Immunocytochemical studies using a monoclonal antibody to bovine cardiac titin on intact and extracted myofibrils.
A monoclonal antibody specific to bovine cardiac titin has been identified. The antibody recognizes a common antigenic site in striated muscles of several species. In relaxed myofibrils, specific staining at the A-I junction resulted in a doublet of fluorescent bands within a sarcomere. The distance between the doublets in successive sarcomeres varied according to the degree of myofibrillar contraction. Staining on formamide-extracted myofibrils has confirmed that this epitope is located near the outer edges of isolated A bands. Selective extraction of myofibrillar proteins resulted in different staining patterns. Disrupting the structural integrity of the M-line or the A-band centre caused a significant amount of titin to translocate toward the Z-line region. In contrast, shortening of the A-band by removal of myosin from the ends of the thick filaments resulted in anti-titin staining moving closer to the M-line region. Several conclusions can be drawn from this study: (a) two aligned groups of titin molecules are placed symmetrically to the M-line in a sarcomere; (b) titin may attach directly or via intermediary protein(s) to sites near the M-line and Z-line such that the protein is under tension and (c) removal of proteins from either region results in titin staining in the opposite region. However, the edges of the A-band give some hindrance to collapse of the titin toward the M-line.
D000818:Animals; D000911:Antibodies, Monoclonal / Q000032:analysis; D000918:Antibody Specificity; D002417:Cattle; D002645:Chickens; D064211:Connectin; D006224:Cricetinae; D004591:Electrophoresis, Polyacrylamide Gel; D005455:Fluorescent Antibody Technique; D006352:Heart Ventricles / Q000032:analysis; D008854:Microscopy, Electron; D009124:Muscle Proteins / Q000032:analysis* / Q000276:immunology / Q000302:isolation & purification; D009210:Myofibrils / Q000378:metabolism* / Q000648:ultrastructure; D011494:Protein Kinases; D011817:Rabbits
D000911:Antibodies, Monoclonal; D064211:Connectin; D009124:Muscle Proteins; D011494:Protein Kinases