Cardiomyopathy-associated mutations in tropomyosin differently affect actin–myosin interaction at single-molecule and ensemble levels
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In the heart, mutations in the TPM1 gene encoding the α-isoform of tropomyosin lead, in particular, to the development of hypertrophic and dilated cardiomyopathies. We compared the effects of hypertrophic, D175N and E180G, and dilated, E40K and E54K, cardiomyopathy mutations in TPM1 gene on the properties of single actin–myosin interactions and the characteristics of the calcium regulation in an ensemble of myosin molecules immobilised on a glass surface and interacting with regulated thin filaments. Previously, we showed that at saturating Ca2+ concentration the presence of Tpm on the actin filament increases the duration of the interaction. Here, we found that the studied Tpm mutations differently affected the duration: the D175N mutation reduced it compared to WT Tpm, while the E180G mutation increased it. Both dilated mutations made the duration of the interaction even shorter than with F-actin. The duration of the attached state of myosin to the thin filament in the optical trap did not correlate to the sliding velocity of thin filaments and its calcium sensitivity in the in vitro motility assay. We suppose that at the level of the molecular ensemble, the cooperative mechanisms prevail in the manifestation of the effects of cardiomyopathy-associated mutations in Tpm.
KeywordsActin–myosin interaction Tropomyosin Cardiomyopathy mutation Calcium regulation In vitro motility assay Optical trap
The authors thank Mr. D.I. Borovkov for assistance in processing the optical trap data.
This work was funded by the Russian Foundation for Basic Research Grants 17-00-00065 (D.L.), 17-00-00070 (S.B.), 17-00-00066 (N.K.), and 18-015-00252 (G.K.); and State Program AAAA-A19-119010590010-3 (D.L.) and AAAA-A18-118020590135-3 (S.B.). This work was performed using the equipment of the Shared Research Center of Scientific Equipment of IIP UB RAS.
Compliance with ethical standards
Conflict of interest
The authors declare no conflict of interest.
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