Cellular distribution of thyroid myosin.
The subcellular localization of myosin in thyroid was investigated by both immunofluorescence and biochemical techniques. Dog thyroid cells stained with antisera to gizzard or thymus myosins showed that epithelial cells from thyroid contain nonmuscle myosin but not smooth muscle myosin. The antimyosin staining appeared at the periphery of the cell and in fibrils within the cell. The nature and subcellular localization of the myosin were further probed using biochemical techniques. Bovine thyroid plasma membranes were isolated by flotation on sucrose density gradients and subsequently extracted with 1% Triton X-100 to prepare an insoluble cytoskeletal fraction. After washing to remove residual Triton X-100, sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the cytoskeletal fraction demonstrated two major bands and several minor bands. The higher molecular weight band of the two major bands comigrated with the 200,000 mol wt heavy chain of myosin. Phosphorylation of the cytoskeletal fraction by thyroid myosin light chain kinase demonstrated a calcium- and calmodulin-dependent phosphorylation of the 20,000 mol wt light chain of myosin. Furthermore, the cytoskeletal fraction contained a myosin-type EDTA-K+ ATPase activity which was not influenced by ouabain and sodium azide. These results demonstrate the association of myosin with thyroid plasma membranes. Little myosin was solubilized by incubation of the thyroid plasma membranes with 0.6 M KCl; however, the addition of 10 mM ATP and 10 mM MgCl2 solubilized most of the myosin, indicating that it is associated with the thyroid plasma membranes through interaction with actin filaments. The presence of myosin in the thyroid plasma membranes may be important in endocytosis and exocytosis involved in thyroid hormone secretion.
D000251:Adenosine Triphosphatases / Q000378:metabolism; D000818:Animals; D002462:Cell Membrane / Q000032:analysis / Q000648:ultrastructure; D002478:Cells, Cultured; D004285:Dogs; D005455:Fluorescent Antibody Technique; D007700:Kinetics; D008970:Molecular Weight; D009218:Myosins / Q000032:analysis* / Q000302:isolation & purification; D010042:Ouabain / Q000494:pharmacology; D013961:Thyroid Gland / Q000166:cytology*
D010042:Ouabain; D000251:Adenosine Triphosphatases; C026049:EDTA-ATPase; D009218:Myosins