Pao L I, Cambier J C
Department of Pediatrics, National Jewish Center for Immunology and Respiratory Medicine, Denver, CO 80206, USA.
J Immunol. 1997 Mar 15;158(6):2663-9.
B cell Ag receptor (BCR) signaling occurs via tyrosine phosphorylation of CD79a and CD79b ITAMs, leading to recruitment and activation of Lyn and Syk tyrosine kinases and subsequent downstream events. CD45 expression is required for BCR triggering of certain of these downstream events, such as calcium mobilization and p21ras activation. However, the site in the BCR signaling cascade at which CD45 impinges is poorly defined. To address this question, we have studied CD45 function in the CD45-deficient (CD45-) and CD45-reconstituted (CD45+) J558L mu m3 plasmacytoma. In both CD45+ and CD45- cells, Ag stimulation led to CD79a and CD79b tyrosine phosphorylation as well as Syk tyrosine phosphorylation, recruitment to the receptors, and activation. In contrast to CD45+ cells, Lyn exhibited high basal tyrosine phosphorylation in the CD45- cells and was not further phosphorylated upon Ag stimulation. Mapping studies indicated that the observed constitutive phosphorylation of Lyn reflects phosphorylation of its C-terminal tyrosine, Y508, at high stoichiometry. Constitutively Y508-phosphorylated Lyn was neither recruited to the BCR nor activated upon Ag stimulation. Moreover, CD79a-ITAM phosphopeptides failed to bind Lyn from the CD45- cells. Thus, Y508 phosphorylation of Lyn occurs in the absence of cellular CD45 expression and appears to render the kinase unable to associate with the phosphorylated receptor complex via its Src homology 2 domain and to participate in signal propagation. Surprisingly, in view of previous findings implicating Src family kinases in ITAM phosphorylation, the data indicate that Ag-induced CD79a and CD79b tyrosine phosphorylation and Syk recruitment and activation can occur in the absence of CD45 expression and, hence, Src-family kinase activation.
B细胞抗原受体（BCR）信号传导通过CD79a和CD79b免疫受体酪氨酸激活基序（ITAM）的酪氨酸磷酸化发生，导致Lyn和Syk酪氨酸激酶的募集和激活以及随后的下游事件。CD45的表达是BCR触发某些这些下游事件所必需的，如钙动员和p21ras激活。然而，CD45在BCR信号级联中发挥作用的位点尚不清楚。为了解决这个问题，我们研究了CD45缺陷（CD45-）和CD45重组（CD45+）的J558Lμm3浆细胞瘤中CD45的功能。在CD45+和CD45-细胞中，抗原刺激均导致CD79a和CD79b酪氨酸磷酸化以及Syk酪氨酸磷酸化、募集到受体并激活。与CD45+细胞相反，Lyn在CD45-细胞中表现出高基础酪氨酸磷酸化，并且在抗原刺激后不再进一步磷酸化。定位研究表明，观察到的Lyn组成型磷酸化反映了其C末端酪氨酸Y508的高化学计量比磷酸化。组成型Y508磷酸化的Lyn既不被募集到BCR也不在抗原刺激后被激活。此外，CD79a-ITAM磷酸肽不能结合来自CD45-细胞的Lyn。因此，Lyn的Y508磷酸化在细胞缺乏CD45表达的情况下发生，并且似乎使该激酶无法通过其Src同源2结构域与磷酸化的受体复合物结合并参与信号传导。令人惊讶的是，鉴于先前的研究结果表明Src家族激酶参与ITAM磷酸化，数据表明在缺乏CD45表达以及因此Src家族激酶激活的情况下，抗原诱导的CD79a和CD79b酪氨酸磷酸化以及Syk募集和激活仍可发生。
