Abstract 5181: SPTBN1D1089Y/+ aggravates liver carcinogenesis via activating SIRT6 and TGF-beta signaling
Authors: Kazufumi Ohshiro, Xiyan Xiang, Xiaochun Yang, Krishanu Bhowmick, Lopa Mishra
Journal: Cancer Research (2025)

Abstract Background: Obese-driven liver cancer (HCC) is alarmingly rising in the US. Yet, key signaling mechanisms underlying this obesity-driven HCC remain unclear. SIRT6 is a NAD+-dependent deacetylase that limits the activity of the lipogenic transcription factors SREBP1/2 and TGF-β signaling. Reduced SIRT6 is associated with human fatty liver and HCC. We found that TGF-β/SMAD3 signaling pathway and SREBP1/2 are modulated by βII-spectrin (encoded by SPTBN1), and a mutation in human HCC, D1089Y abrogates SMAD3 binding (Gastro 2018; 154:195-210). In stressed hepatocytes, βII-spectrin promotes lipogenesis through interactions between Caspase-3-cleaved forms of both SREBP1 and βII-spectrin (Sci TM 2021;13:624). Liver-specific knockout of βII-spectrin blocks fatty liver and HCC. Here, we examined the extent of SPTBN1-D1089Y regulation of SIRT6 expression through SMAD3, and the combined roles of SIRT6 and SPTBN1 in human HCC. Methods: We generated a SPTBN1-D1089Y mutant mouse and examined SIRT6 expression in liver tissues and MEFs. Four-month-old Sptbn1D1089Y/+ mutant and WT male mice were injected with DEN/CCI4 and fed on WD.TGF-β/SMAD3 pathway activity was analyzed by Western blotting and RNA-seq. SIRT6 expression was examined in mouse MASLD/MASH. Structure-based molecular docking simulations using structural coordinates of human SIRT6 and SPTBN1 from the AlphaFold database. Results: SIRT6 expression is significantly decreased in liver macrophages from mice with MASLD/MASH. Sptbn1D1089Y/+ mice receiving DEN/CCI4 plus WD displayed significantly more tumor nodules with severe liver injury, steatosis, and inflammation. TGF-β/SMAD3 pathway activity is suppressed in SPTBN1-D1089Y mutant MEFs with decreased phosphor-SMAD3 levels. SIRT6 levels are decreased in Huh7 cells overexpressing SPTBN1-D1089Y and the D1089Y-mutant mouse liver tissues. Structural modeling predicts that SIRT6 binding sites of SPTBN1 (R429-S439) are close to the region mediating SREBP1/SPTBN1 interaction. In HepG2 cells, overexpression of SIRT6 deacetylates βII-spectrin; SMAD3 and βII-spectrin were found to interact with SIRT6 upon TGF-β treatment by co-immunoprecipitation; SIRT6 knockdown increased nSREBP1, SCD1, and βII-spectrin. SPTBN1-WT overexpression increased slightly HCC cell viability, whereas SPTBN1-D1089Y mutant increased approximately more than two times of the WT viability. Exogenous SPTBN1-WT increased the expression of transcriptional repressor CTCF and SIRT6, and SPTBN1-D1089Y mutant decreased both expressions. Conclusions: Our study indicates a SMAD3 dependent as well as a SMAD3 independent role of SPTBN1 in regulating SIRT6. Sptbn1-D1089Y heterozygous loss aggravates liver carcinogenesis. Sptbn1-D1089Y together with SIRT6 deficiency may increase the susceptibility to MASH/HCC. Citation Format: Kazufumi Ohshiro, Xiyan Xiang, Xiaochun Yang, Krishanu Bhowmick, Lopa Mishra. SPTBN1D1089Y/+ aggravates liver carcinogenesis via activating SIRT6 and TGF-beta signaling [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 5181.