α‐Dystroglycan is involved in positive selection of thymocytes by participating in immunological synapse formation

Abstract

Alpha-dystroglycan has been proved to be involved in lymphocyte activation by participating in immunological synapse (IS) formation. Considering the existence of IS formation in thymic development, we questioned whether alpha-dystroglycan was expressed in thymus and influenced thymic development. In this study, we demonstrated that alpha-dystroglycan was expressed on fetal thymocytes, especially on double-positive (DP, CD4(+)CD8(+)) cells. Blocking alpha-dystroglycan by treatment of fetal thymus organ culture (FTOC) with anti-alpha-dystroglycan antibody IIH6C4 decreased the number of DP cells compared with nontreated or isotype antibody controls. Down-regulation of alpha-dystroglycan by retroviruses carrying antisense cDNA of dystroglycan in reaggregate thymus organ culture (RTOC) further confirmed these results. Enhanced apoptosis of DP cells was observed after blocking alpha-dystroglycan. Interestingly, we found that blocking alpha-dystroglycan reduced IS formation between DP cells and thymic epithelial cells. Furthermore, blocking alpha-dystroglycan up-regulated CD95/CD95L expression and reduced Bcl-2 expression on DP cells in the developing thymus. Finally, the increase in the apoptosis of DP cells was associated with a consequent decrease in the positive selection, as indicated by the reduction of both ERK phosphorylation in DP cells and single-positive (SP, CD4(+) or CD8(+)) cell outcome. Altogether, these results indicated that alpha-dystroglycan was involved in positive selection of thymocytes by participating in the IS formation.

Functional Glycosylation of Dystroglycan Is Crucial for Thymocyte Development in the Mouse

“…Functional α-DG was detectable on early DN thymocytes with highest expression levels on thymocytes at the DN3 and DN4 stages, a critical point of transition in T cell development during which thymocytes undergo β-selection. The pattern of α-DG expression in developing T cells in the thymus of adult mice reported here varies from the results of a recent study on the dynamics of α-DG expression in T cells from fetal thymus [27]. These differences may be due to the different developmental stage of the animals used in the studies, fetal versus adult.…”

“…Apoptosis in DP cells could not be addressed due to the inherently high turnover of this cell population. Our in vivo findings complement recent results employing re-aggregate thymus organ culture (RTOC) to demonstrate that ablation of DG in T cells in vitro enhanced the apoptosis rate in DP and SP thymocytes [27].…”

“…These differences may be due to the different developmental stage of the animals used in the studies, fetal versus adult. In addition, for their FACS staining, Gong et al [27] used a monoclonal antibody to the protein core: mAb 6C1 (mouse IgG) directed against amino acids 572–602 of human dystroglycan, whereas our studies employed mAb IIH6 (mouse IgM), which specifically recognized functionally glycosylated α-DG.…”

Small interference ITGA6 gene targeting in the human thymic epithelium differentially regulates the expression of immunological synapse-related genes

“…Interestingly however, it has been reported that a-distroglycan, a subunit of the distroglycan complex (which binds the ECM molecule laminin), plays a role in the intrathymic immunological synapse, since blocking this molecule through different strategies resulted in altered intrathymic T-cell development, as revealed in fetal mouse thymus organ cultures. 16 Components of ECM are critically involved in the formation of thymic microenvironments and T cell development, by establishing molecular bridges between thymocytes and microenvironmental cells. The major membrane receptors for adhesion to ECM proteins in metazoa are integrins, transmembrane heterodimers composed of a and b subunits, comprising 18 a and 8 b members, so far known to assemble into 24 distinct dimers.…”