JAM3 maintains leukemia-initiating cell self-renewal through LRP5/AKT/β-catenin/CCND1 signaling
Yaping Zhang, … , Guo-Qiang Chen, Junke Zheng
Yaping Zhang, … , Guo-Qiang Chen, Junke Zheng
Published March 26, 2018
Citation Information: J Clin Invest. 2018;128(5):1737-1751. https://doi.org/10.1172/JCI93198.
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Research Article Hematology

JAM3 maintains leukemia-initiating cell self-renewal through LRP5/AKT/β-catenin/CCND1 signaling

Abstract

Leukemia-initiating cells (LICs) are responsible for the initiation, development, and relapse of leukemia. The identification of novel therapeutic LIC targets is critical to curing leukemia. In this report, we reveal that junctional adhesion molecule 3 (JAM3) is highly enriched in both mouse and human LICs. Leukemogenesis is almost completely abrogated upon Jam3 deletion during serial transplantations in an MLL-AF9–induced murine acute myeloid leukemia model. In contrast, Jam3 deletion does not affect the functions of mouse hematopoietic stem cells. Moreover, knockdown of JAM3 leads to a dramatic decrease in the proliferation of both human leukemia cell lines and primary LICs. JAM3 directly associates with LRP5 to activate the downstream PDK1/AKT pathway, followed by the downregulation of GSK3β and activation of β-catenin/CCND1 signaling, to maintain the self-renewal ability and cell cycle entry of LICs. Thus, JAM3 may serve as a functional LIC marker and play an important role in the maintenance of LIC stemness through unexpected LRP5/PDK1/AKT/GSK3β/β-catenin/CCND1 signaling pathways but not via its canonical role in cell junctions and migration. JAM3 may be an ideal therapeutic target for the eradication of LICs without influencing normal hematopoiesis.

Authors

Yaping Zhang, Fangzhen Xia, Xiaoye Liu, Zhuo Yu, Li Xie, Ligen Liu, Chiqi Chen, Haishan Jiang, Xiaoxin Hao, Xiaoxiao He, Feifei Zhang, Hao Gu, Jun Zhu, Haitao Bai, Cheng Cheng Zhang, Guo-Qiang Chen, Junke Zheng

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Figure 6

JAM3 is required for the proliferation of human leukemia cell lines.

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JAM3 is required for the proliferation of human leukemia cell lines. (A)...
(A) Representative flow cytometric analysis of JAM3 expression on different leukemia cell lines including Kasumi-1 (M2), HL-60 (M3), THP-1 (M5), U937 (M5), and MV4-11 (M5). (Isotype control, gray line). (B) FLAG-tagged JAM3 and shRNAs targeting JAM3 (sh997, sh1188, sh359, and sh731) were cotransfected into 293T cells (1:4 ratio), followed by immunoblotting for JAM3. (C) Representative images of JAM3-knockdown (sh731 and sh1188) THP-1 cells after 6 days in culture. (D–G) The numbers of THP-1, U937, Kasumi-1, and HL-60 cells were counted at the indicated days after infection with the JAM3-targeting sh731 or sh1188 or scrambled shRNA (n = 3; *P < 0.05, **P < 0.01, ***P < 0.001, 2-way ANOVA followed by Bonferroni’s post-test). (H) Representative images of colonies formed by the JAM3-knockdown (sh731 and sh1188) THP-1 cells after 9 days of culture in 1640 medium supplemented with 0.9% of methylcellulose and 10% of FBS. (I) Quantification of colony numbers in H (n = 3; **P < 0.01, ***P < 0.001, 1-way ANOVA followed by Bonferroni’s post-test). (J) Representative flow cytometric analysis of the cell cycle distribution in THP-1 cells targeted by sh731, sh1188, or scrambled shRNA, which was determined using BrdU incorporation. (K) Quantitative analysis of the cell cycle distribution results in J (n = 3; *P < 0.05, **P < 0.01, ***P < 0.001, 2-way ANOVA followed by Bonferroni’s post-test). Experiments were conducted 3–5 times for validation.