| Popis: |
A biochemical and ultrastructural stereo-morphological analysis, with special reference to spatial organization and length of nucleonema and Ag-positive zones, was performed for various modifications of nucleolonemal type nucleoli in normal and regenerating (6 and 22 hours after partial hepatectomy) rat hepatocytes. To determine possible disorders on nucleosomal and supranucleosomal levels, chromatin DNA degradation was carried out during micrococcal nuclease hydrolysis, followed by analysis of electrophoretically separated particles. Functional characterization of intranucleolar chromatin was performed by testing the rate of DNA degradation after DNAase I treatment as well as by detection of free G-C pairs during titration with actinomycin D. Transcriptional activity of nucleoli was determined according to the intensity of [14C]-UTP uptake with isolated nucleoli. It is shown that the total chromatin from control nucleoli contains nucleosomal fibrils, although deprived of high compactization level. Nucleosomes themselves are strongly destabilized. In activated nucleoli structural differences of chromatin are more perceptible. In 6 hour preparations the bulk of chromatin fibrils (about 70%) undergo a further relaxation and lose the nucleosomal structure. Therefore at this point of experiment, the maximum length of nucleolonema and Ag-positive zones was registered in addition to the highest quantity of free G-C pairs, and sensibility to DNAase I transcriptional activity of isolated nucleoli. 22 hours after hepatectomy, the transcriptional activity and functional parameters of intranucleolar chromatin markedly decreased compared to the 6 hour period. Simultaneously, the share of chromatin restituting the nucleosomal structure increased, while the length of nucleolonema was shorter than in nucleoli 6 hours after hepatectomy. The main results could be resumed in the following way: the general composition of nucleolonemal type nucleolus variations described in our experimental conditions is in close relation with the with the compactization grade of ribosomal DNP-fibrils. |
