The Journal of Cell Biology, Vol 96, 449-454, Copyright © 1983 by The Rockefeller University Press

N-Ethylmaleimide-modified subfragment-1 and heavy meromyosin inhibit reactivated contraction in motile models of retinal cones

The mechanism of contraction in motile models of teleost retinal cones has been examined by using N-ethylmaleimide (NEM)-modified myosin fragments (NEM-S-1 and NEM-heavy meromyosin [HMM]) to prevent access of native myosin to actin filaments during reactivation of contraction. In the diurnal light/dark cycle, retinal cones of green sunfish (Lepomis cyanellus) and bluegill (lepomis macrochirus) exhibit length changes of more than 90 mum. The motile myoid region of the cone contracts from 100 mum in the dark to 6 mum in the light. Motile models for cone contraction have been obtained by lysis of dark-adapted retinas with the non-ionic detergent, Brij-58. These cone motile models undergo Ca(++)-and ATP-dependent reactivated contraction, with morphology and rate comparable to those observed in vivo (Burnside, B.,B. Smith, M. Nagata, and K. Porrello, 1982, J. Cell Biol., 92:198-206). The cone myoids contain longitudinally oriented actin filaments which bind myosin subfragment-1 (S-1) to form characteristic “arrowhead” complexes which dissociate in the presence of MgATP (Burnside, B., 1978, J. Cell Biol., 78:227-246). Modification of S-1 or HMM with the sulfhydryl reagent, NEM, produces new species, NEM-S-1 or NEM-HMM, which still bind actin but which fail to detach in the presence of MgATP (Meeusen, R.L., and W.Z. Cande, 1979, J. Cell Biol., 82:57-65). We have used NEM-S-1 and NEM-HMM to test whether cone contraction depends on an actomyosin force- generating system. We find that reactivated contraction of cone models is inhibited by NEM-S-1 and NEM-HMM but not by the unmodified species, S-1 and HMM. Thus, reactivated cone contraction exhibits NEM-S-1 and NEM-HMM sensitivity as well as Ca(++)- and ATP- dependence. These observations are consistent with and actimyosin-mediated mechanism for force production during cone contraction.