TRIM35 Monoubiquitinates H2B in Cardiac Cells, Implications for Heart Failure.

BACKGROUND: The tumor suppressor and proapoptotic transcription factor P53 is induced (and activated) in several forms of heart failure, including cardiotoxicity and dilated cardiomyopathy; however, the precise mechanism that coordinates its induction with accessibility to its transcriptional promoter sites remains unresolved, especially in the setting of mature terminally differentiated (nonreplicative) cardiomyocytes. METHODS: Male and female control or TRIM35 (tripartite motif containing 35) overexpression adolescent (aged 1-3 months) and adult (aged 4-6 months) transgenic mice were used for all in vivo experiments. Primary adolescent or adult mouse cardiomyocytes were isolated from control or TRIM35 overexpression transgenic mice for all in vitro experiments. Adenovirus or small-interfering RNA was used for all molecular experiments to overexpress or knockdown, respectively, target genes in primary mouse cardiomyocytes. Patient dilated cardiomyopathy or nonfailing left ventricle samples were used for translational and mechanistic insight. Chromatin immunoprecipitation and DNA sequencing or quantitative real-time polymerase chain reaction (qPCR) was used to assess P53 binding to its transcriptional promoter targets, and RNA sequencing was used to identify disease-specific signaling pathways. RESULTS: Here, we show that E3-ubiquitin ligase TRIM35 can directly monoubiquitinate lysine-120 (K120) on histone 2B in postnatal mature cardiomyocytes. This epigenetic modification was sufficient to promote chromatin remodeling, accessibility of P53 to its transcriptional promoter targets, and elongation of its transcribed mRNA. We found that increased P53 transcriptional activity (in cardiomyocyte-specific Trim35 overexpression transgenic mice) was sufficient to initiate heart failure and these molecular findings were recapitulated in nonischemic human LV dilated cardiomyopathy samples. CONCLUSIONS: These findings suggest that TRIM35 and the K120Ub-histone 2B epigenetic modification are molecular features of cardiomyocytes that can collectively predict dilated cardiomyopathy pathogenesis.

TRIM35-mediated degradation of nuclear PKM2 destabilizes GATA4/6 and induces P53 in cardiomyocytes to promote heart failure.

Pyruvate kinase M2 (PKM2) is a glycolytic enzyme that translocates to the nucleus to regulate transcription factors in different tissues or pathologic states. Although studied extensively in cancer, its biological role in the heart remains unresolved. PKM1 is more abundant than the PKM2 isoform in cardiomyocytes, and thus, we speculated that PKM2 is not genetically redundant to PKM1 and may be critical in regulating cardiomyocyte-specific transcription factors important for cardiac survival. Here, we showed that nuclear PKM2 (S37P-PKM2) in cardiomyocytes interacts with prosurvival and proapoptotic transcription factors, including GATA4, GATA6, and P53. Cardiomyocyte-specific PKM2-deficient mice (Pkm2 Mut Cre+) developed age-dependent dilated cardiac dysfunction and had decreased amounts of GATA4 and GATA6 (GATA4/6) but increased amounts of P53 compared to Control Cre+ hearts. Nuclear PKM2 prevented caspase-1-dependent cleavage and degradation of GATA4/6 while also providing a molecular platform for MDM2-mediated reduction of P53. In a preclinical heart failure mouse model, nuclear PKM2 and GATA4/6 were decreased, whereas P53 was increased in cardiomyocytes. Loss of nuclear PKM2 was ubiquitination dependent and associated with the induction of the E3 ubiquitin ligase TRIM35. In mice, cardiomyocyte-specific TRIM35 overexpression resulted in decreased S37P-PKM2 and GATA4/6 along with increased P53 in cardiomyocytes compared to littermate controls and similar cardiac dysfunction to Pkm2 Mut Cre+ mice. In patients with dilated left ventricles, increase in TRIM35 was associated with decreased S37P-PKM2 and GATA4/6 and increased P53. This study supports a previously unrecognized role for PKM2 as a molecular platform that mediates cell signaling events essential for cardiac survival.

MFN2-driven mitochondria-to-nucleus tethering allows a non-canonical nuclear entry pathway of the mitochondrial pyruvate dehydrogenase complex.

The mitochondrial pyruvate dehydrogenase complex (PDC) translocates into the nucleus, facilitating histone acetylation by producing acetyl-CoA. We describe a noncanonical pathway for nuclear PDC (nPDC) import that does not involve nuclear pore complexes (NPCs). Mitochondria cluster around the nucleus in response to proliferative stimuli and tether onto the nuclear envelope (NE) via mitofusin-2 (MFN2)-enriched contact points. A decrease in nuclear MFN2 levels decreases mitochondria tethering and nPDC levels. Mitochondrial PDC crosses the NE and interacts with lamin A, forming a ring below the NE before crossing through the lamin layer into the nucleoplasm, in areas away from NPCs. Effective blockage of NPC trafficking does not decrease nPDC levels. The PDC-lamin interaction is maintained during cell division, when lamin depolymerizes and disassembles before reforming daughter nuclear envelopes, providing another pathway for nPDC entry during mitosis. Our work provides a different angle to understanding mitochondria-to-nucleus communication and nuclear metabolism.