Myosin: Structure, Function, Regulation and Disease

Abstract

The human myosin heavy chain superfamily comprises 12 different classes. Class 2 is the largest and is made up of 14 myosin heavy chain (MHC) genes (and 1 pseudogene). Class 2 myosin heavy chains form a dimer of two heavy chains, and associate with 2 pairs of light chains (essential and regulatory). Of the 13 types of class 2 myosins, 10 are expressed in striated muscle and the remaining 4 in non-muscle cells and smooth muscle. All of the class 2 myosins form filaments. In striated muscle, the filaments contain precisely 294 molecules. In contrast, non-muscle myosins form much shorter filaments (about 30 molecules). The activity of all of these myosins is regulated by an interaction of the heads (comprising the motor domain and lever) with the first part of the coiled coil (subfragment 2, S2) to form the so-called ‘interacting heads’ motif. Non-muscle and smooth muscle myosin heads are additionally regulated by an interaction of the head with the tail forming region (light meromyosin, LMM) to form a shutdown molecule. We use a combination of cell biology, and structural approaches, including electron microscopy to understand the effects of disease-causing mutations in S2 and in LMM in class 2 myosins. Mutations in striated myosins (MyHCβ, MyHC2a) cause cardiac and skeletal muscle diseases respectively, although mutations in the distal LMM region of MyHCβ are predominantly associated with skeletal muscle disease. Mutations in non-muscle myosin 2A (NM2A) cause a range of bleeding disorders. We determine how specific mutations affect the myosin's ability to form filaments and how they affect formation of its shutdown state. For example, we use circular dichroism to determine if specific mutations affect the secondary structure of LMM. We use negative stain electron microscopy to evaluate filament formation by the LMM (fused to GST, to prevent paracrystal formation) and we use GFP-tagged MHC to evaluate the ability of each mutant to incorporate into filaments in cultured cells, myotubes and adult rat cardiomyocytes (Parker et al. (2018) J. Mol. Biol). We recently solved the structure of full-length smooth muscle myosin (SMM) in its shutdown state (Scarff et al.. (2020) Nature) using CryoEM. This shows how the shutdown state is stabilised and enables us to predict how mutations might disrupt it. Together with additional structural studies, we are starting to understand how specific mutations affect myosin function, to gain a better understanding of the disease process. Support or Funding Information This research is supported by the Medical Research Council, MR/R009406/1 and MR/S023593/1 to MP.