BIOSS Centre for Biological Signalling Studies, Department of Molecular Immunology, Institute of Biology III, Faculty of Biology, University of Freiburg, Freiburg, Germany.
Max Planck Institute of Immunobiology and Epigenetics, Freiburg, Germany.
Biol Chem. 2019 Mar 26;400(4):555-563. doi: 10.1515/hsz-2018-0308.
Upon activation of the B cell antigen receptor (BCR), the spleen tyrosine kinase (Syk) and the Src family kinase Lyn phosphorylate tyrosines of the immunoreceptor tyrosine-based activation motif (ITAM) of Igα and Igβ which further serve as binding sites for the SH2 domains of these kinases. Using a synthetic biology approach, we dissect the roles of different ITAM residues of Igα in Syk activation. We found that a leucine to glycine mutation at the Y+3 position after the first ITAM tyrosine prevents Syk binding and activation. However, a pre-activated Syk can still phosphorylate this tyrosine in trans. Our data show that the formation of a Syk/ITAM initiation complex and trans-ITAM phosphorylation is crucial for BCR signal amplification. In contrast, the interaction of Lyn with the first ITAM tyrosine is not altered by the leucine to glycine mutation. In addition, our study suggests that an ITAM-bound Syk phosphorylates the non-ITAM tyrosine Y204 of Igα only in cis. Collectively, our reconstitution experiments suggest a model whereby first trans-phosphorylation amplifies the BCR signal and subsequently cis-phosphorylation couples the receptor to downstream signaling elements.
当 B 细胞抗原受体 (BCR) 被激活后，脾脏酪氨酸激酶 (Syk) 和Src 家族激酶 Lyn 磷酸化 Igα 和 Igβ 的免疫受体酪氨酸基激活基序 (ITAM) 上的酪氨酸残基，这些酪氨酸残基进一步成为这些激酶的 SH2 结构域的结合位点。我们使用合成生物学方法，解析了 Igα 的不同 ITAM 残基在 Syk 激活中的作用。我们发现，在第一个 ITAM 酪氨酸之后的 Y+3 位置的亮氨酸突变为甘氨酸会阻止 Syk 的结合和激活。然而，预先激活的 Syk 仍然可以在转位时磷酸化这个酪氨酸。我们的数据表明，Syk/ITAM 起始复合物的形成和跨 ITAM 磷酸化对于 BCR 信号放大至关重要。相比之下，Lyn 与第一个 ITAM 酪氨酸的相互作用不受亮氨酸到甘氨酸突变的影响。此外，我们的研究表明，一个结合 ITAM 的 Syk 仅在顺式中磷酸化 Igα 的非 ITAM 酪氨酸 Y204。总的来说，我们的重建实验提出了一个模型，即首先是跨磷酸化放大了 BCR 信号，随后是顺式磷酸化将受体与下游信号元件偶联。
