Umezawa Yoshihiro, Akiyama Hiroki, Okada Keigo, Ishida Shinya, Nogami Ayako, Oshikawa Gaku, Kurosu Tetsuya, Miura Osamu
From the Department of Hematology, Graduate School of Medical and Dental Sciences Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyoku, Tokyo 113-8519, Japan.
From the Department of Hematology, Graduate School of Medical and Dental Sciences Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyoku, Tokyo 113-8519, Japan
J Biol Chem. 2017 Dec 1;292(48):19639-19655. doi: 10.1074/jbc.M117.779603. Epub 2017 Oct 3.
Platelet endothelial cell adhesion molecule 1 (PECAM-1) is a cell adhesion protein involved in the regulation of cell adhesion and migration. Interestingly, several PECAM-1-deficient hematopoietic cells exhibit impaired chemotactic responses to stromal cell-derived factor 1 (SDF-1), a chemokine essential for B lymphopoiesis and bone marrow myelopoiesis. However, whether PECAM-1 is involved in SDF-1-regulated chemotaxis is unknown. We report here that SDF-1 induces tyrosine phosphorylation of PECAM-1 at its immunoreceptor tyrosine-based inhibition motifs in several hematopoietic cell lines via the Src family kinase Lyn, Bruton's tyrosine kinase, and JAK2 and that inhibition of these kinases reduced chemotaxis. Overexpression and knockdown of PECAM-1 enhanced and down-regulated, respectively, SDF-1-induced Gα-dependent activation of the PI3K/Akt/mTORC1 pathway and small GTPase Rap1 in hematopoietic 32Dcl3 cells, and these changes in activation correlated with chemotaxis. Furthermore, pharmacological or genetic inhibition of the PI3K/Akt/mTORC1 pathway or Rap1, respectively, revealed that these pathways are independently activated and required for SDF-1-induced chemotaxis. When coexpressed in 293T cells, PECAM-1 physically associated with the SDF-1 receptor CXCR4. Moreover, PECAM-1 overexpression and knockdown reduced and enhanced SDF-1-induced endocytosis of CXCR4, respectively. Furthermore, when expressed in 32Dcl3 cells, an endocytosis-defective CXCR4 mutant, CXCR4-S324A/S325A, could activate the PI3K/Akt/mTORC1 pathway as well as Rap1 and induce chemotaxis in a manner similar to PECAM-1 overexpression. These findings suggest that PECAM-1 enhances SDF-1-induced chemotaxis by augmenting and prolonging activation of the PI3K/Akt/mTORC1 pathway and Rap1 and that PECAM-1, at least partly, exerts its activity by inhibiting SDF-1-induced internalization of CXCR4.
血小板内皮细胞黏附分子1（PECAM-1）是一种参与细胞黏附和迁移调节的细胞黏附蛋白。有趣的是，几种缺乏PECAM-1的造血细胞对基质细胞衍生因子1（SDF-1）的趋化反应受损，SDF-1是B淋巴细胞生成和骨髓髓细胞生成所必需的趋化因子。然而，PECAM-1是否参与SDF-1调节的趋化作用尚不清楚。我们在此报告，SDF-1通过Src家族激酶Lyn、布鲁顿酪氨酸激酶和JAK2在几种造血细胞系中诱导PECAM-1基于免疫受体酪氨酸的抑制基序发生酪氨酸磷酸化，并且抑制这些激酶会降低趋化作用。PECAM-1的过表达和敲低分别增强和下调了造血32Dcl3细胞中SDF-1诱导的PI3K/Akt/mTORC1途径和小GTP酶Rap1的Gα依赖性激活，并且这些激活变化与趋化作用相关。此外，分别对PI3K/Akt/mTORC1途径或Rap1进行药理学或基因抑制表明，这些途径是独立激活的，并且是SDF-1诱导趋化作用所必需的。当在293T细胞中共表达时，PECAM-1与SDF-1受体CXCR4发生物理相互作用。此外，PECAM-1的过表达和敲低分别减少和增强了SDF-1诱导的CXCR4内吞作用。此外，当在内吞缺陷的CXCR4突变体CXCR4-S324A/S325A中在32Dcl3细胞中表达时，其可以激活PI3K/Akt/mTORC1途径以及Rap1，并以类似于PECAM-1过表达的方式诱导趋化作用。这些发现表明，PECAM-1通过增强和延长PI3K/Akt/mTORC1途径和Rap1的激活来增强SDF-1诱导的趋化作用，并且PECAM-1至少部分地通过抑制SDF-1诱导的CXCR4内化来发挥其活性。
