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. 2021 Jun 11;12:3575. doi: 10.1038/s41467-021-23858-7

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© The Author(s) 2021

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 5 — a, b RNA-Seq analysis revealed differential gene expression in hearts from 5-week-old CAG-BAG3^P209L mice (n = 6 mice) compared to CTRL (n = 6 mice). Volcano plot showing false discovery corrected q-values and log₂(fold)-changes. Blue and red dots depict genes depleted and enriched respectively in CAG-BAG3^P209L mice compared to CTRL. Names of the 20 most differentially expressed genes are shown. b Same hearts as in (a). Gene expression heatmap for significantly (P < 10⁻⁶ Bonferroni step down corrected for multiple comparison) enriched KEGG pathways identified by ClueGO (two-sided hypergeometric test) and selected genes from a fetal cardiac gene program found in pathological hypertrophy (Myh6, Atp2a2, Atp2b4, Hand2, Gata6, Mef2c, Nkx2-5, Gata4, Acta1, Lmna, Myh7, Nppa, Nppb). c Volcano plot from proteome analysis of hearts from 5-week-old CAG-BAG3^P209L vs CTRL mice. Blue and red dots depict the proteins significantly depleted and accumulated respectively in the CAG-BAG3^P209L mice compared to the CTRL (Two-sided Student’s T-test p-value <0.05, FDR < 0.05, S0 = 0.1). d KEGG term analysis of proteins with significantly increased abundance in CAG-BAG3^P209L mice compared to CTRL mice. e Raincloud plots illustrate the change in protein abundance in hearts of CAG-BAG3^P209L compared to CTRL mice for proteins known to interact with hBAG3 (n = 86) and all others (n = 629). In the box plot, center lines show the medians with box plot limits indicate the 25th and 75th percentiles. Box plot whiskers extend to 1.5x interquartile range from the 25th and 75th percentiles. Asterisk indicates a significant difference between both populations (Two-sided Mann–Whitney rank sum test, p-value 5.7E−6). f Immunoblot analysis of hearts from 5-week-old mice revealed the presence of hBAG3 and a significant increase in the amount of mouse Bag3 in CAG-BAG3^P209L mice in comparison to CTRL mice. CAG-BAG3^P209L mice had nearly as much hBAG3 as mouse Bag3. Note significant increases in the expression levels of the heat shock proteins HSPB7 and α-B crystallin, and the autophagy marker p62. Expression was normalized to the expression level of α-actin. The data were obtained from four independent biological replicates and not technically repeated. CTRL, control mice (siblings of CAG-BAG3^P209L mice, which are either WT, PGK-Cre, or CAG-flox-hBAG3^P209L).