Posttranslational modifications of titin from cardiac muscle: how, where, and what for?

Abstract

Titin is a giant elastic protein expressed in the contractile units of striated muscle cells, including the sarcomeres of cardiomyocytes. The last decade has seen enormous progress in our understanding of how titin molecular elasticity is modulated in a dynamic manner to help cardiac sarcomeres adjust to the varying hemodynamic demands on the heart. Crucial events mediating the rapid modulation of cardiac titin stiffness are post‐translational modifications (PTMs) of titin. In this review, we first recollect what is known from earlier and recent work on the molecular mechanisms of titin extensibility and force generation. The main goal then is to provide a comprehensive overview of current insight into the relationship between titin PTMs and cardiomyocyte stiffness, notably the effect of oxidation and phosphorylation of titin spring segments on titin stiffness. A synopsis is given of which type of oxidative titin modification can cause which effect on titin stiffness. A large part of the review then covers the mechanically relevant phosphorylation sites in titin, their location along the elastic segment, and the protein kinases and phosphatases known to target these sites. We also include a detailed coverage of the complex changes in phosphorylation at specific titin residues, which have been reported in both animal models of heart disease and in human heart failure, and their correlation with titin‐based stiffness alterations. Knowledge of the relationship between titin PTMs and titin elasticity can be exploited in the search for therapeutic approaches aimed at softening the pathologically stiffened myocardium in heart failure patients.

Compliant Titin Isoform Content Is Reduced in Left Ventricles of Sedentary Versus Active Rats

“…A 3-or 4-week exposure to voluntary running wheel exercise results in the reverse, i.e., reductions in PKC phosphorylation and increases in PKA phosphorylation, which is consistent with an increase in compliance (Hidalgo et al, 2014;Slater et al, 2017). PKG, ERK, and CaMKII are other known modifiers of cardiac stiffness (Koser et al, 2019). When PKG, ERK, and CaMKII bind to the N2B element of titin, compliance increases, and CaMKII can play a dual role by also phosphorylating the PEVK element to increase passive tension like PKC.…”

“…Titin compliance can be directly modified by a wide array of post-translational modifications (Koser et al, 2019), of which phosphorylation is the most well characterized. For example, a single bout of treadmill exercise reduced compliance by increasing PKC and reducing PKA phosphorylation of titin (Muller et al, 2014).…”

“…In mice, PKG was not shown to be different after 4weeks of exercise (Hidalgo et al, 2014), but the effect of long term exercise on ERK and CamKII phosphorylation targets on titin is not yet described. Similarly, the effect of exercise on other modifiers of titin such as oxidative stress (including disulphide bonding and glutathionylation) (Alegre-Cebollada et al, 2014;Koser et al, 2019) are not known.…”

“…In the current study, we assessed total phosphorylation using ProQ Diamond staining, which has previously helped identify titin phosphorylation sites, but we observed no difference between the sedentary and active group. Two challenges of ProQ Diamond staining are first that it may not reflect PKC phosphorylation of the PEVK element of titin (Hudson et al, 2011) and second that more than 300 total putative phosphorylation sites on titin have been detected by mass spectrometry (Koser et al, 2019) meaning it is not specific. Nonetheless, we might speculate that, consistent with results reported in 4-week studies, exercise is likely to be associated with a reduction in PKC phosphorylation of the PEVK element, along with trends in increases of PKA of the N2B element (Hidalgo et al, 2014;Slater et al, 2017).…”

A HaloTag-TEV genetic cassette for mechanical phenotyping of proteins from tissues

“…4b). We propose that this low cooperativity in native titin's mechanical folding can stem from heterogeneous mechanical properties of the multiple contributing Ig domains 10 and from mechanically active posttranslational modifications, such as disulfide bonds, S-thiolation and phosphorylation 28,60 , which are challenging to incorporate in recombinant proteins. In any case, this low cooperativity comes with the advantage that titin can generate mechanical work at a wider range of forces than previously estimated 15 .…”