Myomesin and M-protein: expression of two M-band proteins in pectoral muscle and heart during development.

Abstract

The expression of the myofibrillar M-band proteins myomesin and M-protein was studied in chicken pectoral muscle and heart during differentiation using monoclonal antibodies in a double-antibody sandwich enzyme-linked immunosorbent assay, immunoblotting, and immunocytochemistry. In presumptive pectoral muscle, myomesin accumulated first, increasing from 2% of the adult concentration at day 7 to 70% by day 16 in ovo. M-protein accumulation lagged 6-7 d behind that of myomesin attaining only 40% of the adult concentration in ovo. The molecular masses of myomesin (185 kD) and M-protein (165 kD) remained constant during embryogenesis. In cultured myogenic cells the accumulation and M-band localization of myomesin preceded that of M-protein by 1.5 d.Chicken heart was shown, in addition to M-protein, to contain unique isoforms of myomesin. In hearts of 6 d embryos, a 195-kD myomesin isoform was the major species; throughout development, however, a transition to a mixture of 195 and 190 kD was observed, the latter being the major species in the adult tissue. During heart differentiation the initial accumulation of myomesin again preceded that of M-protein, albeit on an earlier time scale than in pectoral muscle with M-protein reaching adult proportions first.Recently we have reported on a new 185-kD protein in the M-band of skeletal muscle myofibrils (1). This brings the number of positively identified M-band proteins to three, the other two being MM-CK (2, 3) and M-protein (4). To avoid multiple nomenclature, it has been agreed to call the 165-kD M-band protein "M-protein" and the recently identified 185-kD M-band protein "myomesin" (1, 5).Several lines of evidence indicate the functional significance of the M-band which transverses the center of the sarcomere. M-band structural components appear to provide a scaffolding for the alignment and orientation of myosin thick fdaments within myofibrils (6-10). Furthermore, there is evidence that the M-band, as well as the Z-disc, are cytoskeletal attachment points and that they assist in maintaining intermyofibril register under the stress of lateral tension imposed on muscle fibers during contraction (11-13).Of the known M-band proteins, the substructural localization and function of only one has been elucidated. MM-CK, associated with the M4, 4' substriations (14,15), is present in sufficient quantity to regenerate in isolated chicken myofibrils the ATP required for contraction (16). Attempts have been made to ascertain the roles of other M-band components in

The M-band: an elastic web that crosslinks thick filaments in the center of the sarcomere

“…In this model, M-protein molecules bridge the myosin filaments at the level of the central M1 line (see Box 1), which is consistent with previous EM observations [20,21]. However, the model cannot explain how this connection is established in muscles devoid of M-protein, such as the embryonic heart [22] and slow-twitch fibers of skeletal muscle [23,24].…”

Tissue-specific Isoforms of Chicken Myomesin Are Generated by Alternative Splicing

“…While 94% of the coding sequences are shared in all transcripts, tissue-specific sequences are present in their 3Ј-ends leading to a C-terminal segment of 98 amino acids in cardiac myomesin and 20 amino acids in the skeletal muscle isoform. The faster mobility in SDS-polyacrylamide gel electrophoresis of myomesin found in chicken skeletal muscle versus myomesin isolated from heart muscle (Grove et al, 1985) can now be explained by different C-terminal domains resulting in proteins of distinguishable calculated molecular masses of 182 kDa (1680 residues) for cardiac myomesin and 174 kDa (1552 residues) for skeletal myomesin. The skeletal muscle-specific C terminus of myomesin shares high homology with the C-terminal sequence of rat myomesin derived from an embryonic heart cDNA library as well as the C-terminal sequence of human 190K protein (Vinkemeier et al, 1993) (Fig.…”

“…analysis of tissue extracts showed a slightly higher band for myomesin in chicken heart (190)(191)(192)(193)(194)(195) than in chicken skeletal muscle (185 kDa); therefore, the existence of different myomesin isoforms in skeletal muscle and heart had been postulated (Grove et al, 1985). In order to investigate this isoform diversity, cDNAs of myomesin were isolated from two different libraries.…”

“…I, chicken heart myomesin; II, chicken skeletal muscle myomesin; III, rat myomesin; IV, human 190K protein (human counterpart of myomesin); V, mouse skelemin; VI, chicken M-protein; VII, human 160 K protein (human counterpart of M-protein). The arrow, brackets, and points at the end of sequences II-VII are as indicated for panel A. specific differences (Grove et al, 1985). Sequences derived from the putative skeletal muscle-specific clones SB194 and SB198 were aligned with the heart myomesin cDNA according to the Wilbur-Lipman algorithm.…”

Isoform-specific Interaction of the Myosin-binding Proteins (MyBPs) with Skeletal and Cardiac Myosin Is a Property of the C-terminal Immunoglobulin Domain

“…These include titin or connectin (6,7), MyBP-C (C-protein) (8), MyBP-H (H-protein) (9,10), myomesin (11), M protein (12,13), skelemin (14), MM-creatine kinase (15,16), AMP-deaminase (17), and X-protein (the slow-type isoform of MyBP-C) (18 -20). The MyBPs, first isolated in crude myosin preparations by Offer and colleagues (21) in the early 1970s are a group of proteins distributed in the central two-thirds of the cross-bridge bearing region (C-zone) of the A-band (22).…”