SUMOylation promotes extracellular vesicle–mediated transmission of lncRNA ELNAT1 and lymph node metastasis in bladder cancer
Changhao Chen, Hanhao Zheng, Yuming Luo, Yao Kong, Mingjie An, Yuting Li, Wang He, Bowen Gao, Yue Zhao, Hao Huang, Jian Huang, Tianxin Lin
Changhao Chen, Hanhao Zheng, Yuming Luo, Yao Kong, Mingjie An, Yuting Li, Wang He, Bowen Gao, Yue Zhao, Hao Huang, Jian Huang, Tianxin Lin
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Research Article Cell biology Oncology

SUMOylation promotes extracellular vesicle–mediated transmission of lncRNA ELNAT1 and lymph node metastasis in bladder cancer

Abstract

Small ubiquitin-like modifier (SUMO) binding (termed SUMOylation) emerged as the inducer for the sorting of bioactive molecules into extracellular vesicles (EVs), triggering lymphangiogenesis and further driving tumor lymph node (LN) metastasis, but the precise mechanisms remain largely unclear. Here, we show that bladder cancer (BCa) cell–secreted EVs mediated intercellular communication with human lymphatic endothelial cells (HLECs) through transmission of the long noncoding RNA ELNAT1 and promoted lymphangiogenesis and LN metastasis in a SUMOylation-dependent manner in both cultured BCa cell lines and mouse models. Mechanistically, ELNAT1 induced UBC9 overexpression to catalyze the SUMOylation of hnRNPA1 at the lysine 113 residue, which mediated recognition of ELNAT1 by the endosomal sorting complex required for transport (ESCRT) and facilitated its packaging into EVs. EV-mediated ELNAT1 was specifically transmitted into HLECs and epigenetically activated SOX18 transcription to induce lymphangiogenesis. Importantly, blocking the SUMOylation of tumor cells by downregulating UBC9 expression markedly reduced lymphatic metastasis in EV-mediated, ELNAT1-treated BCa in vivo. Clinically, EV-mediated ELNAT1 was correlated with LN metastasis and a poor prognosis for patients with BCa. These findings highlight a molecular mechanism whereby the EV-mediated ELNAT1/UBC9/SOX18 regulatory axis promotes lymphangiogenesis and LN metastasis in BCa in a SUMOylation-dependent manner and implicate ELNAT1 as an attractive therapeutic target for LN metastatic BCa.

Authors

Changhao Chen, Hanhao Zheng, Yuming Luo, Yao Kong, Mingjie An, Yuting Li, Wang He, Bowen Gao, Yue Zhao, Hao Huang, Jian Huang, Tianxin Lin

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Figure 4

ELNAT1 directly interacts with hnRNPA1.

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ELNAT1 directly interacts with hnRNPA1. (A) RNA pull-down assay of ELNA...
(A) RNA pull-down assay of ELNAT1 in UM-UC-3 cells. (B) MS analysis of the proteins from the RNA pull-down assay. (C and D) RNA pull-down with nuclear extract (C) and Western blotting with purified recombinant hnRNPA1 (D) were performed to evaluate the interaction between ELNAT1 and hnRNPA1. (E) Immunofluorescence was performed to assess the colocalization of ELNAT1 and hnRNPA1 in UM-UC-3 and T24 cells. Scale bars: 5 μm. (F) RIP assay using anti-hnRNPA1 to assess the enrichment of ELNAT1 by hnRNPA1. IgG was used as a negative control; U1 was used as a nonspecific control. A 2-tailed Student’s t test was performed to determine statistical significance. (G and H) RNA pull-down assays using serial deletions of ELNAT1 were performed to evaluate the regions required for the binding of ELNAT1 and hnRNPA1. (I) Prediction for the stem-loop structures of hnRNPA1-binding sites in ELNAT1. (J) RIP assay after deletion of 610–680 nt of ELNAT1 in UM-UC-3 cells. A 2-tailed Student’s t test was used to assess statistical significance. Error bars show the SD of 3 independent experiments. **P < 0.01.