Investigating Histone Acetylation in Lipogenic Triple Negative Breast Cancer Cells using Cell Engineering and Bioinformatics Público
Wu, Yifei (Spring 2025)
Abstract
Triple negative breast cancer (TNBC) is an aggressive breast cancer subtype that has poor outcomes when compounded with obesity. Adipocyte secreted factors (ASFs) activate metabolic pathways that change the level of acetyl-CoA, a substrate involved in histone acetylation and fatty acid synthesis, suggesting an association between epigenetic remodeling and ASFs. Although aberrant histone acetylation is a marker for cancer progression, the relationship between lipogenesis and histone acetylation is yet to be elucidated in TNBC cells. To investigate the association between chromatin structure and tumor suppressor gene (TSG) expression, we used RNA-seq and ATAC-seq to identify if chromosomal accessibility is reduced around the promoter site of downregulated TSGs in lipogenic TNBC cells. We found no significant difference in peak signal value between cells treated with adipocyte-conditioned media (ACM) vs unconditioned media (UCM), suggesting an alternative mechanism for downregulation of TSGs other than chromosomal accessibility. By performing transcription factor enrichment analysis, we found that downregulated TSGs are regulated by a set of TFs that uniquely regulates downDEGs. This suggests downregulation of TSGs is induced by dysregulation of TFs. In future work, we will determine how TNBC aggressiveness is impacted by TFs by treating cells with specific TF inhibitors. To visualize histone acetylation level and nuclear distribution in lipogenic TNBC cells, we use fusion protein technology to engineer plasmids that express histone acetylation reader probes with bromodomain (BRD) dimers fused to fluorescent proteins. We found that BT549 transfection rate with the engineered plasmids was low, likely due to transgene silencing induced by CMV promoter in the GGDestX4 expression vector. In future experiments, we will engineer a new Golden Gate compatible plasmid based on the pSBtetTA-YP_CFP vector, which has been shown to achieve high transfection rate in BT549 cells. To determine histone acetylation level and its nuclear redistribution in lipogenic TNBC cells, we will re-do transfection with the new expression vector and treat the cells with ACM or UCM.
Table of Contents
Abstract………………………………………………………..1
Background……………………………………………………2
Hypothesis……………………………………………………..6
Research aims…………………………………………………6
Methodology…………………………………………………..7
Results…………………………………………………………9
Conclusion and Future Directions…………………………….19
References…………………………………………………….22
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