Julien Ochala (1), Anders Oldfors (2), & Lars Larsson (1, 3)

1: Department of Clinical Neurophysiology, Uppsala University Hospital, Sweden
2: Department of Pathology, Sahlgrenska University Hospital, Göteborg, Sweden
3: Center for Development and Health Genetics, The Pennsylvania State University, University Park, USA

https://doi.org/10.36866/pn.69.20

In skeletal muscle fibres, tropomyosin, a regulatory protein, coils around the actin filament, relaying the information from the Ca2+ sensor, the troponin complex to the actin-myosin interactions, the cross-bridges. Tropomyosin alternates between three positions: the blocked, closed and open states.

In the absence of Ca2+, tropomyosin is in a position on the outer domain of actin that sterically hinders the docking of cross-bridges – the blocked state. Full activation by reversal of steric blocking involves two additional states, requiring successive tropomyosin movements away from the blocked configuration. Ca2+ bindings to the troponin complex cause tropomyosin movement toward the inner domain of actin, exposing sites that allow weak binding of myosin heads while still inhibiting isomerization to the strong binding state. Following this change, tropomyosin still covers an essential part of the site, leaving it inaccessible to myosin heads – the closed state. Weak-to-strong myosin binding transition induces a second tropomyosin shift, permitting cooperative binding of additional myosin heads by exposing neighbouring sites – the open state.

Three major tropomyosin isoforms are expressed, α, β and γ, which are encoded by the TPM1, TPM2, and TPM3 genes, respectively (Perry, 2001). The α isoform encoded by TPM1 is predominantly expressed in cardiac and fast-twitch skeletal muscle fibres. The β isoform encoded by TPM2 is mainly expressed in slow-twitch skeletal muscle fibres. The γ isoform encoded by TPM3 is predominantly expressed in cardiac and slow-twitch skeletal muscle fibres. A number of mutations in the TPM1, TPM2, and TPM3 genes have been reported to cause inappropriate α, β and γ tropomyosin isoforms and inherited myopathies. Eight mutations in the TPM1 gene induce amino acids changes on the α tropomyosin isoform (Glu40Lys, Glu54Lys, Ala63Val, Lys70Thr, Val95Ala, Asp175Asn, Glu180Gly, Glu180Val), and are not associated with skeletal myopathies but solely with cardiomyopathies. Eight mutations in the TPM2 and TPM3 genes have been associated with amino acids modifications on the β (Arg91Gly, Glu117Lys, Arg133Trp, Glu139del, Glu147Pro) and γ (Met9Arg, Glu31stop, Arg167His) tropomyosin isoforms and with general muscle weakness and skeletal myopathies (Fig.1).

Recent studies approach the mechanisms by which three mutations in β (Arg91Gly, Arg133Trp) and γ (Met9Arg) tropomyosin isoforms cause general weakness in inherited diseases, in the absence of muscle wasting.

The Arg91Gly and Arg133Trp mutations on the β tropomyosin isoform appear to preserve the information from the Ca2+ sensor, the troponin complex to the cross­bridges. The Ca2+-activation of the contractile proteins is unaltered as attested by the force-pCa relationships (Ochala et al. 2007; Robinson et al. 2007) in contrast to the Met9Arg mutation on the γ tropomyosin isoform where the force-pCa curve was shifted to the right (Michele et al., 1999). The lack of effects of the Arg91Gly and Arg133Trp mutations on force-pCa relationships is surprising but may be related to their location in a region of the tropomyosin segment that is not directly associated with the troponin complex Ca2+ binding sites (Fig. 2). In skeletal muscle fibres, the critical interactions between tropomyosin and troponin T take place between tropomyosin residues 0–30 and 190–284. The Arg91Gly and Arg133Trp mutations on the β tropomyosin isoform are 60 residues away from the nearest potential Troponin T­anchoring region, in contrast to the Met9Arg mutation on the γ tropomyosin isoform (Fig. 2) (Perry, 2001).

The Arg91Gly and Arg133Trp mutations on the β tropomyosin isoform appear to directly modify the cross-bridge kinetics as attested by the decreased maximal apparent rate constant of force redevelopment and increased maximum unloaded shortening velocity in fibres containing Arg133Trp (Ochala et al. 2007) or increased maximal ATPase rates in fibres expressing Arg91Gly (Robinson et al. 2007). The mutations from Arg to Gly or Arg to Trp replace charges predicted to stabilize tropomyosin coiled-coil (Tajsharghi et al. 2007). Therefore, the Arg133Trp mutation may affect tropomyosin movement toward the inner domain of actin and the transition from the blocked state to the closed and open states. This would disrupt attachment and/or transition of myosin heads from the weakly to the strongly bound force­generating state, decreasing the cross-bridge attachment rate and the maximal apparent rate constant of force redevelopment. Moreover, the Arg91Gly and Arg133Trp mutations may affect tropomyosin movement from the open to the blocked state, slowing myosin attachment and promoting dissociation of myosin heads from the actin filament; thereby increasing the cross-bridge detachment rate and maximum unloaded shortening velocity or maximal ATPase rates. The combination of a decreased apparent rate of force redevelopment and an increased maximum unloaded shortening velocity in fibres expressing Arg133Trp induces a shortened cross-bridge duty cycle, i.e. a shorter time spent by myosin heads in a strongly bound force­generating state, resulting in a decreased force generating capacity.

In conclusion, studies on the three Arg91Gly, Arg133Trp and Met9Arg mutations in β and γ tropomyosin isoforms suggest specific mechanisms underlying the weakness, in the absence of muscle wasting, in patients carrying these mutations. However, the mechanism is mutation-specific and results obtained from one mutation cannot be generalized to other tropomyosin mutations. Further, the results from these mutations analyses indicate the critical role of tropomyosin isoform expression in modulating muscle contraction under physiological conditions.

Acknowledgements

The project was supported by grants from the Swedish Institute and Association Française contre les Myopathies to J O, from the Association Française contre les Myopathies and Swedish Research Council (07122) to A O, and from the Swedish Research Council (08651), Association Française contre les Myopathies, Cancer Foundation and National Institutes of Health (AR045627, AR047318) to L L.

References

Michele DE Albayya FP & Metzger JM (1999). A nemaline myopathy mutation in alpha­tropomyosin causes defective regulation of striated muscle force production. J Clin Invest 104, 1575-1581.

Ochala J, Li M, Tajsharghi H, Kimber E, Tulinius M, Oldfors A & Larsson L (2007). Effects of a R133W beta-tropomyosin mutation on regulation of muscle contraction in single human muscle fibres. J Physiol 581, 1283­1292.

Perry SV (2001). Vertebrate tropomyosin: distribution, properties and function. J Muscle Res Cell Motil 22, 5-49.

Robinson P, Lipscomb S, Preston LC, Altin E, Watkins H, Ashley CC & Redwood CS (2007). Mutations in fast skeletal troponin I, troponin T, and beta-tropomyosin that cause distal arthrogryposis all increase contractile function. Faseb J 21, 896-905.

Tajsharghi H, Kimber E, Holmgren D, Tulinius M & Oldfors A (2007). Distal arthrogryposis and muscle weakness associated with a beta­tropomyosin mutation. Neurology 68, 772­775.