Knapp Amy R, Wang Huanyu, Parthun Mark R
Department of Molecular and Cellular Biochemistry, The Ohio State University, Columbus, Ohio, United States of America.
PLoS One. 2014 Jul 29;9(7):e100299. doi: 10.1371/journal.pone.0100299. eCollection 2014.
Hif1p is an H3/H4-specific histone chaperone that associates with the nuclear form of the Hat1p/Hat2p complex (NuB4 complex) in the yeast Saccharomyces cerevisiae. While not capable of depositing histones onto DNA on its own, Hif1p can act in conjunction with a yeast cytosolic extract to assemble nucleosomes onto a relaxed circular plasmid.
To identify the factor(s) that function with Hif1p to carry out chromatin assembly, multiple steps of column chromatography were carried out to fractionate the yeast cytosolic extract. Analysis of partially purified fractions indicated that Hif1p-dependent chromatin assembly activity resided in RNA rather than protein. Fractionation of isolated RNA indicated that the chromatin assembly activity did not simply purify with bulk RNA. In addition, the RNA-mediated chromatin assembly activity was blocked by mutations in the human homolog of Hif1p, sNASP, that prevent the association of this histone chaperone with histone H3 and H4 without altering its electrostatic properties.
These results suggest that specific RNA species may function in concert with histone chaperones to assemble chromatin.
Hif1p是一种H3/H4特异性组蛋白伴侣，在酿酒酵母中与Hat1p/Hat2p复合物（NuB4复合物）的核形式相关联。虽然Hif1p自身不能将组蛋白沉积到DNA上，但它可以与酵母胞质提取物协同作用，将核小体组装到松弛的环状质粒上。
为了鉴定与Hif1p共同发挥作用以进行染色质组装的因子，进行了多步柱色谱法对酵母胞质提取物进行分级分离。对部分纯化级分的分析表明，Hif1p依赖性染色质组装活性存在于RNA而非蛋白质中。对分离出的RNA进行分级分离表明，染色质组装活性并非简单地与总RNA一起纯化。此外，RNA介导的染色质组装活性被Hif1p的人类同源物sNASP中的突变所阻断，这些突变阻止了这种组蛋白伴侣与组蛋白H3和H4的结合，而不改变其静电性质。
这些结果表明，特定的RNA种类可能与组蛋白伴侣协同作用以组装染色质。
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