Comparative study of myosins present in the lateral muscle of some fish: species variations in myosin isoforms and their distribution in red, pink and white muscle

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Abstract
Myosin isoforms and their distribution in the various fibre types of the lateral muscle of eight teleost fish (representing a wide range of taxonomic groups and lifestyles) were investigated electrophoretically, histochemically and immunohistochemically. Polyclonal antisera were raised against slow (red muscle) and fast (white muscle) myosins of the mullet, and used to stain sections of lateral muscle. Antisera specific for fast and slow myosin heavy chains only (anti-FHC and anti-SHC respectively) and for whole fast and slow myosins (anti-F and anti-S respectively) were obtained, and their specificity was confirmed by immunoblotting against electrophoretically separated myofibrillar proteins. The ATPase activity of myosin isoforms was examined histochemically using methods to demonstrate their acid- and alkali-lability and their Ca-Mg dependent actomyosin ATPase. As expected, the predominant myosin (and fibre) type in the red muscle showed an alkali-labile ATPase activity, reacted with the anti-S and anti-SHC sera (but not anti-F or anti-FHC) and contained two ‘slow’ light chains, whereas the predominant myosin (and fibre) type in the white muscle showed an alkali-stable ATPase activity, reacted with anti-F and anti-FHC sera (but not anti-S or anti-SHC) and contained three ‘fast’ light chains. However, superimposed upon this basic pattern were a number of variations, many of them species-related. On analysis by two-dimensional gel electrophoresis fish myosin light chains LC1s, LC2s and LC2f migrated like the corresponding light chains of mammalian myosins, but fish LC1f consistently had a more acidic pI value than mammalian LC1f. Fish LC3f varied markedly inMr in a species-related manner: in some fish (e.g. eel and mullet) theMr value of LC3f was less than that for the other two light chains (as in mammalian myosin), whereas in others it was similar to that of LC2f (e.g. cat-fish) or even greater (e.g. goldfish). Species differences were also seen in the relative intensity of LC1f and LC3f spots given by the fish fast myosins. In most of the fish examined the red muscle layer showed some micro-heterogeneity, containing (in addition to the typical slow fibres) small numbers of fibres with a histo- and immunohistochemical profile typical of white muscle (fast) fibres. However, other immunohistochemically distinct minority fibres were found in the red muscle of the goldfish. Three types of pink muscle were distinguished: (1) a mosaic of immunohistochemically typical red and white fibres (e.g. grey mullet); (2) a ‘transition zone’ with properties intermediate between those of red and white muscle (e.g. guppy); and (3) a layer of fibres which appeared on the basis of their myosin and actomyosin ATPase activities to contain a distinct myosin type, although this could not be distinguished from the white muscle fast myosin by any of the antisera used (e.g. goldfish, cat-fish, carp). Four of the fish examined had a mosaic white muscle consisting of fibres of a very wide range of diameters. The larger sized fibres always had the histo- and immunohistochemical profile of fast (white) fibres, but the characteristics of the small fibres varied according to the species. In the trout no histo- or immunohistochemical difference between large and small fibres could be detected, and in the mullet there was a histochemical difference only. In the eel some of the smallest fibres reacted with the anti-S serum as well as with anti-fast sera, and in the carp the small fibres reacted with both anti-S and anti-SHC in addition to the anti-fast sera, but in neither species could any trace of slow light chains be found in this muscle. In these two cases the small fibres may contain a cross-reacting form of myosin analogous to mammalian embryonic myosin. The significance of the small fibres of mosaic white muscles in the context of postlarval hyperplastic growth mechanisms in fish muscle is discussed.