The effects of selective histone extraction on the structure of erythrocyte chromatin


Brasch, Klaus R.




The fine structure of chromatin appears to be at least partially determined by the close association between DNA and histone proteins (Ris, 1966b). Some evidence suggests that various histone components may play different roles In the gross configuration of chromatin within the cell nucleus (Littau et al., 1965). The present work consisted of a light and electron microscopic study of the structural organization of chromatin in isolated fowl erythrocyte nuclei, both before and after selective acid extraction of histone proteins. The effects of histone removal on the electron staining characteristics of various heavy metals were also examined. The chromatin of both in situ and isolated nuclei was localized in large condensed masses. An increase in condensation of chromatin was observed, when isolated nuclei were washed in 0.01 M citric acid (pH 2.6) prior to histone extraction. Suspension of nuclei in 0.1 M citric acid (pH 2.0), accompanied by the removal of histones predominantly rich in lysine and partial removal of histones rich in serine, resulted in complete gelation of all chromatin. Subsequent extraction of the remaining histones rich in lysine and serine, followed by removal of arginine-rich components, led to a progressive loosening of the chromatin. Ultimately, only a skeletonized remnant of the original o chromatin masses remained, which consisted of 20 to 30 A thick fibrils and scattered centres of electron-dense material. At this point, as during all stages of the histone extraction sequence, the gross distribution of large chromatin masses was retained. The results in general suggest that histone proteins contribute mainly to the longitudinal integrity of chromatin fibrils and that agents other than histones may be responsible for both the interlocking of DNA-strand terminals and the gross configuration of chromatin within the nucleus. Investigations with indium trichloride and uranyl acetate indicated that only the former stains nucleic acids selectively, while histone proteins contribute substantially to uranyl acetate staining.






Carleton University

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