Motto D G, Ross S E, Wu J, Hendricks-Taylor L R, Koretzky G A
Department of Physiology, University of Iowa College of Medicine, Iowa City 52242, USA.
J Exp Med. 1996 Apr 1;183(4):1937-43. doi: 10.1084/jem.183.4.1937.
Recently we described the molecular cloning of SLP-76, a hematopoietic cell-specific 76-kD protein that was first identified through its association with GST/Grb2 fusion proteins. The primary sequence of SLP-76 predicts a protein of 533 amino acids comprising an amino-terminal region with numerous potential tyrosine phosphorylation sites, a central region rich in proline residues, and a single carboxy-terminal SH2 domain. Here we demonstrate formally that Grb2 associates with unphosphorylated SLP-76 and map the Grb2 binding site on SLP-76 undergoes rapid tyrosine phosphorylation and associates with tyrosine phosphoproteins of 36, 62, and 130 kD. In vitro experiments show that the SH2 domain of SLP-76 associates with the 62- and 130-kD proteins and additionally with a serine/threonine kinase. Finally, we demonstrate that transient overexpression of SLP-76 results in dramatically enhanced TCR-mediated induction of nuclear factor of activated T cells (NFAT) and interleukin (IL) 2 promoter activity; and we provide evidence that a functional SLP-76 SH2 domain is required for this effect. Our data document the in vivo associations of SLP-76 with several proteins that potentially participate in T cell activation and implicate SLP-76 itself as an important molecule in TCR-mediated IL-2 production.
最近我们描述了SLP-76的分子克隆，它是一种造血细胞特异性的76-kD蛋白，最初是通过其与GST/Grb2融合蛋白的关联而被鉴定出来的。SLP-76的一级序列预测该蛋白由533个氨基酸组成，包括一个具有众多潜在酪氨酸磷酸化位点的氨基末端区域、一个富含脯氨酸残基的中央区域以及一个单一的羧基末端SH2结构域。在这里，我们正式证明Grb2与未磷酸化的SLP-76结合，并确定SLP-76上的Grb2结合位点会迅速发生酪氨酸磷酸化，并与36、62和130 kD的酪氨酸磷酸化蛋白结合。体外实验表明，SLP-76的SH2结构域与62-kD和130-kD蛋白结合，此外还与一种丝氨酸/苏氨酸激酶结合。最后，我们证明SLP-76的瞬时过表达导致TCR介导的活化T细胞核因子（NFAT）和白细胞介素（IL）2启动子活性显著增强；并且我们提供证据表明这种效应需要一个功能性的SLP-76 SH2结构域。我们的数据记录了SLP-76在体内与几种可能参与T细胞活化的蛋白的关联，并表明SLP-76本身是TCR介导的IL-2产生中的一个重要分子。
