Sorting protein VPS33B regulates exosomal autocrine signaling to mediate hematopoiesis and leukemogenesis
Hao Gu, … , Guo-Qiang Chen, Junke Zheng
Hao Gu, … , Guo-Qiang Chen, Junke Zheng
Published October 31, 2016
Citation Information: J Clin Invest. 2016;126(12):4537-4553. https://doi.org/10.1172/JCI87105.
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Research Article Hematology

Sorting protein VPS33B regulates exosomal autocrine signaling to mediate hematopoiesis and leukemogenesis

Abstract

Certain secretory proteins are known to be critical for maintaining the stemness of stem cells through autocrine signaling. However, the processes underlying the biogenesis, maturation, and secretion of these proteins remain largely unknown. Here we demonstrate that many secretory proteins produced by hematopoietic stem cells (HSCs) undergo exosomal maturation and release that is controlled by vacuolar protein sorting protein 33b (VPS33B). Deletion of VPS33B in either mouse or human HSCs resulted in impaired exosome maturation and secretion as well as loss of stemness. Additionally, VPS33B deficiency led to a dramatic delay in leukemogenesis. Exosomes purified from either conditioned medium or human plasma could partially rescue the defects of HSCs and leukemia-initiating cells (LICs). VPS33B co-existed in exosomes with GDI2, VPS16B, FLOT1, and other known exosome markers. Mechanistically, VPS33B interacted with the GDI2/RAB11A/RAB27A pathway to regulate the trafficking of secretory proteins as exosomes. These findings reveal an essential role for VPS33B in exosome pathways in HSCs and LICs. Moreover, they shed light on the understanding of vesicle trafficking in other stem cells and on the development of improved strategies for cancer treatment.

Authors

Hao Gu, Chiqi Chen, Xiaoxin Hao, Conghui Wang, Xiaocui Zhang, Zhen Li, Hongfang Shao, Hongxiang Zeng, Zhuo Yu, Li Xie, Fangzhen Xia, Feifei Zhang, Xiaoye Liu, Yaping Zhang, Haishan Jiang, Jun Zhu, Jiangbo Wan, Chun Wang, Wei Weng, Jingjing Xie, Minfang Tao, Cheng Cheng Zhang, Junling Liu, Guo-Qiang Chen, Junke Zheng

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Figure 7

VPS33B supports mouse AML development.

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VPS33B supports mouse AML development. (A) Percentages of YFP+ leukemia ...
(A) Percentages of YFP+ leukemia cells in the peripheral blood of the recipients 4 weeks after primary transplantation (n = 5; **P < 0.01 using Student’s t test). (B) The representative flow cytometric analysis of Mac-1+c-Kit+ LICs in the BM of primary recipient mice is shown. (C) Quantification of the percentages of Mac-1+c-Kit+ cells in WT and VPS33B-null recipient mice (n = 5; ***P < 0.001 using Student’s t test). (D) Mice transplanted with MLL-AF9–infected VPS33B-null HSCs/progenitors had significantly extended survival upon primary transplantation compared with the survival of mice transplanted with control cells (n = 5; **P < 0.01 using log-rank test). (E) Secondary transplantation of 1 × 104 leukemia cells displayed a significantly delayed onset of leukemogenesis by VPS33B-null cells compared with control cells (n = 6; ***P < 0.001 using log-rank test). (F) Representative transmission electron micrographs of Mac-1+c-Kit+ WT and VPS33B-null LICs showing representative images of EVs, MVB I (blue arrows), and MVB II (green arrows). Scale bars: 2 μm; 500 nm (insets). (G) Quantification of organelles (EVs, MVB I, and MVB II) present in WT and VPS33B-null Mac-1+c-Kit+ LICs (n = 20; ***P < 0.001 using Student’s t test). (H) Representative images of Mac-1+c-Kit+ WT and VPS33B-null LICs cultured in basic medium (SCF+IL-3+IL-6) with or without ANGPTL2- and ANGPTL3-conditioned medium (Ctrl, A2, and A3) or their purified exosomes (CExo, A2Exo, and A3Exo), as well as human plasma–derived exosomes (hExo) for 6 days (n = 5). Experiments were conducted 3 times for validation.