PPIL2 is a target of the JAK2/STAT5 pathway and promotes myeloproliferation via degradation of p53
Pan Wang, Xu Han, Kehan Ren, Ermin Li, Honghao Bi, Inci Aydemir, Madina Sukhanova, Yijie Liu, Jing Yang, Peng Ji
Pan Wang, Xu Han, Kehan Ren, Ermin Li, Honghao Bi, Inci Aydemir, Madina Sukhanova, Yijie Liu, Jing Yang, Peng Ji
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Research Article Cell biology Hematology

PPIL2 is a target of the JAK2/STAT5 pathway and promotes myeloproliferation via degradation of p53

Abstract

The activated JAK2/STAT pathway is characteristic of myeloproliferative neoplasms (MPNs). The pleckstrin 2 (PLEK2) signalosome is downstream of the JAK2/STAT5 pathway and plays an important role in MPN development. The detailed molecular composition of this signalosome is unclear. Here, we reveal peptidylprolyl isomerase-like 2 (PPIL2) as a critical component of the complex in regulating human and murine erythropoiesis. PPIL2 was a direct target of STAT5 and was upregulated in patients with MPN and in a Jak2V617F MPN mouse model. Mechanistically, PPIL2 interacted with and catalyzed p53 polyubiquitination and proteasome-mediated degradation to promote cell growth. Ppil2 deficiency, or inhibition by cyclosporin A, led to a marked upregulation of p53 in vivo and ameliorated myeloproliferative phenotypes in Jak2V617F mice. Cyclosporin A also markedly reduced JAK2-mutated erythroid and myeloid proliferation in an induced pluripotent stem cell–derived human bone marrow organoid model. Our findings reveal PPIL2 as a critical component of the PLEK2 signalosome in driving MPN pathogenesis through negative regulation of p53, thus providing a target and opportunity for drug repurposing using cyclosporin A to treat MPNs.

Authors

Pan Wang, Xu Han, Kehan Ren, Ermin Li, Honghao Bi, Inci Aydemir, Madina Sukhanova, Yijie Liu, Jing Yang, Peng Ji

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Figure 1

PPIL2 is a component of the PLEK2 signalosome and regulates erythropoiesis in vitro.

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PPIL2 is a component of the PLEK2 signalosome and regulates erythropoies...
(A) Western blotting of indicated proteins after streptavidin pulldown of lysates from HEK293T cells transfected with control or BirA-PLEK2. (B) Western blotting of indicated proteins following anti-Flag IP of lysate from HEK293T cells transfected with control or Flag-PLEK2. (C) Relative PPIL2 mRNA levels at the indicated stages of cultured human (upper) and mouse (lower, T: hours) terminal erythroblasts. (D) Western blotting of PPIL2 on different days (designated as D) of EPO medium–cultured CD34+ cells. HSC70 was used as a loading control. (E) Western blotting of Ppil2 at different days of EPO medium–cultured mouse bone marrow lineage-negative cells. (F) Western blotting of PPIL2 on day 9 EPO medium–cultured CD34+ cells transduced with control or CRISPR-PPIL2 sgRNAs. (G) Proliferation assay of cells from F. (H) Flow cytometric analysis of CD235a (glycophorin A) on day 7 cultured cells from F. Representative images of the flow cytometric analyses of CD235a and CD71 on cells of different days are on the right. (I) Quantification of cell apoptosis using flow cytometry in cells from F. (J) Western blotting of Ppil2 in EPO medium–cultured mouse lineage-negative cells transduced with control or CRISPR-Ppil2 sgRNAs. (K) Proliferation assays of cells from J cultured for 48 hours. (L) Quantification of CD71+Ter119+ erythroid progenitors and CD71-Ter119+ mature erythrocytes by flow cytometric assays in cells from J. (M) Apoptosis assays of cells from J. Data are shown as the mean ± SEM. The comparison between 2 groups was evaluated by 2-tailed t test (G), and the comparison among multiple groups was evaluated with 1-way ANOVA (I, K, L, and M). *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001.