Cytochimie ultrastructurale du nucléole: III. Action de l'actinomycine D sur le métabolisme du RNA nucléolaire

Abstract

The distribution of newly synthesized RNA among the components of the nucleolus was studied by ultrastructural autoradiography after incorporation of labeled precursor. Monkey kidney cells in culture were labeled with tritiated uridine for 5 min. After a thorough wash with buffer, they were cultivated in the presence of cold uridine for 10, 30 or 60 min. After 5 min incorporation and immediate fixation, the activity resulting from the incorporation of the radioactive precursor into new RNA is localized only in the fibrillar elements of the nucleoli. During the chase on the other hand, the reduced silver grains appear primarily over the granules of the nucleoli. Experimental corroboration of this localization of labeled RNA was obtained in cells treated by Actinomycin D with the intention of obtaining a spatial segregation of the fibrillar, granular and amorphous components of this organelle. The cells were labeled with the same precursor for 5 or 15 min and then cultivated in cold medium with Actinomycin D for 30 min, 2 h or 7 h. After 5 min incorporation and 2 h or 7 h subsequent treatment with Actinomycin D, the nucleoli have segregated into their three distinct zones: granular, fibrillar and amorphous. The radioactivity is concentrated in the granular zone. Only a few grains are localized over the fibrillar zone. The third zone is not labeled. These results are consistent with the hypothesis of a migration of RNA from the fibrillar zone to the granular zone, this migration not being blocked by actinomycin D.