Journal of Muscle Research and Ceil Motility 7, 537-549 (1986) The effect of calcium on the aggregation of chicken gizzard thin filaments WILLIAM LEHMAN Department of Physiology, Boston University School of Medicine, Boston, Massachusetts 02118, U.S.A. Received 27 January and in revised form 20 June 1986 Summary Electron microscopy demonstrates that thin filaments isolated from chicken gizzard smooth muscle in the absence of Ca 2+ are aggregated into networks. In contrast, thin filaments isolated in the presence of Ca 2+ are dissociated from each other. Electron microscopy also reveals that the respective state of aggregation in each type of preparation is reversible and dependent on Ca2+ concentration. Corresponding viscosity measurements indicate that network formation is associated with an increase in thin filament viscosity. We propose that thin filament aggregation in vivo may be responsible for the tension maintenance of smooth muscle during relaxation. Introduction Actin filaments appear to have a dual role in cell function. They have a structural role in the main- tenance and control of cell shape and organization and another role in many cellular motile processes. The extent to which actin filaments serve in one role or the other is determined, in part, by a variety of accessory binding proteins. Certain actin binding proteins control the balance between assembly and disassembly of filaments, others cross-link assembled filaments into bundles and networks or anchor them to other structures and yet others modulate cellular motility. In many instances actin-filament organiza- tion and actin-based cell motility is also controlled by the levels of cytoplasmic free Ca 2+ (for a review see Weeds, 1983). The structural and contractile roles of actin filaments in vertebrate muscle have been extensively studied and are frequently used as models for comparison with other tissues. Contractile activity in all muscles results from a relative sliding of interdigi- tating actin-based thin filaments and myosin- containing thick filaments. This sliding is driven by the hydrolysis of ATP by myosin-crossbridges, accompanied by their cyclic and repetitive interaction with the actin subunits of thin filaments. In all muscles, actin filaments contain the accessory protein tropomyosin. Moreover, in striated muscle the troponin-complex binds to tropomyosin and in concert they confer Ca2+-dependency on the acto- myosin interaction, regulating muscular activity. The thin filaments in striated muscle appear to be stable structures, and unlike actin-filament assemblies in many non-muscle cells which are labile, are not thought to be modulated by actin-binding proteins other than troponin-tropomyosin (for a review see Huxley, 1983). Whereas the composition of vertebrate striated muscle thin filaments is well known, the constituents of smooth muscle thin filaments are not well defined, and the distribution and function of actin-binding proteins in this tissue remain uncertain and con- troversial. Recent work, however, on isolated 'native' smooth muscle thin filaments has begun to resolve some of these uncertainties. Although such thin filaments do not contain troponin they are capable of conferring Ca 2§ dependence on myosin ATPase (Marston & Smith, 1984; Marston & Lehman, 1985). In addition to actin and tropomyosin the native filaments contain caldesmon, a potential regulator of actomyosin ATPase, and small amounts of filamin (Marston & Lehman, 1985; Lehman, 1986). Each of these actin-binding proteins is also abundant in non-muscle cells (Kakiuchi et al., 1983; Ban et al., 1984; Ngai & Walsh, 1985; Weihing, 1985), suggesting that actin filament interactions in smooth muscle and in non-muscle tissue may be related. The influence of actin-binding proteins on thin filament organization in smooth muscle is not clear. In vitro reconstitution experiments by many investi- gators (Bretscher, 1984; Dabrowska et aI., 1985; Moody et al., 1985; Chalovich et al., 1986) suggest that in addition to modulating ATPase activity caldesmon, under certain conditions, is capable of crossqinking actin filaments and that this bundling is blocked in 0142-4319/86 $03.00 + .12 9 1986 Chapman and Hall Ltd.