SCYL3, as a novel binding partner and regulator of ROCK2, promotes hepatocellular carcinoma progression

Martina Mang Leng Lei; Carmen Oi Ning Leung; Eunice Yuen Ting Lau; Rainbow Wing Hei Leung; Victor Wan San Ma; Man Tong; Yin Ying Lu; Chen Yang Huang; Qiao Hua Zhu; Irene Oi Lin Ng; Stephanie Ma; Terence Kin Wah Lee

doi:10.1016/j.jhepr.2022.100604

SCYL3, as a novel binding partner and regulator of ROCK2, promotes hepatocellular carcinoma progression

JHEP Rep. 2022 Oct 13;5(1):100604. doi: 10.1016/j.jhepr.2022.100604. eCollection 2023 Jan.

Authors

Affiliations

¹ Department of Applied Biology and Chemical Technology, The Hong Kong Polytechnic University, Hong Kong.
² Department of Clinical Oncology, Queen Elizabeth Hospital, Hong Kong.
³ School of Biomedical Sciences, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong.
⁴ Comprehensive Liver Cancer Center, The Fifth Medical Center of PLA General Hospital, Beijing, China.
⁵ Department of Interventional Radiology and Oncology, Shunde Hospital, Southern Medical University, Shunde, China.
⁶ Department of Pathology, Queen Mary Hospital, The University of Hong Kong, Hong Kong.
⁷ State Key Laboratory of Liver Research, The University of Hong Kong, Hong Kong.
⁸ Research Institute for Future Food, The Hong Kong Polytechnic University, Hong Kong.
⁹ State Key Laboratory of Chemical Biology and Drug Discovery, The Hong Kong Polytechnic University, Hong Kong.

Abstract

Background & aims: SCY1-like pseudokinase 3 (SCYL3) was identified as a binding partner of ezrin, implicating it in metastasis. However, the clinical relevance and functional role of SCYL3 in cancer remain uncharacterized. In this study, we aimed to elucidate the role of SCYL3 in the progression of hepatocellular carcinoma (HCC).

Methods: The clinical significance of SCYL3 in HCC was evaluated in publicly available datasets and by qPCR analysis of an in-house HCC cohort. The functional significance and mechanistic consequences of SCYL3 were examined in SCYL3-knockdown/overexpressing HCC cells. In vivo tumor progression was evaluated in Tp53 ^KO/c-Myc ^OE mice using the sleeping beauty transposon system. Potential downstream pathways were investigated by co-immunoprecipitation, western blotting analysis and immunofluorescence staining.

Results: SCYL3 is often overexpressed in HCC; it is preferentially expressed in metastatic human HCC tumors and is associated with worse patient survival. Suppression of SCYL3 in HCC cells attenuated cell proliferation and migration as well as in vivo metastasis. Intriguingly, endogenous SCYL3 overexpression increased tumor development and metastasis in Tp53 ^KO/c-Myc ^OE mice. Mechanistic investigations revealed that SCYL3 physically binds and regulates the stability and transactivating activity of ROCK2 (Rho kinase 2) via its C-terminal domain, leading to the increased formation of actin stress fibers and focal adhesions.

Conclusions: These findings reveal that SCYL3 plays a critical role in promoting the progression of HCC and have implications for developing new therapeutic strategies to tackle metastatic HCC.

Impact and implications: SCYL3 was first reported to be a binding partner of a metastasis-related gene, ezrin. To date, the clinical relevance and functional role of SCYL3 in cancer remain uncharacterized. Herein, we uncover its crucial role in liver cancer progression. We show that it physically binds and regulates the stability and transactivating activity of ROCK2 leading to HCC tumor progression. Our data provide mechanistic insight that SCYL3-mediated ROCK2 protein stability plays a pivotal role in growth and metastasis of HCC cells. Targeting SCYL3/ROCK2 signaling cascade may be a novel therapeutic strategy for treatment of HCC patients.

Keywords: CHX, cycloheximide; EV, empty vector; GEO, Gene Expression Omnibus; HCC, hepatocellular carcinoma; HRD motif, histidine-arginine-aspartic acid motif; HTVI, hydrodynamic tail vein injection; Hepatocellular carcinoma; MLC2, myosin light chain 2; Metastasis; NTC, non-target control; OE, overexpression; Protein stability; ROCK2; ROCK2, Rho kinase 2; SB, sleeping beauty; SCYL3; SCYL3 OE, SCYL3-overexpressing; SCYL3, SCY1-like pseudokinase 3; SCYL3-ΔC, SCYL3 mutant with deletion of the C-terminal domain; TCGA, The Cancer Genome Atlas; qPCR, quantitative PCR; sg, single-guide; sh, short-hairpin.