Abstract 104: Gene Regulation By RNA Granule Formation In Atherosclerosis

Abstract

Background: Atherosclerosis, the leading cause of cardiovascular disease related death, is a chronic inflammatory condition incited by the accumulation of low-density lipoproteins (LDL) and recruitment of macrophages and smooth muscle cells (SMCs) in the arterial intima. Both macrophages and SMCs become lipid laden foam cells that persist in the arterial vasculature, inciting chronic inflammation and plaque formation. A better understanding of the mechanisms underlying this maladaptive inflammatory response may reveal novel therapeutic strategies for intervention. Hypothesis: We hypothesize that the transformation into a foam cell state, a process that requires transcriptional rewiring, is facilitated by stress granules (SGs). SGs are cytoplasmic membrane-less organelles that sequester translation initiation factors, mRNAs, and noncoding RNAs as a result of stress. We previously reported that SG formation is increased in macrophages and vascular smooth muscle cells (SMCs) in plaques of atherosclerotic mice, and can be induced in vitro during cellular cholesterol loading. However, the consequences of SG formation and the transcripts sequestered by these organelles remain unknown Methods and Results: A current limitation in studying SG biology is the lack of in vivo models to track and isolate these dynamic structures. Using CRISPR-Cas9, we generated mice with a 3x-HA-tag on the SG marker G3BP1 in C57BL6 mice, allowing us to identify and purify SGs in vivo and in isolated primary cells. In HA-tagged G3BP1 mice fed western diet, we observe staining for HA-G3BP1 in macrophages and smooth muscle cells localized to the necrotic core and fibrous cap of atherosclerotic plaque. Furthermore, qRT-PCR on lipid laden cells revealed changes in known stress granule markers and transcripts involved in immune regulation. Conclusion: Our data support a role for SGs in repressing the translation of RNA transcripts in lipid-laden foam cells that accumulate in atherosclerotic plaque. Further study of the specific proteins and RNA transcripts sequestered in SGs of foam cells may reveal how they alter gene expression to facilitate cellular phenotypic changes in the plaque.