Post-translational modification patterns on β-myosin heavy chain are altered in ischemic and non-ischemic human hearts

Phosphorylation and acetylation of sarcomeric proteins are important for fine-tuning myocardial contractility. Here, we used bottom-up proteomics and label-free quantification to identify novel post-translational modifications (PTMs) on beta-myosin heavy chain (β-MHC) in normal and failing human heart tissues. We report six acetylated lysines and two phosphorylated residues: K34-Ac, K58-Ac, S210-P, K213-Ac, T215-P, K429-Ac, K951-Ac, and K1195-Ac. K951-Ac was significantly reduced in both ischemic and non-ischemic failing hearts compared to non-diseased hearts. Molecular dynamics simulations show that K951-Ac may impact stability of thick filament tail interactions and ultimately myosin head positioning. K58-Ac altered the solvent exposed SH3 domain surface – known for protein-protein interactions – but did not appreciably change motor domain conformation or dynamics under conditions studied. Together, K213-Ac/T215-P altered loop 1’s structure and dynamics – known to regulate ADP-release, ATPase activity, and sliding velocity. Our study suggests that β-MHC acetylation levels may be influenced more by the PTM location than the type of heart disease since less protected acetylation sites are reduced in both heart failure groups. Additionally, these PTMs have potential to modulate interactions between β-MHC and other regulatory sarcomeric proteins, ADP-release rate of myosin, flexibility of the S2 region, and cardiac myofilament contractility in normal and heart failure hearts.

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Abstract P421: Analysis Of The Functional Relevance Of Human Beta-myosin Heavy Chain Post-translational Modifications

Circulation Research ◽

10.1161/res.129.suppl_1.p421 ◽

2021 ◽

Vol 129 (Suppl_1) ◽

Author(s):

Maicon Landim-Vieira ◽

Matthew C Childers ◽

Amanda Wacker ◽

Michelle C Rodriguez Garcia ◽

Rakesh Singh ◽

...

Keyword(s):

Heart Failure ◽

Myosin Heavy Chain ◽

Heavy Chain ◽

Functional Significance ◽

Fine Tuning ◽

Muscle Performance ◽

Tail Domain ◽

Sarcomeric Proteins ◽

High Confidence ◽

Post Translational Modifications

Sarcomeric proteins have been shown to be a target of post-translational modifications (PTMs). Phosphorylation and acetylation of several sarcomeric proteins have been reported to be important for fine-tuning of myocardial contractility. Given the emerging importance of understanding the potential role of PTMs on cardiac muscle performance in healthy and diseased states, we sought to identify novel PTMs on human cardiac beta-myosin heavy chain (beta-MHC). We found several high confidence beta-MHC peptides modified by K-acetylation and S- and T-phosphorylation found in non-diseased, ischemic, and non-ischemic human heart samples. Using bottom-up proteomics and label-free quantification, we identified seven high-confidence peptides (K34, K58, S210, T215, K429, K951, K1195) with K951 displaying significant reduction in acetylation levels in both ischemic and non-ischemic failing hearts compared to donor hearts. Molecular dynamics simulations were performed to better understand the functional significance of the beta-MHC PTMs. Focus was placed on modifications in the regions with greatest potential functional significance as well as modified residues with significantly altered abundance in diseased states (K951-Ac at the myosin tail nearby a binding site for myosin heads in the super-relaxed state). K951 is located in the myosin tail (S2) at the C-terminal end of simulated structure. In both unmodified and modified simulations, the tail fragment showed significant flexibility and partial unfolding at the C-terminus. In the unmodified simulations, the inter- and intra-helical contacts were maintained. However, when beta-MHC is acetylated at residue 951, these helical contacts were altered as the uncharged acetylated residue no longer formed strong hydrogen bonds with a residue of the opposite chain. This facilitated changes increase in inter-helical contacts, an increase in inter-helical distance, and disruption of the coiled-coil tail domain structure. Our study suggests that there are distinct differences in beta-MHC acetylation levels that appear to be influenced more by location of the modified residues than the type of heart disease (ischemic- and non-ischemic heart failure). Additionally, we speculate that these PTMs have the potential to modulate the interactions between beta-MHC and other regulatory sarcomeric proteins, as well as ADP-release rate of myosin, flexibility of S2 fragment, and cardiac myofilament contractility under normal and heart failure condition.

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The Ku Protein Complex Interacts with YY1, Is Up-Regulated in Human Heart Failure, and Represses α Myosin Heavy-Chain Gene Expression

Molecular and Cellular Biology ◽

10.1128/mcb.24.19.8705-8715.2004 ◽

2004 ◽

Vol 24 (19) ◽

pp. 8705-8715 ◽

Cited By ~ 31

Author(s):

Carmen C. Sucharov ◽

Steve M. Helmke ◽

Stephen J. Langer ◽

M. Benjamin Perryman ◽

Michael Bristow ◽

...

Keyword(s):

Heart Failure ◽

Myosin Heavy Chain ◽

Heavy Chain ◽

Human Heart ◽

Ventricular Myocytes ◽

Major Effect ◽

Neonatal Rat ◽

Chain Gene ◽

Human Heart Failure ◽

Peptide Mass

ABSTRACT Human heart failure is accompanied by repression of genes such as α myosin heavy chain (αMyHC) and SERCA2A and the induction of fetal genes such as βMyHC and atrial natriuretic factor. It seems likely that changes in MyHC isoforms contribute to the poor contractility seen in heart failure, because small changes in isoform composition can have a major effect on the contractility of cardiac myocytes and the heart. Our laboratory has recently shown that YY1 protein levels are increased in human heart failure and that YY1 represses the activity of the human αMyHC promoter. We have now identified a region of the αMyHC promoter that binds a factor whose expression is increased sixfold in failing human hearts. Through peptide mass spectrometry, we identified this binding activity to be a heterodimer of Ku70 and Ku80. Expression of Ku represses the human αMyHC promoter in neonatal rat ventricular myocytes. Moreover, overexpression of Ku70/80 decreases αMyHC mRNA expression and increases skeletal α-actin. Interestingly, YY1 interacts with Ku70 and Ku80 in HeLa cells. Together, YY1, Ku70, and Ku80 repress the αMyHC promoter to an extent that is greater than that with YY1 or Ku70/80 alone. Our results suggest that Ku is an important factor in the repression of the human αMyHC promoter during heart failure.

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Effects of myosin heavy chain manipulation in experimental heart failure

Journal of Molecular and Cellular Cardiology ◽

10.1016/j.yjmcc.2009.10.013 ◽

2010 ◽

Vol 48 (5) ◽

pp. 999-1006 ◽

Cited By ~ 14

Author(s):

Jeanne James ◽

Kan Hor ◽

Michael-Alice Moga ◽

Lisa Ann Martin ◽

Jeffrey Robbins

Keyword(s):

Heart Failure ◽

Myosin Heavy Chain ◽

Heavy Chain ◽

Experimental Heart Failure

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Coexistence of mitochondrial DNA and beta myosin heavy chain mutations in hypertrophic cardiomyopathy with late congestive heart failure

Heart ◽

10.1136/hrt.80.6.548 ◽

1998 ◽

Vol 80 (6) ◽

pp. 548-558 ◽

Cited By ~ 49

Author(s):

E Arbustini ◽

R Fasani ◽

P Morbini ◽

M Diegoli ◽

M Grasso ◽

...

Keyword(s):

Heart Failure ◽

Congestive Heart Failure ◽

Mitochondrial Dna ◽

Hypertrophic Cardiomyopathy ◽

Myosin Heavy Chain ◽

Heavy Chain

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Reduced Beta Myosin Heavy Chain K213 Acetylation and T215 Phosphorylation in Human Heart Failure

Biophysical Journal ◽

10.1016/j.bpj.2019.11.2447 ◽

2020 ◽

Vol 118 (3) ◽

pp. 436a

Author(s):

Amanda Wacker ◽

Michelle C. Rodriguez Garcia ◽

Maicon Landim Vieira ◽

Rakesh K. Singh ◽

Elizabeth A. Brundage ◽

...

Keyword(s):

Heart Failure ◽

Myosin Heavy Chain ◽

Heavy Chain ◽

Human Heart ◽

Human Heart Failure

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Sodium Intake and Heart Failure

International Journal of Molecular Sciences ◽

10.3390/ijms21249474 ◽

2020 ◽

Vol 21 (24) ◽

pp. 9474

Author(s):

Yash Patel ◽

Jacob Joseph

Keyword(s):

Heart Failure ◽

Myosin Heavy Chain ◽

Heavy Chain ◽

Mechanical Performance ◽

Sodium Intake ◽

Heart Association ◽

Dietary Practices ◽

High Sodium ◽

Sodium Diet ◽

High Sodium Diet

Sodium is an essential mineral and nutrient used in dietary practices across the world and is important to maintain proper blood volume and blood pressure. A high sodium diet is associated with increased expression of β—myosin heavy chain, decreased expression of α/β—myosin heavy chain, increased myocyte enhancer factor 2/nuclear factor of activated T cell transcriptional activity, and increased salt-inducible kinase 1 expression, which leads to alteration in myocardial mechanical performance. A high sodium diet is also associated with alterations in various proteins responsible for calcium homeostasis and myocardial contractility. Excessive sodium intake is associated with the development of a variety of comorbidities including hypertension, chronic kidney disease, stroke, and cardiovascular diseases. While the American College of Cardiology/American Heart Association/Heart Failure Society of America guidelines recommend limiting sodium intake to both prevent and manage heart failure, the evidence behind such recommendations is unclear. Our review article highlights evidence and underlying mechanisms favoring and contradicting limiting sodium intake in heart failure.

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