Localization of Myosin in the Internodal Cell of Nitella as Suggested by Differential Treatment with N-Ethylmaleimide

Abstract

ABSTRACT. Cytoplasmic streaming in the internodal cell of Nitella ceased quickly following the application of the SH-group modifying reagent Nethylmaleimide (NEM). Centrifugation and NEM application were combined in a single cell mounted in a double-chambered vessel. This arrangement made possible the separate treatment of the streaming endoplasm and stationary cortex with NEM. Streaming occurred normally when the endoplasm was kept intact while the cortex was treated with the reagent. In the reciprocal case, in which endoplasm was treated while cortex was untreated, no streaming occurred. The results obtained suggest that myosin molecules are not intermingled with actin filaments localized on the cortex but reside in the endoplasm, since it is known that NEM inhibits actomyosin-type ATPase conspicuously while it has little effect on F-actin. It was tentatively concluded that the cytoplasmic streaming in Nitella results from the sliding of the endoplasmic myosin molecules alongside the stationary F-actin filaments lying on the cortex.

Differential Heat Treatment of the <I>Nitella</I> Internodal Cell and Its Relation to Cytoplasmic Streaming

“…When only the endoplasm was treated with the same reagent, and the ectoplasm was left untreated, no streaming took place (2). When cytochalasin B (CB) was used as the reagent, the stivation was reversed, i.e., cytoplasmic streaming was stopped only when the cortex was treated (13).…”

III-5 Cytoplasmic Streaming

“…To find the intracellular localization of the putative myosin in Nitella, Chen and Kamiya (1975) developed a new method in which the flowing endoplasm and the s81 s82 stationary cortex could be treated separately with an appropriate agent. By combining this method with treatment with N-ethylmaleimide (NEM) (Chen and Kamiya, 1975), cytochalasin B (CB) (Nagai and Kamiya, 1977) or supraoptimal temperature (Chen and Kamiya, 1981), myosin was shown to be localized in the flowing endoplasm and not in the stationary cortical layer, and the actin participating actively in producing the motive force was found to reside in the cortex.…”

“…By combining this method with treatment with N-ethylmaleimide (NEM) (Chen and Kamiya, 1975), cytochalasin B (CB) (Nagai and Kamiya, 1977) or supraoptimal temperature (Chen and Kamiya, 1981), myosin was shown to be localized in the flowing endoplasm and not in the stationary cortical layer, and the actin participating actively in producing the motive force was found to reside in the cortex. Using a somewhat different technique, Kamitsubo (1981) came to the same conclusion.…”

Dependency of Cytoplasmic Streaming on Intracellular ATP and Mg<SUP>2+</SUP> Concentrations

“…The relationship between the direction of the arrowheads and that of cytoplasmic streaming has also been clarified (9). Chen and Kamiya (2), and Kuroda and Kamiya (11) showed evidence suggesting that another (putative) component, myosin, is localized in the sol layer. Kato and Tonomura (8) purified myosin from Nitella flexillis.…”