The Role of SREBP1-FGL1 Axis in Metabolism and Cancer Immunity

Background: The connection between cancer and lipid dysregulation is well established, as abnormal lipid metabolism has been linked to tumor growth and progression. The sterol regulatory element binding protein (SREBP) pathway plays an important role in regulating lipid metabolism and cancer. Dysregulated lipid metabolism affects cellular signaling pathways to enhance synthesis and uptake of lipids, contributing to the proliferation and development of cancerous cells. Fibrinogen-like protein 1 (FGL1) is a liver secreted hepatokine that has been shown to support hepatocyte proliferation and liver regeneration. Abnormal FGL1 expression has been linked to lipid accumulation and inflammation and is also associated with the development of metabolic diseases. Interestingly, FGL1 was recently identified as a strong and specific ligand of lymphocyte activation gene 3 (LAG3), an immune inhibitory receptor that is found on activated T-cells. FGL1 expression in tumor cells was found to inhibit the anti-tumor activity of T-cells, and thus acts as an immune-checkpoint for cancer survival. Importantly, inactivation of FGL1 was shown to enhance the efficiency of immunotherapy and restore T-cell function both in vitro and in vivo. While a few studies suggest a correlation between SREBP1 and FGL1, a direct link between the two remains unexplored. Publicly available ChIP-Seq data have identified potential binding of SREBP1 at the FGL1 promoter. Here, we investigate the role of SREBP1 in the regulation of FGL1. Objective: The aim of this project is:1- To characterize the role of SREBP1 in regulating FGL1 expression. 2- To investigate the role of the SREBP1-FGL1 axis on lipid metabolism and tumor immunity. Methods: The SREBP-FGL1 axis is explored in HepG2, BxPC3, and A549 cells, derived from hepatocellular, pancreatic, and non-small cell lung carcinoma, respectively. The expression of FGL1, both intracellular and secreted, in response to SREBP1/2 inactivation is determined by real-time PCR and/or Western blotting. In parallel, the activity of the SREBP pathway was monitored in the same cell lines following FGL1 knockdown. Results: We have demonstrated that SREBP1 inactivation results in the downregulation of FGL1 expression, both at the transcriptional and protein levels, in all three cancer cell lines (HepG2, A549, BxPC3). SREBP1 inactivation reduced FGL1 expression significantly in all three cell-lines. These results suggest that SREBP1 regulates the expression of FGL1 in cancer multiple cancer cells. Moreover, FGL1 knockdown in HepG2 cells showed accumulation of activated SREBP1 at the protein level, and a significant increase in mRNA levels of SREBP1c. SREBP1c is a critical target of insulin signaling. Interestingly, we found that FGL1 inactivation sensitized cells to insulin as evident by the increased levels of SREBP1c mRNA and the accumulation of active SREBP1 at the protein level. Additionally, we observed an upregulation of enzymes involved in gluconeogenesis in both SREBP1 and FGL1 knockdown HepG2 cells. This was especially obvious for phosphoenolpyruvate carboxykinase 1 (PCK1/PEPCK1), the rate-limiting enzyme in gluconeogenesis. Conclusion: Enhanced lipid synthesis and dysregulation of metabolism is a hallmark of cancer, which makes the SREBP pathway a viable target for cancer therapy. It has been demonstrated that the hepatokine FGL1 has a role in cancer immunity and the response to immunotherapy. In conclusion, this project identifies an SREBP1-FGL1 signaling axis in several FGL1-expressing cancer cell lines, suggesting that SREBP1 could regulate tumor immunity. Thus, a further understanding of this signaling axis could lay the groundwork for the development of targeted cancer therapies.