RESUMO
Recent thymic emigrants (RTEs) must undergo phenotypic and functional maturation to become long-lived mature naive T cells. In CD4-cre NKAP conditional knockout mice, NKAP-deficient RTEs fail to complete T cell maturation. In this study, we demonstrate that NKAP-deficient immature RTEs do not undergo apoptosis, but are eliminated by complement. C3, C4, and C1q are bound to NKAP-deficient peripheral T cells, demonstrating activation of the classical arm of the complement pathway. As thymocytes mature and exit to the periphery, they increase sialic acid incorporation into cell surface glycans. This is essential to peripheral lymphocyte survival, as stripping sialic acid with neuraminidase leads to the binding of natural IgM and complement fixation. NKAP-deficient T cells have a defect in sialylation on cell surface glycans, leading to IgM recruitment. We demonstrate that the defect in sialylation is due to aberrant α2,8-linked sialylation, and the expression of three genes (ST8sia1, ST8sia4, and ST8sia6) that mediate α2,8 sialylation are downregulated in NKAP-defcient RTEs. The maturation of peripheral NKAP-deficient T cells is partially rescued in a C3-deficient environment. Thus, sialylation during T cell maturation is critical to protect immature RTEs from complement in the periphery.
Assuntos
Movimento Celular/imunologia , Proteínas do Sistema Complemento/imunologia , Subpopulações de Linfócitos T/imunologia , Timo/imunologia , Animais , Antígenos CD/metabolismo , Antígenos de Diferenciação de Linfócitos B/metabolismo , Apoptose/genética , Apoptose/imunologia , Antígenos CD55/genética , Antígenos CD55/metabolismo , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Movimento Celular/genética , Sobrevivência Celular/genética , Sobrevivência Celular/imunologia , Ativação do Complemento/imunologia , Complemento C3/deficiência , Complemento C3/genética , Complemento C3/imunologia , Proteínas do Sistema Complemento/metabolismo , Expressão Gênica , Imunoglobulina M/imunologia , Imunoglobulina M/metabolismo , Imunofenotipagem , Antígenos Comuns de Leucócito/genética , Antígenos Comuns de Leucócito/metabolismo , Camundongos , Camundongos Knockout , Ácido N-Acetilneuramínico/metabolismo , Fenótipo , Ligação Proteica/imunologia , Proteínas Repressoras/deficiência , Proteínas Repressoras/genética , Subpopulações de Linfócitos T/citologia , Subpopulações de Linfócitos T/metabolismo , Timo/metabolismoRESUMO
B-cell-biased lymphoid progenitors (BLPs) and Pre-pro B cells lie at a critical juncture between B cell specification and commitment. However, both of these populations are heterogenous, which hampers investigation into the molecular changes that occur as lymphoid progenitors commit to the B cell lineage. Here, we demonstrate that there are PDCA-1(+)Siglec H(+) plasmacytoid dendritic cells (pDCs) that co-purify with BLPs and Pre-pro B cells, which express little or no CD11c or Ly6C. Removal of PDCA-1(+) pDCs separates B cell progenitors that express high levels of a Rag1-GFP reporter from Rag1-GFP(low/neg) pDCs within the BLP and Pre-pro B populations. Analysis of Flt3-ligand knockout and IL-7Rα knockout mice revealed that there is a block in B cell development at the all-lymphoid progenitor (ALP) stage, as the majority of cells within the BLP or Pre-pro B gates were PDCA-1(+) pDCs. Thus, removal of PDCA-1(+) pDCs is critical for analysis of BLP and Pre-pro B cell populations. Analysis of B cell potential within the B220(+)CD19(-) fraction demonstrated that AA4.1(+)Ly6D(+)PDCA-1(-) Pre-pro B cells gave rise to CD19(+) B cells at high frequency, while PDCA-1(+) pDCs in this fraction did not. Interestingly, the presence of PDCA-1(+) pDCs within CLPs may help to explain the conflicting results regarding the origin of these cells.
