Molecular mechanisms of dysfunction of muscle fibres associated with Glu139 deletion in TPM2 gene

Abstract

Deletion of Glu139 in β-tropomyosin caused by a point mutation in TPM2 gene is associated with cap myopathy characterized by high myofilament Ca2+-sensitivity and muscle weakness. To reveal the mechanism of these disorders at molecular level, mobility and spatial rearrangements of actin, tropomyosin and the myosin heads at different stages of actomyosin cycle in reconstituted single ghost fibres were investigated by polarized fluorescence microscopy. The mutation did not alter tropomyosin’s affinity for actin but increased strongly the flexibility of tropomyosin and kept its strands near the inner domain of actin. The ability of troponin to switch actin monomers “on” and “off” at high and low Ca2+, respectively, was increased, and the movement of tropomyosin towards the blocked position at low Ca2+ was inhibited, presumably causing higher Ca2+-sensitivity. The mutation decreased also the amount of the myosin heads which bound strongly to actin at high Ca2+ and increased the number of these heads at relaxation; this may contribute to contractures and muscle weakness.

Looking for Targets to Restore the Contractile Function in Congenital Myopathy Caused by Gln147Pro Tropomyosin

“…In control, the activation of thin filaments by troponin-high Ca 2+ and the myosin heads is accompanied by the increase in the Φ E value for actin-FITC, and by the decrease in this value for Tpm-AF and S1-AEDANS. This pattern of changes has also been observed in our previous studies [ 3 , 26 , 28 ]. The changes in the value of Φ E for Tpm-AF relative to actin are interpreted here according to a widespread theory of the Tpm shifting over actin surface from the outer to the inner actin domains [ 4 ].…”

“…If S1 are introduced into fibers, and thereby induce the open functional state of thin filaments, then the Φ E values amount to 49.1 degrees (+1.3) for actin-FITC and 55.5 degrees (‒2.6) for Tpm-AF—the highest and lowest values in the histogram, respectively. Thus, as in the previous studies of conformational rearrangements of muscle proteins in ghost fibers [ 3 , 25 , 26 ], activation of thin filaments is accompanied by the increase in the Φ E value for actin-FITC and by the decrease in this value for Tpm-AF as compared to the simulation of the blocked state.…”

“…Since the actin conformation determines the ATPase activity of myosin, it is important to monitor how the binding of the mutant Tpm affects it. An increase in the Φ E value for actin-FITC was characteristic for the switching actin monomers on, i.e., for their activation ( Figure 1 a), when they contribute to the activation of myosin ATPase [ 26 ]. From the data given in Figure 3 a it follows that the mutant Tpm increases the Φ E value by 2.0 degrees at a low Ca 2+ concentration in the absence of myosin heads and up to 1.4 degrees in the presence of strongly bound myosin heads (in the presence of ADP).…”

“…The polarized fluorescence technique can detect the angular changes of the probe orientation, but not their radial shift [ 49 , 50 ]. However, we found [ 3 , 26 , 29 ] that the changes in the value of Φ E for Tpm-AF correlate with the movement of Tpm observed by EM studies [ 5 , 7 , 51 ]. Since the decrease in the value of Φ E for Tpm-AF is observed in the control under activation conditions, then such changes are considered by us as a shift of Tpm strands towards the inner actin domains [ 3 , 26 , 29 ].…”

“…However, we found [ 3 , 26 , 29 ] that the changes in the value of Φ E for Tpm-AF correlate with the movement of Tpm observed by EM studies [ 5 , 7 , 51 ]. Since the decrease in the value of Φ E for Tpm-AF is observed in the control under activation conditions, then such changes are considered by us as a shift of Tpm strands towards the inner actin domains [ 3 , 26 , 29 ]. The change in a position of the Tpm strands relative to the inner domain of actin may be associated with a disparity in the alterations in the persistence length of Tpm strands and actin filaments that presumably cause azimuthal shifting of the Tpm strands [ 6 , 26 , 28 , 29 ].…”