Research on the B cell development using mutant mice generated through gene targeting.

免疫球蛋白(Ig)MU链作为与Vpre-B和lambda5蛋白相关的受体表达于前B细胞表面。为了解前B细胞受体的生理功能，利用基因打靶技术在胚胎干细胞中分别打乱了mU链基因和lambda5基因的膜外显子，并将其导入小鼠生殖系，产生了突变小鼠MUMT和lambda5T。在杂合子MUMT小鼠的B细胞中，MUMT等位基因并不排除其他重链等位基因。纯合子木乃伊小鼠缺乏B细胞，B细胞发育停滞在大的前B细胞阶段或之前。同样，在纯合子lambda5T小鼠中，B细胞的发育受到抑制，尽管这种抑制是有泄漏的，这表明存在不依赖lambda5的B细胞发育途径。在这些突变体的未成熟细胞中，L链基因重排的发生程度与正常小鼠相同，这表明L链基因重排的启动不依赖于前B细胞受体。HS1蛋白是Src样蛋白酪氨酸激酶的主要底物之一，在B细胞表面IgM(SIGM)交联后立即磷酸化。在T细胞受体(TCR)的刺激下，HS1也被酪氨酸磷酸化。通过基因打靶获得的HS1基因缺陷小鼠，表现出胸腺负选择能力的降低以及sIgM的交联性诱导B细胞的凋亡细胞死亡。这一结果清楚地表明，HS1是一个参与了由抗原受体触发的细胞凋亡信号转导途径的分子。

Immunoglobuline(Ig) mu chain is expressed on the surface of pre-B cells as a receptor associated with Vpre-B and lambda5 proteins. To understand the physiological function of the pre-B cell receptor, the membrane exon of the mu chain gene and lambda5 gene were disrupted separately in embryonic stem cells by gene targeting, and through the transmission of these mutations into the mouse germ lines, the mutant mice, muMT and lambda5T, were generated. In the B cells from heterozygous muMT mice, muMT allele does not exclude the other heavy chain allele. Homozygous muMT mice lack B cells and B cell development is arrested at or before the stage of large pre-B cells. Similarly, B cell development is inhibited in homozygous lambda5T mice, although this inhibition is leaky, suggesting the presence of lambda5-independent pathway of the B cell development. L chain gene rearrangement occurs in the immature cells in these mutants to the same extent as those cells in normal mice, indicating the initiation of the L chain gene rearrangement is independent of the pre-B cell receptor.HS1 protein is one of the major substrates of Src-like protein tyrosine kinases and phosphorylated immediately after crosslinking of surface IgM(sIgM)on B cells. The HS1 is also tyrosine-phosphorylated upon the stimulation of T cell receptors(TCR). The HS1 deficient mice, which were generated through gene targeting, showed the impaired ability for thymic negative selection as well as the impaired apoptotic cell death of the B cells induced by the crosslinking of sIgM.This results clearly demonstrated that HS1 is a molecule involved in the signal transduction pathway toward apoptosis that is triggered the antigen receptors.