7 da Superfamília de Receptores de Fatores de Necrose Tumoral/imunologia , Adulto , Linfócitos B/citologia , Linfócitos B/metabolismo , Proliferação de Células , Células Cultivadas , Citometria de Fluxo , Rearranjo Gênico de Cadeia Pesada de Linfócito B/genética , Rearranjo Gênico de Cadeia Pesada de Linfócito B/imunologia , Humanos , Cadeias Pesadas de Imunoglobulinas/genética , Cadeias Pesadas de Imunoglobulinas/imunologia , Região Variável de Imunoglobulina/genética , Região Variável de Imunoglobulina/imunologia , Pessoa de Meia-Idade , Mielite Transversa/sangue , Mielite Transversa/líquido cefalorraquidiano , Plasmócitos/citologia , Plasmócitos/metabolismo , Reação em Cadeia da Polimerase , Receptores de Antígenos de Linfócitos B/imunologia , Receptores de Antígenos de Linfócitos B/metabolismo , Hipermutação Somática de Imunoglobulina/genética , Membro 7 da Superfamília de Receptores de Fatores de Necrose Tumoral/metabolismo , Adulto JovemRESUMO
T cell receptor genes are assembled in developing T lymphocytes from discrete V, D, and J genes by a site-specific somatic rearrangement mechanism. A flanking recombination signal, composed of a conserved heptamer and a semiconserved nonamer separated by 12 or 23 variable nucleotides, targets the activity of the rearrangement machinery to the adjoining V, D, and J genes. Following the rearrangement of V, D, or J genes, their respective recombination signals are ligated together. Although these signal joints are allegedly invariant, created by the head-to-head abuttal of the heptamers, some do exhibit junctional diversity. Recombination signals were initially identified by comparison and alignment of germ-line sequences with the sequence of rearranged genes. However, their overall low level of sequence conservation makes their characterization solely from sequence data difficult. Recently, computational analysis unraveled correlations between nucleotides at several positions scattered within the spacer and recombination activity, so that it is now possible to identify putative recombination signals and determine and predict their recombination efficiency. In this paper, we analyzed the variability introduced in signal joints generated after rearrangement of the TRDD1 and TRDD2 genes in murine thymocytes. The recurrent presence of identical nucleotides inserted in these signal joints led us to reconsider the location and sequence of the TRDD1 recombination signal. By combining molecular characterization and computational analysis, we show that the functional TRDD1 recombination signal is shifted inside the putative coding sequence of the TRDD1 gene and, consequently, that this gene is shorter than indicated in the databases.
Assuntos
Rearranjo Gênico do Linfócito T/genética , Proteínas/genética , Animais , Sequência de Bases , Camundongos , Dados de Sequência Molecular , Recombinação Genética/genética , Análise de Sequência de DNA , Timo/imunologiaRESUMO
We have compared the microsequence specificity of mutations introduced during somatic hypermutation (SH) and those introduced meiotically during neutral evolution. We have minimized the effects of selection by studying nonproductive (hence unselected) Ig V region genes for somatic mutations and processed pseudogenes for meiotic mutations. We find that the two sets of patterns are very similar: the mutabilities of nucleotide triplets are positively correlated between the somatic and meiotic sets. The major differences that do exist fall into three distinct categories: 1) The mutability is sharply higher at CG dinucleotides under meiotic but not somatic mutation. 2) The complementary triplets AGC and GCT are much more mutable under somatic than under meiotic mutation. 3) Triplets of the form WAN (W = T or A) are uniformly more mutable under somatic than under meiotic mutation. Nevertheless, the relative mutabilities both within this set and within the SAN (S = G or C) triplets are highly correlated with those under meiotic mutation. We also find that the somatic triplet specificity is strongly symmetric under strand exchange for A/T triplets as well as for G/C triplets in spite of the strong predominance of A over T mutations. Thus, we suggest that somatic mutation has at least two distinct components: one that specifically targets AGC/GCT triplets and another that acts as true catalysis of meiotic mutation.
Assuntos
Análise Mutacional de DNA/métodos , Meiose/genética , Meiose/imunologia , Mutação , Sequência de Bases , Distribuição Binomial , Biologia Computacional/métodos , Biologia Computacional/estatística & dados numéricos , Análise Mutacional de DNA/estatística & dados numéricos , DNA Complementar/genética , Evolução Molecular , Mutação em Linhagem Germinativa , Humanos , Região Variável de Imunoglobulina/genética , Modelos Imunológicos , Pseudogenes/genética , Homologia de Sequência do Ácido Nucleico , Estatísticas não ParamétricasRESUMO
Somatic mutation is a fundamental component of acquired immunity. Although its molecular basis remains undetermined, the sequence specificity with which mutations are introduced has provided clues to the mechanism. We have analyzed data representing over 1700 unselected mutations in V gene introns and nonproductively rearranged V genes to identify the sequence specificity of the mutation spectrum-the distribution of resultant nucleotides. In other words, we sought to determine what effects the neighboring bases have on what a given base mutates "to." We find that both neighboring bases have a significant effect on the mutation spectrum. Their influences are complicated, but much of the effect can be characterized as enhancing homogeneity of the mutated DNA sequence. In contrast to what has been reported for the sequence specificity of the "targeting" mechanism, that of the spectrum is notably symmetric under complementation, indicating little if any strand bias. We compared the spectrum to that found previously for germline mutations as revealed by analyzing pseudogene sequences. We find that the influences of nearest neighbors are quite different in the two datasets. Altogether, our findings suggest that the mechanism of somatic hypermutation is complex, involving two or more stages: introduction of mis-pairs and their subsequent resolution, each with distinct sequence specificity and strand bias.
Assuntos
Mutação em Linhagem Germinativa/imunologia , Nucleotídeos/imunologia , Adenina/imunologia , Animais , Composição de Bases , Sequência de Bases , Análise Mutacional de DNA/métodos , Análise Mutacional de DNA/estatística & dados numéricos , Rearranjo Gênico , Humanos , Região Variável de Imunoglobulina/genética , Camundongos , Nucleotídeos/genética , Pseudogenes/imunologia , Timina/imunologiaRESUMO
Darwinian theory requires that mutations be produced in a nonanticipatory manner; it is nonetheless consistent to suggest that mutations that have repeatedly led to nonviable phenotypes would be introduced less frequently than others-if under appropriate genetic control. Immunoglobulins produced during infection acquire point mutations that are subsequently selected for improved binding to the eliciting antigen. We and others have speculated that an enhancement of mutability in the complementarity-determining regions (CDR; where mutations have a greater chance of being advantageous) and/or decrement of mutability in the framework regions (FR; where mutations are more likely to be lethal) may be accomplished by differential codon usage in concert with the known sequence specificity of the hypermutation mechanism. We have examined 115 nonproductively rearranged human Ig sequences. The mutation patterns in these unexpressed genes are unselected and therefore directly reflect inherent mutation biases. Using a chi2 test, we have shown that the number of mutations in the CDRs is significantly higher than the number of mutations found in the FRs, providing direct evidence for the hypothesis that mutations are preferentially targeted into the CDRs.
Assuntos
Genes de Imunoglobulinas , Região Variável de Imunoglobulina/genética , Mutação , Artrite/genética , Artrite/imunologia , Artrite Reumatoide/genética , Artrite Reumatoide/imunologia , DNA Complementar/genética , Evolução Molecular , Frequência do Gene , Humanos , Seleção Genética , Serina/genéticaRESUMO
Diversity in the antigen-binding receptors of the immune system has long been a primary interest of biologists. Recently it has been suggested that polymorphism in regulatory (noncoding) gene segments is of substantial importance as well. Here, we survey the level of variation in MHC class II gene promoters in man and mouse using extensive collections of published sequences together with unpublished sequences recently deposited by us in the EMBL gene bank using the Shannon entropy to quantify diversity. For comparison, we also apply our analysis to distantly related MHC class II promoters, as well as to class I promoters and to class II coding regions. We observe a high level of intraspecies variability, which in mouse but not in man is localized to a significant extent near the binding sites of transcription factors-sites that are conserved over longer evolutionary distances. This localization may both indicate and enhance heterozygote advantage, as the presence of two functionally different promoters would be expected to confer flexibility in the immune response.