Nuclear PFKP promotes CXCR4-dependent infiltration by T cell acute lymphoblastic leukemia
Xueliang Gao, … , Xue-Zhong Yu, Haizhen Wang
Xueliang Gao, … , Xue-Zhong Yu, Haizhen Wang
Published July 13, 2021
Citation Information: J Clin Invest. 2021;131(16):e143119. https://doi.org/10.1172/JCI143119.
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Research Article Cell biology Oncology Article has an altmetric score of 12

Nuclear PFKP promotes CXCR4-dependent infiltration by T cell acute lymphoblastic leukemia

Abstract

PFKP (phosphofructokinase, platelet), the major isoform of PFK1 expressed in T cell acute lymphoblastic leukemia (T-ALL), is predominantly expressed in the cytoplasm to carry out its glycolytic function. Our study showed that PFKP is a nucleocytoplasmic shuttling protein with functional nuclear export and nuclear localization sequences (NLSs). Cyclin D3/CDK6 facilitated PFKP nuclear translocation by dimerization and by exposing the NLS of PFKP to induce the interaction between PFKP and importin 9. Nuclear PFKP stimulated the expression of C-X-C chemokine receptor type 4 (CXCR4), a chemokine receptor regulating leukemia homing/infiltration, to promote T-ALL cell invasion, which depended on the activity of c-Myc. In vivo experiments showed that nuclear PFKP promoted leukemia homing/infiltration into the bone marrow, spleen, and liver, which could be blocked with CXCR4 antagonists. Immunohistochemical staining of tissues from a clinically well-annotated cohort of T cell lymphoma/leukemia patients showed nuclear PFKP localization in invasive cancers, but not in nonmalignant T lymph node or reactive hyperplasia. The presence of nuclear PFKP in these specimens correlated with poor survival in patients with T cell malignancy, suggesting the potential utility of nuclear PFKP as a diagnostic marker.

Authors

Xueliang Gao, Shenghui Qin, Yongxia Wu, Chen Chu, Baishan Jiang, Roger H. Johnson, Dong Kuang, Jie Zhang, Xi Wang, Anand Mehta, Kenneth D. Tew, Gustavo W. Leone, Xue-Zhong Yu, Haizhen Wang

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Figure 4

Nuclear PFKP upregulates CXCR4 expression.

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Nuclear PFKP upregulates CXCR4 expression. (A) Expression levels of cell...
(A) Expression levels of cell surface proteins (upper: CXCR4, by flow cytometry; lower: HMMR, FGFR3 and VEGFA, by immunoblotting [IB]) functioning in cell invasion/migration were measured in leukemia cells treated with palbociclib (Palbo) (1 μM, 48 hours for DND41 and KOPTK1; 1 μM, 24 hours for MOLT4 and MOLT16). (B) CXCR4 expression measured by flow cytometry was decreased in KOPTK1 cells treated with the CDK6 degrader (CDK6D; 10 μM BSJ-03-123, 48 hours), or in which CDK6/cyclin D3 was knocked down with shRNA. IB shows the efficiency of the CDK6 degrader. Actin and vinculin were loading controls. (C) Correlation analysis of CCLE transcriptomic data reveals that expression of CCND3 (encoding cyclin D3) and CDK6, but not of CCND1 (encoding cyclin D1) or CDK4, is correlated with that of CXCR4 in all cancer cell lines (1,019 cell lines in total) and in hematopoietic cell lines (176 cell lines in total). Purple color represents positive correlation, and blue color represents negative correlation. Larger size and darker color of the dots represent stronger correlation between two genes. Pearson’s correlation coefficients for each pair of genes were calculated using R software. (D) CXCR4 expression measured by flow cytometry was increased in cells expressing NLS-PFKP but not PFKP-RXL (upper). IB shows HMMR expression was not changed in cells expressing NLS-PFKP or PFKP-RXL mutant (lower). (E) Flow cytometry analysis of CXCR4 expressed on the surface of KOPTK1 cells (upper), in which the median value of the reading was used for quantification (lower). CXCR4 expression was normalized to KOPTK1 cells transfected with empty vector. (F) CDK6 knockdown in leukemia cells expressing NLS-PFKP did not significantly affect nuclear PFKP accumulation (upper). IB of whole cell lysate (WCL) showed that the expression of NLS-PFKP or CXCR4 was not affected by CDK6 knockdown (lower). (G) Fold changes in mRNA levels of CXCR4 in cells expressing NLS-PFKP compared with WT-PFKP measured using qRT-PCR. n = 3 (A, B, and D–G). Data represent mean ± SEM. **P < 0.01, ***P < 0.001, ****P < 0.0001 by 2-tailed Student’s t test (A) or 1-way ANOVA (B, D, E, and G).