Cullin5 deficiency promotes small-cell lung cancer metastasis by stabilizing integrin β1
Gaoxiang Zhao, … , Daming Gao, Hongbin Ji
Gaoxiang Zhao, … , Daming Gao, Hongbin Ji
Published January 28, 2019
Citation Information: J Clin Invest. 2019;129(3):972-987. https://doi.org/10.1172/JCI122779.
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Research Article Oncology

Cullin5 deficiency promotes small-cell lung cancer metastasis by stabilizing integrin β1

Abstract

Metastasis is the dominant cause of patient death in small-cell lung cancer (SCLC), and a better understanding of the molecular mechanisms underlying SCLC metastasis may potentially improve clinical treatment. Through genome-scale screening for key regulators of mouse Rb1–/– Trp53–/– SCLC metastasis using the pooled CRISPR/Cas9 library, we identified Cullin5 (CUL5) and suppressor of cytokine signaling 3 (SOCS3), two components of the Cullin-RING E3 ubiquitin ligase complex, as top candidates. Mechanistically, the deficiency of CUL5 or SOCS3 disrupted the functional formation of the E3 ligase complex and prevented the degradation of integrin β1, which stabilized integrin β1 and activated downstream focal adhesion kinase/SRC (FAK/SRC) signaling and eventually drove SCLC metastasis. Low expression levels of CUL5 and SOCS3 were significantly associated with high integrin β1 levels and poor prognosis in a large cohort of 128 clinical patients with SCLC. Moreover, the CUL5-deficient SCLCs were vulnerable to the treatment of the FDA-approved SRC inhibitor dasatinib. Collectively, this work identifies the essential role of CUL5- and SOCS3-mediated integrin β1 turnover in controlling SCLC metastasis, which might have therapeutic implications.

Authors

Gaoxiang Zhao, Liyan Gong, Dan Su, Yujuan Jin, Chenchen Guo, Meiting Yue, Shun Yao, Zhen Qin, Yi Ye, Ying Tang, Qibiao Wu, Jian Zhang, Binghai Cui, Qiurong Ding, Hsinyi Huang, Liang Hu, Yuting Chen, Peiyuan Zhang, Guohong Hu, Luonan Chen, Kwok-Kin Wong, Daming Gao, Hongbin Ji

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Figure 2

Genetic deletion of Cul5 enhances mouse SCLC metastasis.

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Genetic deletion of Cul5 enhances mouse SCLC metastasis. (A) Diagram of ...
(A) Diagram of validation of target genes from mGeCKOa screening using individual sgRNAs. (B) Cell lysates from RT cells infected with lentiviral sgCul5 or sgSocs3 vectors were subjected to IB analysis. GAPDH served as a loading control. (C and D) Representative images and H&E staining of the lung from mice transplanted with RT cells transduced with sgCtrl (n = 5), sgCul5 (n = 5), or sgSocs3 (n = 5). Metastatic tumors are indicated by blue arrows. Scale bars: 50 μm. (E and F) Representative photos and H&E staining of liver, chest wall, and LN tissue from the different mice transplanted with RT cells transduced with sgCul5 or sgSocs3. Blue arrows indicate metastatic tumors. Scale bar: 50 μm. (G) Colony formatiaon capability of RT cells transduced with sgCtrl or sgCul5 in a soft agar assay. Data represent the mean ± SEM (n = 3). ***P < 0.001, by Student’s t test. (H) Postsurgical metastasis was assessed 4 weeks after resection of comparably sized primary allograft tumors. The surgery was performed 3 weeks after transplantation for the sgCtrl group and 2 weeks after transplantation for the sgCul5 group. Representative H&E staining of lungs is shown. Blue arrow indicates metastasis. Scale bar: 50 μm. (I) Diagram of CTC analysis. The GFP+ RT cells were transduced with sgCtrl or sgCul5 and then transplanted into the flanks of nude mice. Quantification of circulating GFP+ cells per 100 μl blood was performed by flow cytometry 4 weeks (sgCtrl) or 3 weeks (sgCul5) after implantation. Dots represent the number of CTCs per 10,000 analyzed blood cells (CD45+) for each mouse. Data represent the mean ± SEM (n = 5 mice). *P < 0.05, by Student’s t test.