Nelson D, Covault J, Chalkley R
Nucleic Acids Res. 1980 Apr 25;8(8):1745-63. doi: 10.1093/nar/8.8.1745.
It has been previously shown that micrococcal nuclease digestion and subsequent fractionation of hen oviduct nuclei generates fractions enriched (first supernatant fraction - 1SF) and depleted (second supernatant fraction - 2SF) in ovalbumin genes, while a third fraction, the pellet fraction, contains about the same level of this gene as whole chromatin (Bloom and Anderson (1978) Cell 15, 141-150). We have utilized this fractionation method in an attempt to assess the extent and kinetics of histone acetylation associated with chromatin from the 1SF, 2SF, and pellet fraction. Hepatoma Tissue Culture (HTC) cells were labelled for 30 minutes in vivo with 3H-acetate, nuclei isolated and the chromatin fractionated. The specific activity of the histones in the 1SF was slightly greater than that of the 2SF (1.2 to 1.6 fold difference) independent of the length of nuclease digestion. If the labelling period is followed by short (10 to 60 minute) treatment of the cells with sodium butyrate, the more rapidly as well as more extensively acetylated histones are also preferentially found in the 1SF. This is in part the result of segregation of chromatin particles into the 1SF as the histones associated with these particles become hyperacetylated. That is, the extent of histone acetylation regulates the distribution of chromatin in the 1SF, 2SF and pellet fraction.
先前的研究表明，用微球菌核酸酶消化鸡输卵管细胞核并随后进行分级分离，可产生富含卵清蛋白基因的级分（第一上清级分-1SF）和贫含卵清蛋白基因的级分（第二上清级分-2SF），而第三个级分，即沉淀级分，所含该基因的水平与整个染色质中的水平大致相同（Bloom和Anderson（1978年），《细胞》15卷，141 - 150页）。我们利用这种分级分离方法来评估与1SF、2SF和沉淀级分中的染色质相关的组蛋白乙酰化程度和动力学。将肝癌组织培养（HTC）细胞在体内用³H - 乙酸标记30分钟，分离细胞核并对染色质进行分级分离。1SF中组蛋白的比活性略高于2SF（相差1.2至1.6倍），与核酸酶消化的时长无关。如果在标记期后用丁酸钠对细胞进行短时间（10至60分钟）处理，在1SF中也优先发现乙酰化更快且更广泛的组蛋白。这部分是由于随着与这些颗粒相关的组蛋白高度乙酰化，染色质颗粒分离到1SF中的结果。也就是说，组蛋白乙酰化程度调节染色质在1SF、2SF和沉淀级分中的分布。
