LOXL2-induced PEAR1 Ser891 phosphorylation suppresses CD44 degradation and promotes triple-negative breast cancer metastasis
Yingzhi Shen, … , Junling Liu, Xuemei Fan
Yingzhi Shen, … , Junling Liu, Xuemei Fan
Published August 15, 2024
Citation Information: J Clin Invest. 2024;134(16):e177357. https://doi.org/10.1172/JCI177357.
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Research Article Cell biology Oncology

LOXL2-induced PEAR1 Ser891 phosphorylation suppresses CD44 degradation and promotes triple-negative breast cancer metastasis

Abstract

CD44 is associated with a high risk of metastasis, recurrence, and drug resistance in various cancers. Here we report that platelet endothelial aggregation receptor 1 (PEAR1) is a CD44 chaperone protein that protected CD44 from endocytosis-mediated degradation and enhances cleavage of the CD44 intracellular domain (CD44-ICD). Furthermore, we found that lysyl oxidase–like protein 2 (LOXL2), an endogenous ligand of PEAR1, bound to the PEAR1-EMI domain and facilitated the interaction between PEAR1 and CD44 by inducing PEAR1 Ser891 phosphorylation in a manner that was independent of its enzyme activity. Levels of PEAR1 protein and PEAR1 phosphorylation at Ser891 were increased in patients with triple-negative breast cancer (TNBC), were positively correlated with expression of LOXL2 and CD44, and were negatively correlated with overall survival. The level of PEAR1 Ser891 phosphorylation was identified as the best independent prognostic factor in TNBC patients. The prognostic efficacy of the combination of PEAR1 phosphorylation at Ser891 and CD44 expression was superior to that of PEAR1 phosphorylation at Ser891 alone. Blocking the interaction between LOXL2 and PEAR1 with monoclonal antibodies significantly inhibited TNBC metastasis, representing a promising therapeutic strategy for TNBC.

Authors

Yingzhi Shen, Jie Yan, Lin Li, Huiyan Sun, Lin Zhang, Guoming Li, Xinxia Wang, Ruoyan Liu, Xuefeng Wu, Baosan Han, Xueqing Sun, Junling Liu, Xuemei Fan

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Figure 5

LOXL2 triggers PEAR1 phosphorylation.

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LOXL2 triggers PEAR1 phosphorylation. (A) The interaction between LOXL2 ...
(A) The interaction between LOXL2 and PEAR1 was detected by pulldown with Dynabeads in the supernatant of MDA-MB-231 cells, to which 15 μg exogenous PEAR1-ECD-his was added. (B) LOXL2 levels in the supernatants of various TNBC cell lines were quantified by ELISA (MDA-MB-231, red; MDA-MB-468, orange; SUM159, yellow). The complete culture medium served as a negative control (blue) (n = 2; mean ± SD). (C and D) Quantification of invasive cells in Transwell assay; and relative migration area in wound healing assay of MDA-MB-231 cells treated with 0, 5, or 10 ng/mL LOXL2 (n = 3; mean ± SEM). (E) Phosphorylated PEAR1, full-length CD44, and CD44-ICD levels were detected in MDA-MB-231 cells and SUM159 cells treated with 0, 5, 10, 20, or 50 ng/mL LOXL2. (F) Schematic showing the structures of full-length LOXL2 protein, the LOXL2-SRCR1-3 truncation, and the LOXL2-SRCR1-2 truncation. (G–I) Quantification of invasive cells in Transwell assay and relative migration area in wound healing assay (n = 3; mean ± SEM) and detection of PEAR1 phosphorylation levels with Western blotting in MDA-MB-231 cells treated with 0.2 nM full-length LOXL2 or its truncation construct. (J) Interaction of PEAR1-ECD–His with LOXL2 in the supernatant of MDA-MB-231 cells was inhibited by 20 μg/mL simtuzumab. (K and L) Quantification of invasive cells in Transwell assay and relative migration area in wound healing assay in MDA-MB-231 cells treated with 0, 5, 10, or 20 μg/mL simtuzumab (n = 3; mean ± SEM). (M) Levels of phosphorylated PEAR1, CD44, and CD44-ICD were determined in MDA-MB-231 cells treated with 0, 5, 10, or 20 μg/mL simtuzumab. One-way ANOVA followed by Dunnett’s test was used for C, D, G, H, K, and L. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. The Western blotting results are representative of 3 independent experiments.