CBL is a RING type E3 ubiquitin ligase that functions as a negative regulator of tyrosine kinase signaling and loss of CBL E3 function is implicated in several forms of leukemia. The Src-like adaptor proteins (SLAP/SLAP2) bind to CBL and are required for CBL-dependent downregulation of antigen receptor, cytokine receptor, and receptor tyrosine kinase signaling. Despite the established role of SLAP/SLAP2 in regulating CBL activity, the nature of the interaction and the mechanisms involved are not known. To understand the molecular basis of the interaction between SLAP/SLAP2 and CBL, we solved the crystal structure of CBL tyrosine kinase binding domain (TKBD) in complex with SLAP2. The carboxy-terminal region of SLAP2 adopts an α-helical structure which binds in a cleft between the 4H, EF-hand, and SH2 domains of the TKBD. This SLAP2 binding site is remote from the canonical TKBD phospho-tyrosine peptide binding site but overlaps with a region important for stabilizing CBL in its autoinhibited conformation. In addition, binding of SLAP2 to CBL in vitro activates the ubiquitin ligase function of autoinhibited CBL. Disruption of the CBL/SLAP2 interface through mutagenesis demonstrated a role for this protein-protein interaction in regulation of CBL E3 ligase activity in cells. Our results reveal that SLAP2 binding to a regulatory cleft of the TKBD provides an alternative mechanism for activation of CBL ubiquitin ligase function.

中文翻译：

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SLAP2衔接子结合破坏c-CBL自抑制以激活泛素连接酶功能

CBL是一种RING型E3泛素连接酶，可作为酪氨酸激酶信号的负调节剂，而CBL E3功能的丧失与多种形式的白血病有关。Src样衔接蛋白（SLAP / SLAP2）与CBL结合，是CBL依赖性抗原受体，细胞因子受体和受体酪氨酸激酶信号转导下调所必需的。尽管SLAP / SLAP2在调节CBL活性中起着确定的作用，但相互作用的性质和涉及的机制尚不清楚。为了了解SLAP / SLAP2与CBL之间相互作用的分子基础，我们解决了与SLAP2复合的CBL酪氨酸激酶结合域（TKBD）的晶体结构。SLAP2的羧基末端区域采用α-螺旋结构，该结构在TKBD的4H，EF-hand和SH2结构域之间的缝隙中结合。该SLAP2结合位点远离经典的TKBD磷酸酪氨酸肽结合位点，但与稳定CBL的自抑制构象重要区域重叠。另外，SLAP2与CBL的体外结合激活了自抑制CBL的泛素连接酶功能。通过诱变破坏CBL / SLAP2接口证明了这种蛋白相互作用在调节细胞中CBL E3连接酶活性中的作用。我们的结果表明，SLAP2与TKBD的调节性裂口结合为CBL泛素连接酶功能的激活提供了另一种机制。体外SLAP2与CBL的结合激活了自抑制CBL的泛素连接酶功能。通过诱变破坏CBL / SLAP2接口证明了这种蛋白相互作用在调节细胞中CBL E3连接酶活性中的作用。我们的结果表明，SLAP2与TKBD的调节性裂口结合为CBL泛素连接酶功能的激活提供了另一种机制。体外SLAP2与CBL的结合激活了自抑制CBL的泛素连接酶功能。通过诱变破坏CBL / SLAP2接口证明了这种蛋白相互作用在调节细胞中CBL E3连接酶活性中的作用。我们的结果表明，SLAP2与TKBD的调节性裂口结合为CBL泛素连接酶功能的激活提供了另一种机制。