In vivo cleaved CDCP1 promotes early tumor dissemination via complexing with activated β1 integrin and induction of FAK/PI3K/Akt motility signaling.
In vivo cleaved CDCP1 promotes early tumor dissemination via complexing with activated β1 integrin and induction of FAK/PI3K/Akt motility signaling.
复制标题
作者：
纤溶酶样丝氨酸蛋白酶特异性切割跨膜分子，含CUB结构域的蛋白-1（CDCP 1），诱导由外向内的信号转导，促进自发转移的早期阶段，导致肿瘤细胞内渗，即细胞逃离原发肿瘤，间质入侵和跨内皮迁移。我们鉴定了活性β1整联蛋白作为膜保留70-kDa CDCP 1片段的生物化学和功能伴侣，该片段通过丝氨酸蛋白酶的蛋白水解切割从其全长135-kDa前体新产生。在细胞培养物和活体动物中，活性β1整联蛋白优先与功能活化的磷酸化70 kDa CDCP 1复合。β1整合素70-kDa与CDCP 1片段的复合物诱导细胞内磷酸化信号传导，涉及粘着斑激酶-1（FAK）和PI 3激酶（PI 3 K）依赖性Akt激活。因此，在CDCP 1被丝氨酸蛋白酶加工的条件下，特异性抑制剂对FAK/PI 3 K活性的抑制以及β1整联蛋白的短发夹RNA下调显着减少了FAK/Akt磷酸化，这表明FAK/PI 3 K/Akt途径在与β1整联蛋白复合的切割CDCP 1的下游运行。此外，这种复合物依赖性信号传导与高水平的肿瘤细胞浸润和扩散呈正相关。相应地，通过独特的切割阻断性单克隆抗体10-D 7或通过用抑肽酶抑制纤溶酶样丝氨酸蛋白酶的蛋白水解活性来消除体内CDCP 1切割，阻止了β1整联蛋白/CDCP 1复合和下游FAK/Akt信号传导，同时显著降低了基质侵袭和自发转移。因此，β1整联蛋白似乎充当运动调节伴侣，介导蛋白水解切割、膜保留的CDCP 1与FAK/PI 3 K/Akt途径成员之间的串扰。这种CDCP 1裂解诱导的信号级联反应构成了一种独特的机制，独立于细胞外基质重塑，由此蛋白水解裂解的CDCP 1通过β1整联蛋白伴侣作用调节肿瘤细胞的体内运动和转移。我们的研究结果表明，发生在β1整联蛋白/FAK/PI 3 K/Akt信号级联的顶点的CDCP 1切割可能代表CDCP 1阳性癌症的治疗靶点。
Specific cleavage of the transmembrane molecule, CUB domain-containing protein-1 (CDCP1), by plasmin-like serine proteases induces outside–in signal transduction that facilitates early stages of spontaneous metastasis leading to tumor cell intravasation, namely cell escape from the primary tumor, stromal invasion and transendothelial migration. We identified active β1 integrin as a biochemical and functional partner of the membrane-retained 70-kDa CDCP1 fragment, newly generated from its full-length 135-kDa precursor though proteolytic cleavage by serine proteases. Both in cell cultures and in live animals, active β1 integrin complexed preferentially with functionally activated, phosphorylated 70-kDa CDCP1. Complexing of β1 integrin the 70-kDa with CDCP1 fragment induced intracellular phosphorylation signaling, involving focal adhesion kinase-1 (FAK) and PI3 kinase (PI3K)-dependent Akt activation. Thus, inhibition of FAK/PI3K activities by specific inhibitors as well as short-hairpin RNA downregulation of β1 integrin significantly reduced FAK/Akt phosphorylation under conditions where CDCP1 was processed by serine proteases, indicating that FAK/PI3K/Akt pathway operates downstream of cleaved CDCP1 complexed with β1 integrin. Furthermore, this complex-dependent signaling correlated positively with high levels of tumor cell intravasation and dissemination. Correspondingly, abrogation in vivo of CDCP1 cleavage either by unique cleavage-blocking monoclonal antibody 10-D7 or by inhibition of proteolytic activity of plasmin-like serine proteases with aprotinin prevented β1 integrin/CDCP1 complexing and downstream FAK/Akt signaling concomitant with significant reduction of stromal invasion and spontaneous metastasis. Therefore, β1 integrin appears to serve as a motility-regulating partner mediating cross-talk between proteolytically cleaved, membrane-retained CDCP1 and members of FAK/PI3K/Akt pathway. This CDCP1 cleavage-induced signaling cascade constitutes a unique mechanism, independent of extracellular matrix remodeling, whereby a proteolytically cleaved CDCP1 regulates in vivo locomotion and metastasis of tumor cells through β1 integrin partnering. Our findings indicate that CDCP1 cleavage, occurring at the apex of a β1 integrin/FAK/PI3K/Akt signaling cascade, may represent a therapeutic target for CDCP1-positive cancers.