Caldesmon exhibits a clustered distribution along individual chicken gizzard native thin filaments

Our earlier immuno-gold electron microscopic study indicated that the distr ibution of caldesmon (CaD) on actin filaments is not uniform and is restric ted to the vicinity of the myosin filaments (Mabuchi K, Li Y, Tao T, Wang C -LA (1996) J Muscle Res Cell Motil 17: 243). This suggested that CaD could effectively inhibit muscle contraction, if those actin filaments in the vic inity of myosin filaments were saturated with CaD. In the present study we further examined the distribution of CaD along isolated, crude and purified native thin filaments (NTF). Individual CaD molecules on purified NTF were visualized with the aid of a chemical crosslinker, 5,5'-dithiobis(2-nitrob enzoic acid), which efficiently crosslinks CaD to actin (Graceffa P, Adam L P, Lehman W (1993) Biochem J 294: 63), and of a monoclonal anti-CaD antibod y. The results indicated that individual NTF had alternating CaD-rich and C aD-deficient regions. Moreover, we found that the N-termini of all CaD mole cules in a given cluster appeared on the same side of an actin filament. El ectron microscopic images of crude NTF immunoprecipitated by a polyclonal a ntibody clearly indicated that the spacing between the CaD clusters is wide enough for myosin heads to interact with actin subunits. Similar clusterin g of CaD was also observed in plastic embedded tissue sections. These obser vations raise the possibility that CaD is not acting as a simple on/off swi tch, but more likely as a modulator, of smooth muscle contraction.