Penkov Dmitry, Palazzolo Martina, Mondino Anna, Blasi Francesco
Molecular Genetics Unit, San Raffaele Scientific Institute and Università Vita Salute San Raffaele, Milan, Italy.
PLoS One. 2008 Jun 18;3(6):e2424. doi: 10.1371/journal.pone.0002424.
Prep1 and Pbx2 are the main homeodomain DNA-binding proteins of the TALE (three amino acid loop extension) family expressed in the thymus. We previously reported reduced Pbx2 expression and defective thymocyte maturation in Prep1 hypomorphic mice. To further investigate the role of this homeodomain DNA-binding protein in T cell development, we generated transgenic mice expressing the N-terminal fragment of Pbx1 (Pbx1NT) under the control of the Lck proximal promoter.
Pbx1NT causes Prep1 cytosolic sequestration, abolishes Prep1-dependent DNA-binding activity and results in reduced Pbx2 expression in developing thymocytes. Transgenic thymi reveal increased numbers of CD4(-) CD8(-) CD44(-) (DN3 and DN4) thymocytes, due to a higher frequency of DN2 and DN4 Pbx1NT thymocytes in the S phase. Transgenic thymocytes however do not accumulate at later stages, as revealed by a normal representation of CD4/CD8 double positive and single positive thymocytes, due to a higher rate of apoptotic cell death of DN4 Pbx1NT thymocytes.
The results obtained by genetic (Prep1 hypomorphic) and functional (Pbx1NT transgenic) inactivation of Prep1 support nonredundant roles for this homeodomain protein during different stages of T cell development.
Prep1和Pbx2是在胸腺中表达的TALE（三氨基酸环延伸）家族主要的同源结构域DNA结合蛋白。我们之前报道了Prep1低表达小鼠中Pbx2表达降低以及胸腺细胞成熟缺陷。为了进一步研究这种同源结构域DNA结合蛋白在T细胞发育中的作用，我们构建了在Lck近端启动子控制下表达Pbx1 N端片段（Pbx1NT）的转基因小鼠。
Pbx1NT导致Prep1在胞质中隔离，消除Prep1依赖的DNA结合活性，并导致发育中的胸腺细胞中Pbx2表达降低。转基因胸腺中CD4(-)CD8(-)CD44(-)（DN3和DN4）胸腺细胞数量增加，这是由于S期DN2和DN4 Pbx1NT胸腺细胞频率更高。然而，转基因胸腺细胞在后期并未积累，如CD4/CD8双阳性和单阳性胸腺细胞的正常比例所示，这是由于DN4 Pbx1NT胸腺细胞凋亡率更高。
通过Prep1的基因（Prep1低表达）和功能（Pbx1NT转基因）失活获得的结果支持了这种同源结构域蛋白在T细胞发育不同阶段的非冗余作用。
