CHK1 targets spleen tyrosine kinase (L) for proteolysis in hepatocellular carcinoma
Jian Hong, … , Yi-Xin Zeng, Tiebang Kang
Jian Hong, … , Yi-Xin Zeng, Tiebang Kang
Published May 15, 2012
Citation Information: J Clin Invest. 2012;122(6):2165-2175. https://doi.org/10.1172/JCI61380.
View: Text | PDF
Research Article Oncology

CHK1 targets spleen tyrosine kinase (L) for proteolysis in hepatocellular carcinoma

Abstract

Hepatocellular carcinoma (HCC) is one of the most prevalent malignancies resistant to current chemotherapies or radiotherapies, which makes it urgent to identify new therapeutic targets for HCC. In this study, we found that checkpoint kinase 1 (CHK1) was frequently overexpressed and correlated with poor clinical outcome in patients with HCC. We further showed that the CHK1 inhibitor GÖ6976 was capable of sensitizing HCC cells to cisplatin, indicating that CHK1 may have oncogenic function in HCC. We found that CHK1 phosphorylated the tumor suppressor spleen tyrosine kinase (L) (SYK[L]) and identified the phosphorylation site at Ser295. Furthermore, CHK1 phosphorylation of SYK(L) promoted its subsequent proteasomal degradation. Expression of a nonphosphorylated mutant of SYK(L) was more efficient at suppressing proliferation, colony formation, mobility, and tumor growth in HCC lines. Importantly, a strong inverse correlation between the expression levels of CHK1 and SYK(L) was observed in patients with HCC. Collectively, our data demonstrate that SYK(L) is a substrate of CHK1 in tumor cells and suggest that targeting the CHK1/SYK(L) pathway may be a promising strategy for treating HCC.

Authors

Jian Hong, Kaishun Hu, Yunfei Yuan, Yi Sang, Qiangui Bu, Guihua Chen, Longjun Yang, Binkui Li, Pinzhu Huang, Dongtai Chen, Yi Liang, Ruhua Zhang, Jingxuan Pan, Yi-Xin Zeng, Tiebang Kang

×

Figure 2

SYK(L) is regulated by CHK1 through phosphorylation on Ser295.

Options: View larger image (or click on image) Download as PowerPoint
SYK(L) is regulated by CHK1 through phosphorylation on Ser295. (A) His-t...
(A) His-tagged SYK(L) WT or the S295A mutant that was translated in vitro were incubated with samples immunoprecipitated from cells with Flag-CHK1 in the presence or the absence of ATP (as indicated) for 2 hours. The samples were then analyzed by Western blot. (B) His-tagged SYK(L) WT protein translated in vitro was incubated with Flag-CHK1 WT or its kinase dead (KD) form, which was immunoprecipitated from cells as indicated for 2 hours. The samples were then analyzed by Western blot. (C) SMMC7721 cells transfected with the indicated plasmids for 24 hours were analyzed by Western blot. (D) SMMC7721 cells were stably transfected with the WT SYK(L) or the mutant SYK(L)-S295A as indicated, and the cells were treated with GÖ6976 (100 nM) or HU (2.5 mM) for 10 hours as indicated and subjected to Western blot. (E) Huh7 cells were transfected with scramble or 3 different CHK1 siRNA duplexes as indicated for 48 hours. The cell lysates were resolved directly by SDS-PAGE (WCL) or were first incubated with anti-SYK(L) antibodies (IP: SYK) and then analyzed by Western blot.