Role of Histone modification in the development of heart failure

M. Elkenani (Göttingen)¹, X. Xu (Göttingen)¹, S. Mobarak (Göttingen)², S. Gersch (Göttingen)¹, R. L. Kosinsky (MN 55905)³, S. A. Johnsen (Göttingen)⁴, G. Hasenfuß (Göttingen)¹, K. Toischer (Göttingen)¹, B. A. Mohamed (Göttingen)^1
1Universitätsmedizin Göttingen Herzzentrum, Klinik für Kardiologie und Pneumologie Göttingen, Deutschland; ²Universitätsmedizin Göttingen Department of Clinical Chemistry Göttingen, Deutschland; ³Mayo Clinic Division of Gastroenterology and Hepatology MN 55905, USA; ⁴Universitätsmedizin Göttingen Department of General, Visceral and Pediatric Surgery Göttingen, Deutschland

Background: Monoubiquitination of histones H2B (H2Bub1), through Ring-finger protein 20 (RNF20) and Ring-finger protein 40 (RNF40), plays important roles in regulating gene expression and is an epigenetic regulator for cardiomyocyte maturation.  Our pervious next-generation sequencing results showed altered expression of Rnf40 in cardiac hypertrophy and heart failure.

Aims: To investigate whether RNF40-mediated H2B ubiquitination would affects postnatal gene expression and cardiac homeostasis.

Methods: Rnf40^flox/flox/α-MHC-MerCreMer^+ve mice were treated with tamoxifen to generate cardiomyocyte-specific Rnf40 knockout (KO) mouse model. Rnf40^flox/flox/α-MHC-MerCreMer^-ve mice with tamoxifen and Rnf40^flox/flox/α-MHC-MerCreMer^+ve mice without tamoxifen were used as controls.

Results: Mice lacking cardiomyocyte RNF40 showed cardiac hypertrophy and severe deterioration in cardiac function, accompanied by considerable interstitial fibrosis and high mortality rates vs. controls. On the molecular level, loss of RNF40 evoked re‐expression of the fetal gene program, activation of calcium/calmodulin‐dependent protein kinase II (CaMKII) and histone deacetylase 4 (HDAC4). Using the state-of-the-art single-nucleus RNA sequencing (sNucRNA-seq), we detected 22 transcriptionally distinct cell clusters that exhibited highly consistent expression patterns, and identified 12 cell populations based on cell-specific markers and significantly enriched genes. Within the cardiomyocyte clsuters, 437 genes were differentially expressed (DEGs) in Rnf40-KO vs. control hearts. DEGs were enriched in biological processes, molecular function and KEGG pathways of hypertrophic cardiomyopathy, cardiac muscle contraction, chromatin modification and mRNA splicing.

Conclusion: Our data provide evidence that altered RNF40-mediated ubiquitination of H2B is involved in heart failure development and might be a novel therapeutic target.

Outlook: We are currently performing single nuclear transposase-accessible chromatin profiling (sNucATAC-seq) on Rnf40-KO hearts to identify the chromatin accessibility in Rnf40- KO cardiomyocytes. Furthermore, integrated analysis of sNucRNA- and sNucATAC-seq will be performed to map gene expression with chromatin accessibility in Rnf40-KO hearts at single nuclear resolution. This will aid in uncovering the role of RNF40-mediated H2B ubiquitination in regulating cardiac gene expression and will provide a multidimensional picture of the impact of chromatin accessibility on gene expression in the context of heart failure development.