Endo T, Masaki T
J Cell Biol. 1984 Dec;99(6):2322-32. doi: 10.1083/jcb.99.6.2322.
Antibodies to chicken fast skeletal muscle (pectoralis) alpha-actinin and to smooth muscle (gizzard) alpha-actinin were absorbed with opposite antigens by affinity chromatography, and four antibody fractions were thus obtained: common antibodies reactive with both pectoralis and gizzard alpha-actinins ([C]anti-P alpha-An and [C]anti-G alpha-An), antibody specifically reactive with pectoralis alpha-actinin ([S]anti-P alpha-An), and antibody specifically reactive with gizzard alpha-actinin ([S]anti-G alpha-An). In indirect immunofluorescence microscopy, (C)anti-P alpha-An, (S)anti-P alpha-An, and (C)anti-G alpha-An stained Z bands of skeletal muscle myofibrils, whereas (S)anti-G alpha-An did not. Although (S)anti-G alpha-An and two common antibodies stained smooth muscle cells, (S)anti-P alpha-An did not. We used (S)anti-P alpha-An and (S)anti-G alpha-An for immunofluorescence microscopy to investigate the expression and distribution of skeletal- and smooth-muscle-type alpha-actinins during myogenesis of cultured skeletal muscle cells. Skeletal-muscle-type alpha-actinin was found to be absent from myogenic cells before fusion but present in them after fusion, restricted to Z bodies or Z bands. Smooth-muscle-type alpha-actinin was present diffusely in the cytoplasm and on membrane-associated structures of mononucleated and fused myoblasts, and then confined to membrane-associated structures of myotubes. Immunoblotting and peptide mapping by limited proteolysis support the above results that skeletal-muscle-type alpha-actinin appears at the onset of fusion and that smooth-muscle-type alpha-actinin persists throughout the myogenesis. These results indicate (a) that the timing of expression of skeletal-muscle-type alpha-actinin is under regulation coordination with other major skeletal muscle proteins; (b) that, with respect to expression and distribution, skeletal-muscle-type alpha-actinin is closely related to alpha-actin, whereas smooth-muscle-type alpha-actinin is to gamma- and beta-actins; and (c) that skeletal- and smooth-muscle-type alpha-actinins have complementary distribution and do not co-exist in situ.
针对鸡的快速骨骼肌（胸肌）α-辅肌动蛋白和平滑肌（砂囊）α-辅肌动蛋白的抗体，通过亲和层析用相对的抗原进行吸收，从而获得了四个抗体组分：与胸肌和砂囊α-辅肌动蛋白均反应的共同抗体（[C]抗-Pα-An和[C]抗-Gα-An）、特异性与胸肌α-辅肌动蛋白反应的抗体（[S]抗-Pα-An）以及特异性与砂囊α-辅肌动蛋白反应的抗体（[S]抗-Gα-An）。在间接免疫荧光显微镜检查中，（C）抗-Pα-An、（S）抗-Pα-An和（C）抗-Gα-An可对骨骼肌肌原纤维的Z带进行染色，而（S）抗-Gα-An则不能。尽管（S）抗-Gα-An和两种共同抗体可对平滑肌细胞进行染色，但（S）抗-Pα-An则不能。我们使用（S）抗-Pα-An和（S）抗-Gα-An进行免疫荧光显微镜检查，以研究培养的骨骼肌细胞在肌生成过程中骨骼肌型和平滑肌型α-辅肌动蛋白的表达和分布情况。结果发现，骨骼肌型α-辅肌动蛋白在融合前的成肌细胞中不存在，但在融合后存在于其中，且局限于Z体或Z带。平滑肌型α-辅肌动蛋白弥漫性地存在于单核和成肌细胞融合后的细胞质以及与膜相关的结构上，然后局限于肌管的与膜相关的结构上。免疫印迹和有限蛋白酶解的肽图谱分析支持了上述结果，即骨骼肌型α-辅肌动蛋白在融合开始时出现，而平滑肌型α-辅肌动蛋白在整个肌生成过程中持续存在。这些结果表明：（a）骨骼肌型α-辅肌动蛋白的表达时间受到与其他主要骨骼肌蛋白的调节协调；（b）就表达和分布而言，骨骼肌型α-辅肌动蛋白与α-肌动蛋白密切相关，而平滑肌型α-辅肌动蛋白与γ-和β-肌动蛋白密切相关；（c）骨骼肌型和平滑肌型α-辅肌动蛋白具有互补分布，且在原位不共存。
