Molecular Mechanism of the E99K Mutation in Cardiac Actin (ACTC Gene) That Causes Apical Hypertrophy in Man and Mouse

Abstract

We generated a transgenic mouse model expressing the apical hypertrophic cardiomyopathy-causing mutation ACTC E99K at 50% of total heart actin and compared it with actin from patients carrying the same mutation. The actin mutation caused a higher Ca 2؉ sensitivity in reconstituted thin filaments measured by in vitro motility assay (2.3-fold for mice and 1.3-fold for humans) and in skinned papillary muscle. The mutation also abolished the change in Ca 2؉ sensitivity normally linked to troponin I phosphorylation. MyBP-C and troponin I phosphorylation levels were the same as controls in transgenic mice and human carrier heart samples. ACTC E99K mice exhibited a high death rate between 28 and 45 days (48% females and 22% males). At 21 weeks, the hearts of the male survivors had enlarged atria, increased interstitial fibrosis, and sarcomere disarray. MRI showed hypertrophy, predominantly at the apex of the heart. End-diastolic volume and end-diastolic pressure were increased, and relaxation rates were reduced compared with nontransgenic littermates. End-systolic pressures and volumes were unaltered. ECG abnormalities were present, and the contractile response to ␤-adrenergic stimulation was much reduced. Older mice (29-week-old females and 38-week-old males) developed dilated cardiomyopathy with increased end-systolic volume and continuing increased end-diastolic pressure and slower contraction and relaxation rates. ECG showed atrial flutter and frequent atrial ectopic beats at rest in some ACTC E99K mice. We propose that the ACTC E99K mutation causes higher myofibrillar Ca 2؉ sensitivity that is responsible for the sudden cardiac death, apical hypertrophy, and subsequent development of heart failure in humans and mice.

Mechanical and energetic properties of papillary muscle fromACTCE99K transgenic mouse models of hypertrophic cardiomyopathy

“…A number of lines of evidence support this interpretation. Here, we report measurements of isometric force in isolated myofibrils showing that Ca 2ϩ sensitivity was 2.5 times higher, in agreement with previous assays of the ACTC E99K mouse using an in vitro motility assay and permeabilized papillary muscle (36). These results are also consistent with those from other HCM mutations, most of which have myofibrillar Ca 2ϩ sensitivity increased by 1.5 to 2.5-fold [as summarized by Values are means Ϯ SE, based on 1 value/muscle, where the value for each muscle is the mean of all the repeats performed by that muscle; n ϭ 13 ACTC E99K muscles and 12 non-TG muscles except for velocity ϭ 0.67 and 1.0, where n ϭ 11 non-TG muscles.…”

“…We used muscles from ACTC E99K transgenic mice (36) and control non-TG littermates. The heart was quickly removed and rinsed in standard saline solution plus 30 mmol/l 2,3-butanedione 2-monoxime (BDM) to remove blood.…”

“…Our experiments were based on the thermopile methodology recently adapted by Barclay and Widen (49) to measure heat output by papillary muscle of mouse heart. We have also developed a transgenic mouse model of HCM with the ␣-cardiac actin (ACTC) E99K mutation that has been demonstrated to reproduce many of the characteristics of HCM in the human heart, including enhanced myofibrillar Ca 2ϩ sensitivity, myocyte disarray, fibrosis, atrial enlargement and ventricular wall thickening, arrhythmia, and a high probability of sudden cardiac death and progression to a heart failure phenotype (36).…”

Actin in striated muscle: recent insights into assembly and maintenance

“…While the effects of specific point mutations on the functional characteristics of the proteins are beginning to be elucidated (e.g. Dyer et al 2009;Song et al 2011), and can in some cases be correlated to a certain type of disease (Chang et al 2005;Debold et al 2010;Lombardi et al 2008), the complexity of thin filament assembly and regulation eluded to in this review would indicate that we are just at the beginning of understanding its mechanisms.…”

Sarcomeric Auto-Oscillations in Single Myofibrils From the Heart of Patients With Dilated Cardiomyopathy

“…29 Namely, the magnitude of the effects of PKA on the mechanical properties is less in diseased hearts. [47][48][49][50] Therefore, this uncoupling may be preserved in cTnI-K36Q myofibrils even after replacement of Tn, resulting in slowing of the kinetics of Ca-SPOC. We consider the mechanism of the uncoupling as follows: increased or decreased basal PKA-based phosphorylation levels of other sarcomere proteins (ie, MyBP-C and titin: see discussions below) within the sarcomere occur as compensatory responses because of the chronic history of the cTnI-K36Q mutation that is maintained even after pcTn reconstitution.…”