Regulation of thymocyte β-selection, development and positive selection by glycogen synthase kinase-3

Glycogen synthase kinase-3 (GSK-3) is a ubiquitously expressed serine/threonine kinase, that exists as two isoforms in mammals, GSK-3α and GSK-3β, that are key downstream mediators of the phosphatidylinositol 3’ kinase, Wnt, Notch and other pathways. Here, we report that simultaneous inactivation of both GSK-3α and GSK-3β during early thymocyte ontogeny has profound effects on both β-selection and positive selection, key checkpoints essential to producing functionally mature αβ T cells. Conditional GSK-3α/β knockout animals (LckCre+ GSK-3αβfl/fl) possessed pre-double positive (pre-DP) thymocytes (CD4−CD8−CD117−CD25−) with compromised TCRβ chain expression along with elevated levels of β-catenin and reduced Notch activity. β-selection was impaired allowing pre-DP thymocytes to differentiate to DP thymocytes (CD4+CD8+) while bypassing strict requirements for productive TCRβ chain rearrangements and functional expression. Also impaired was the requisite pre-TCR and Notch-mediated expansion that normally precedes differentiation to the DP stage. Consequently, LckCre+ GSK-3αβfl/fl mice initially generated fewer DP thymocytes that expressed significantly reduced levels of mature TCR. The aberrant DP thymocytes expressed high levels of the pro-survival Bcl-2 family member Mcl-1, failed positive selection and accumulated as CD4hiCD8lo positive selection intermediates resulting in loss of both mature CD4 and CD8 lineages. LckCre+ GSK-3αβfl/fl mice succumbed to oligoclonal peripheral lymphomas with high penetrance. These data reveal essential roles for GSK-3 in several checkpoints of early T cell development.

糖原合成酶激酶 - 3（GSK - 3）是一种广泛表达的丝氨酸/苏氨酸激酶，在哺乳动物中以两种异构体形式存在，即GSK - 3α和GSK - 3β，它们是磷脂酰肌醇3’激酶、Wnt、Notch和其他通路的关键下游介质。在此，我们报道在早期胸腺细胞发生过程中同时使GSK - 3α和GSK - 3β失活对β选择和阳性选择都具有深远影响，这两个选择是产生功能成熟的αβ T细胞所必需的关键检查点。条件性GSK - 3α/β基因敲除动物（LckCre⁺ GSK - 3αβfl/fl）具有前双阳性（pre - DP）胸腺细胞（CD4⁻CD8⁻CD117⁻CD25⁻），其TCRβ链表达受损，同时β - 连环蛋白水平升高，Notch活性降低。β选择受损，使得前DP胸腺细胞能够分化为DP胸腺细胞（CD4⁺CD8⁺），同时绕过了对有功能的TCRβ链重排和功能性表达的严格要求。同样受损的还有通常在分化为DP阶段之前所需的前TCR和Notch介导的扩增。因此，LckCre⁺ GSK - 3αβfl/fl小鼠最初产生的DP胸腺细胞较少，且这些细胞表达的成熟TCR水平显著降低。异常的DP胸腺细胞高表达促存活的Bcl - 2家族成员Mcl - 1，阳性选择失败，并积累为CD4hiCD8lo阳性选择中间体，导致成熟的CD4和CD8谱系均缺失。LckCre⁺ GSK - 3αβfl/fl小鼠死于高外显率的寡克隆外周淋巴瘤。这些数据揭示了GSK - 3在早期T细胞发育的几个检查点中的重要作用。