Deletion of Murine Endothelial Bag3 Alters the Cellular Proteome Among Organs of Origin

Terwilliger, Z. S.; Li, F.; Pentakota, A.; Zeczycki, T. N.; Green, T. D.; Kolasa, M.; Edwards, N.; Goldman, M.; Fisher-Wellman, K.; McClung, J.

doi:10.1101/2025.09.24.678094

bioRxiv · 10.1101/2025.09.24.678094

Deletion of Murine Endothelial Bag3 Alters the Cellular Proteome Among Organs of Origin

Terwilliger, Z. S.·Li, F.·Pentakota, A.·Zeczycki, T. N.·Green, T. D.·Kolasa, M.·Edwards, N.·Goldman, M.·Fisher-Wellman, K.·McClung, J.

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Abstract

BAG3 (Bcl-2 associated athanogene 3) is a multifunctional protein with pleiotropic effects in multiple cell types. Despite our knowledge of its role in cardiovascular disease, its specific role in endothelial cells (ECs) is unknown. The purpose of this study was to identify differences in the EC proteome of multiple tissues before and after cell specific deletion of Bag3. We hypothesized that BAG3 loss would uniquely alter the baseline proteome landscape of ECs in each tissue. Cdh5(PAC)-CreERT2;Bag3f/f mice received tamoxifen (KO; n=18) or vehicle (WT; n=18) and tissues (brain, heart, lung, and peripheral skeletal muscle-skm) were collected for FACS sorting of ECs from equal numbers of male and female mice at >22 weeks of age, followed by LC-MS/MS label-free proteomics. Initial comparisons of the WT proteomes between tissues revealed differential abundance of EC proteins (p<0.05), including: 379 brain v heart; 325 brain v lung; 501 brain v skm; 354 heart v lung; 107 heart v skm; and 442 lung v skm. Additionally, the EC mitochondrial proteome was unique to each tissue of origin, with significantly (p<0.05) higher proportions dedicated to complexes I, III, IV and V, in heart compared to the other tissues. KO demonstrated the largest effect on skm ECs, increasing 30 proteins (Mybpc2 L2FC=4.81; PFKM L2FC=3.02) and decreasing 65 (CDC42 L2FC=-2.44; Prpf8 L2FC=-2.24). Overall, these results demonstrate that the EC proteome of different tissues is unique and that the loss of BAG3 in these cells differentially alters a small proportion of the EC specific proteome.

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Terwilliger, Z. S., Li, F., Pentakota, A., Zeczycki, T. N., Green, T. D., Kolasa, M., Edwards, N., Goldman, M., Fisher-Wellman, K., McClung, J.. 2025-09-26. Deletion of Murine Endothelial Bag3 Alters the Cellular Proteome Among Organs of Origin. https://doi.org/10.1101/2025.09.24.678094

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