RESUMO
Mutations in fukutin-related protein (FKRP) underlie a group of muscular dystrophies associated with the hypoglycosylation of α-dystroglycan (α-DG), a proportion of which show central nervous system involvement. Our original FKRP knock-down mouse (FKRP(KD)) replicated many of the characteristics seen in patients at the severe end of the dystroglycanopathy spectrum but died perinatally precluding its full phenotyping and use in testing potential therapies. We have now overcome this by crossing FKRP(KD) mice with those expressing Cre recombinase under the Sox1 promoter. Owing to our original targeting strategy, this has resulted in the restoration of Fkrp levels in the central nervous system but not the muscle, thereby generating a new model (FKRP(MD)) which develops a progressive muscular dystrophy resembling what is observed in limb girdle muscular dystrophy. Like-acetylglucosaminyltransferase (LARGE) is a bifunctional glycosyltransferase previously shown to hyperglycosylate α-DG. To investigate the therapeutic potential of LARGE up-regulation, we have now crossed the FKRP(MD) line with one overexpressing LARGE and show that, contrary to expectation, this results in a worsening of the muscle pathology implying that any future strategies based upon LARGE up-regulation require careful management.
Assuntos
Distroglicanas/metabolismo , N-Acetilglucosaminiltransferases/biossíntese , N-Acetilglucosaminiltransferases/genética , Proteínas/genética , Síndrome de Walker-Warburg/genética , Animais , Membrana Basal/metabolismo , Membrana Basal/patologia , Sistema Nervoso Central/metabolismo , Modelos Animais de Doenças , Glicosilação , Laminina/biossíntese , Camundongos , Camundongos Knockout , Músculo Esquelético/metabolismo , Músculo Esquelético/patologia , Mutação , Pentosiltransferases , Transferases , Regulação para Cima , Síndrome de Walker-Warburg/mortalidadeRESUMO
Negative regulation of B cell activation by cognate immune complexes plays an important homeostatic role in suppressing B cell hyperactivity and preventing consequent autoimmunity. Immune complexes co-ligate the BCR and FcγRIIB resulting in both growth arrest and apoptosis. We now show that such apoptotic signalling involves induction and activation of p53 and its target genes, the pro-apoptotic Bcl-2 family members, Bad and Bid, as well as nuclear export of p53. Collectively, these events result in destabilisation of the mitochondrial and lysosomal compartments with consequent activation and interplay of executioner caspases and endosomal-derived proteases. In addition, the upregulation of Fas and FasL with consequent activation of caspase 8-dependent death receptor signalling is required to facilitate efficient apoptosis of B cells. Consistent with this role for Fas death receptor signalling, apoptosis resulting from co-ligation of the BCR and FcγRIIB is defective in B cells from Fas-deficient MRL/MpJ-Fas(lpr) mice. As these mice develop spontaneous, immune complex-driven lupus-like glomerulonephritis, targeting this FcγRIIB-mediated apoptotic pathway may therefore have novel therapeutic implications for systemic autoimmune disease.
Assuntos
Complexo Antígeno-Anticorpo/imunologia , Linfócitos B/imunologia , Nefrite Lúpica/imunologia , Receptores de Antígenos de Linfócitos B/imunologia , Receptores de IgG/imunologia , Receptor fas/imunologia , Animais , Complexo Antígeno-Anticorpo/genética , Apoptose , Linfócitos B/metabolismo , Linfócitos B/patologia , Proteína Agonista de Morte Celular de Domínio Interatuante com BH3/genética , Proteína Agonista de Morte Celular de Domínio Interatuante com BH3/imunologia , Caspase 8/genética , Caspase 8/imunologia , Modelos Animais de Doenças , Proteína Ligante Fas/genética , Proteína Ligante Fas/imunologia , Regulação da Expressão Gênica/imunologia , Humanos , Nefrite Lúpica/genética , Nefrite Lúpica/patologia , Lisossomos/imunologia , Lisossomos/patologia , Masculino , Camundongos , Camundongos Endogâmicos MRL lpr , Camundongos Knockout , Mitocôndrias/imunologia , Mitocôndrias/patologia , Receptores de Antígenos de Linfócitos B/genética , Receptores de IgG/genética , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/imunologia , Proteína de Morte Celular Associada a bcl/genética , Proteína de Morte Celular Associada a bcl/imunologia , Receptor fas/genéticaRESUMO
Recognition by scavenger receptor cysteine-rich domains on membrane proteins regulates innate and adaptive immune responses. Two receptors expressed primarily on T cells, CD5 and CD6, are linked genetically and are structurally similar, both containing three scavenger receptor cysteine-rich domains in their extracellular regions. A specific cell surface interaction for CD5 has been difficult to define at the molecular level because of the susceptibility of CD5 protein to denaturation. By using soluble CD5 purified at neutral pH to preserve biological activity, we show that CD5 mediates species-specific homophilic interactions. CD5 domain 1 only is involved in the interaction. CD5 mAbs that have functional effects in humans, rats, and mice block homophilic binding. Ag-specific responses by mouse T cells in vitro were increased when engagement of human CD5 domain 1 was inhibited by mutation or by IgG or Fab fragment from a CD5 mAb. This showed that homophilic binding results in productive engagement. Enhancement of polyclonal immune responses of rat lymph node cells by a Fab fragment from a CD5 mAb shown to block homophilic interactions provided evidence that the extracellular region of CD5 regulates inhibition in normal cells. These biochemical and in vitro functional assays provide evidence that the extracellular region of CD5 regulates immunity through species-specific homophilic interactions.