Defining the function of c/EBP-beta and its downstream target PSMB9 in atherosclerosis

The National Heart, Lung, and Blood Institute awarded Emory University $162,080 on March 13, 2026, under the Cardiovascular Diseases Research program (CFDA 93.837) to fund research defining the function of C/EBP-beta and its downstream target PSMB9 in atherosclerosis.

The research investigates how the protein C/EBP-beta (CCAAT/enhancer binding protein B) and its isoforms contribute to atherosclerosis development, particularly in response to disturbed blood flow. Prior work identified C/EBP-beta upregulation in inflamed endothelial cells exposed to disturbed flow using a partial carotid ligation model. The funded work examines the functional roles of C/EBP-beta's three isoforms—LAP*, LAP, and LIP—and their interaction with PSMB9 (proteasome subunit beta type-9 immunoproteasome component), which may inform new therapeutic targets for cardiovascular disease.

The assistance type is a Project Grant. Performance of work occurs in Atlanta, Georgia. The period of performance runs through September 11, 2027. Emory University's Office of Sponsored Programs Division serves as the prime recipient.

Abstract

Despite countless significant advances in the field of cardiovascular disease, IT remains the number one cause of death worldwide. Several systemic risk factors which increase disease risk and include hypertension, high cholesterol, obesity, diabetes, smoking, and chronic inflammation. An additional local risk factor is disturbed blood flow (d-flow), which activates the endothelium to permit of atherosclerosis, and is accelerated when other risk factors are present. Despite our best efforts to treat these conditions to improve outcomes, cardiovascular disease remains the number one killer, which suggests that there remain residual unaddressed risk factors. We have identified through previous studies using the partial carotid ligation (PCL) model of d-flow causing atherogenesis that ccaat/enhancer binding protein β (c/ebpβ), a protein previously identified to play critical roles in development, cell differentiation, and inflammation, was upregulated at the mrna level in d-flow exposed inflamed endothelial cells in vivo. We found in orthogonal studies that c/ebpβ was upregulated at the protein level in regions of d-flow, and that the compared to regions of stable flow, that the protein had localized to the nucleus, suggested that IT was playing a role in downstream signaling and transcription. Three isoforms comprise c/ebpβ, which is translated from a single intron. The first isoform is LAP*, the full length protein, with activating and repression domains, the second isoform is LAP, which is nearly identical to LAP* but lacks 23 amino acids (and one of the activating domains) at the n terminal region. The short isoform is LIP, which lacks all activating domains and is thought to have a repressive role on the function of LAP* and LAP. We found that flag-tagged LIP isoform CO-immunoprecipitated with the protein subunit beta type-9 immunoproteasome component (PSMB9, alternatively named LMP2) in vitro, which notably, has also been implicated in atherosclerosis. Finally, we found that in contrast to stable flow, d-flow induced PSMB9 activity in vitro. Given these findings, we will set out to define the role of c/ebpβ and PSMB9 in atherosclerosis. We will determine whether c/ebpβ and PSMB9 are required for endothelial cell inflammation caused by d-flow in vitro using crispr-i and small molecule inhibition. We will employ a mouse model of endothelial-specific c/ebpβ or PSMB9 deletion combined with PCL and hypercholesterolemia to determine if these proteins are required by endothelial cells for the propagation of disturbed blood flow leading to endothelial inflammation and atherosclerosis in vivo. Finally we will determine whether those two proteins are over-expressed in endothelial cells from atherosclerotic regions compared to healthy control regions in human coronary arteries. We hypothesize that c/ebpβ is a key protein required for atherosclerosis, and that IT exerts its pro-atherogenic effects by binding and activating the downstream PSMB9 immunoproteasome. We hope that our findings will further elucidate unaddressed risk factors which contribute to cardiovascular disease, and will lead to improved treatments for the number one cause of death.