Integrins regulate survival of pre-B-ALL cells through differential IAP and caspase-7 ubiquitination and degradation.
Integrins regulate survival of pre-B-ALL cells through differential IAP and caspase-7 ubiquitination and degradation.
复制标题
整合素通过差异 IAP 和 caspase-7 泛素化和降解来调节前 B-ALL 细胞的存活。
DOI：
10.1038/sj.leu.2403298
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发表时间：
2004
期刊：
影响因子：
11.4
通讯作者：
Freedman,AS
中科院分区：
文献类型：
--
作者：
Astier,AL;Svoboda,M;Hinds,E;DeBeaumont,R;Munoz,O;Freedman,AS
整合素和它们的配体之间的相互作用拯救正常和肿瘤性B细胞免于凋亡。特别地，已经观察到前体B白血病细胞的粘附依赖性细胞存活。虽然从大多数前ALL患者分离的细胞在单独培养基中培养时发生凋亡，但在骨髓基质细胞存在下，凋亡被阻止。此外，ALL细胞存活的延长取决于基质细胞和白血病细胞之间的细胞-细胞接触信号，并直接涉及b1整联蛋白。1.恶性肿瘤细胞上的α 4 β 1单克隆抗体或基质细胞上的VCAM-1单克隆抗体的粘附破坏导致白血病细胞的凋亡。同样，粘附调节化疗期间B系白血病细胞的存活。2-4重要的是，儿童ALL中白血病细胞的整合素依赖性存活与疾病的侵袭性有关。5因此，确定整合素介导的存活机制对于潜在的临床影响是重要的。已经提出了若干机制。整合素调节前B ALL细胞中的bcl-2家族成员。3在AML细胞系中，α 4 β 1整合素与纤连蛋白（Fn）的粘附导致PI-3 K/AKT的激活，并上调bcl-2，从而赋予抗失巢凋亡和耐药性。此外，在鼠模型中，抗VLA-4抗体和阿糖胞苷的组合赋予100%的存活率，而单独化疗仅具有很小的影响。6在pre-B ALL中，我们最近发现b1整合素刺激抑制caspase-3和caspase-7的诱导，并增加凋亡抑制因子（IAP）家族成员XIAP和Survivin的表达。4 IAP蛋白结合并抑制半胱天冬酶。大多数IAP蛋白在C末端具有环指基序，其赋予内在的泛素连接酶活性，并在凋亡信号后促进底物非依赖性泛素化。7另一方面，IAP还可以泛素化几种底物，如caspase-3和caspase-7，保护细胞免于凋亡。[8]因此，凋亡受促凋亡分子和抗凋亡分子数量之间的精细平衡控制，一旦泛素化，则受蛋白酶体降解的调节。在本研究中，我们研究了b1整合素在这些蛋白的泛素化和降解中的作用。REH（pre-B ALL）细胞在无血清培养基中培养，Fn是b1整合素的配体，或多聚赖氨酸（PLL）作为阴性对照粘附细胞。首先通过膜联蛋白-PI染色分析细胞，并通过流式细胞术分析，以评估整联蛋白刺激的存活效果。如图1a所示和先前报道的，4整合素刺激拯救了前B细胞免于凋亡。然后进行实验以比较两个IAP家族成员XIAP和IAP 2在整合素介导的人肿瘤细胞存活中的诱导作用。
The interactions between integrins and their ligands rescue normal and neoplastic B cells from apoptosis. In particular, adhesion-dependent cell survival has been observed for precursor B leukemia cells. Although cells isolated from most patients with pre-ALL undergo apoptosis when cultured in media alone, apoptosis is prevented in the presence of bone marrow stromal cells. Moreover, the prolongation of ALL cell survival is dependent on cell–cell contact signals between stromal and leukemic cells, with direct involvement of b1 integrins. 1 Disruption of the adhesion with mAbs against a4b1 on the malignant cell or VCAM-1 on the stromal cell leads to apoptosis of the leukemia cells. Similarly, adhesion regulates survival of B-lineage leukemia cells during chemotherapy. 2–4 Importantly, the integrin-dependent survival of leukemic cells in childhood ALL has been linked to the aggressiveness of the disease. 5 It is therefore important to determine the mechanisms of integrin-mediated survival for potential clinical impact. Several mechanisms have been proposed. Integrins modulate bcl-2 family members in pre-B ALL cells. 3 In AML cell lines, a4b1 integrin adhesion to fibronectin (Fn) leads to the activation of PI-3K/AKT with upregulation of bcl-2, which confers resistance to anoikis and drug resistance. Moreover, in a murine model, the combination of anti-VLA-4 antibodies and cytosine arabinoside conferred 100% survival, whereas chemotherapy alone had only a small impact. 6 In pre-B ALL, we recently demonstrated that b1 integrin stimulation inhibited caspase-3 and-7 induction, and increased the expression of XIAP and Survivin, two inhibitors of apoptosis (IAP) family members. 4 IAP proteins bind to and inhibit caspases. Most IAP proteins possess a Ring-finger motif at the C terminus, which confers an intrinsic ubiquitin ligase activity and promotes substrate-independent ubiquitination, upon apoptotic signals. 7 On the other hand, IAP can also ubiquitinate several substrates, such as caspase-3 and-7, protecting the cells from apoptosis. 8 Thus, apoptosis is controlled by a fine balance between the amounts of pro-and antiapoptotic molecules, regulated by degradation in the proteasome once ubiquitinated. In this study, we investigated the role of b1 integrins in the ubiquitination and degradation of these proteins.REH (pre-B ALL) cells were cultured in a serum-free medium on Fn, a ligand for b1 integrin, or Poly-L-Lysine (PLL) used as a negative control to adhere the cells. Cells were first analyzed by Annexin-PI staining and analyzed by flow cytometry in order to assess the survival effect of integrin stimulation. As shown in Figure 1a and as previously reported, 4 integrin stimulation rescued pre-B cells from apoptosis. Experiments were then performed to compare the induction of XIAP and IAP2, two IAP family members, in integrin-mediated survival of human