Joint Study on Nature of Histone Variants by Gene Disruption

在44个鸡H1和核心组蛋白基因中，有39个位于一个110 kb的主要组蛋白基因簇中。在本研究中，利用基因打靶技术，我们分别产生了几个DT4O突变体，它们分别缺乏一个或两个特定的组蛋白基因，一个110kb的主基因簇的等位基因，大约57kb的携带21个基因的簇片段，以及12个H1基因拷贝中的11个。对所产生的突变体的系统分析使我们得到了一些值得注意的结论，如下：1)所有的组蛋白基因家族都具有固有的能力，可以补偿大约一半自己成分的缺失，并基于剩余基因表达的增加，保持每种组蛋白亚型的数量在化学计量平衡中，无论主要基因簇的一个等位基因完全中断，还是两个等位基因的大约一半片段中断。因此，主要组蛋白基因簇的一个等位基因足以使细胞增殖。2)12个H1基因拷贝中的11个的缺失不会影响细胞的功能，即生长速度和整体染色质结构。这些结果表明，仅H1基因的一个拷贝就足以用于细胞增殖。3)由于任何组蛋白变体的组成没有改变，因此在簇的一个等位基因缺失的情况下，蛋白质图谱没有改变，但在簇的大约一半片段缺失的情况下，蛋白质图谱发生变化，并且由于H1和核心变体的质量的变化，后者加上更多的h1基因(S)。4)在突变体中，蛋白质图谱发生改变，分别缺少特定的h1和h2B变体。这些可变蛋白是相应突变体所特有的，这表明H1和核心变异体应该通过改变位于特定基因组区域的染色质结构来分别参与特定基因的表达调控。

Thirty-nine of the 44 chicken H1 and core histone genes are located in a major histone gene cluster of 110 kb. In this study, using gene targeting techniques, we generated several DT4O mutants, respectively, which are devoid of one or two particular histone genes, one allele of the major gene cluster of 110 kb, an approximately 57 kb segment of the cluster carrying 21 genes, and 11 of the 12 H1 gene copies. Systematic analyses of the resultant mutants led us to some noticeable conclusions, as follows.1)All of the histone gene families have the inherent ability to compensate for the deletion of approximately half of their own constituents, and to maintain the amount of each of the histone subtypes in stoichiometric balance, based on increases in the expression of the remaining genes, regardless of the complete disruption of one allele of the major gene cluster or the disruption of approximately half segments of the two alleles. Therefore, one allele of the major histone gene cluster is enough for cell proliferation.2)The deletion of 11 of the 12 H1 gene copies does not affect cell functions, i.e. the growth rate and global chromatin structure. These results indicate that only one copy of the H1 genes is enough for cell proliferation.3)The protein patterns are not altered in the case of the deletion of one allele of the cluster, due to no changes in the composition of any histone variant, but vary in the case of the deletion of approximately half segments of the cluster, and the latter plus more Hl gene(s), due to changes in the quality of the H1 and core variants.4)The protein patterns are altered in the mutants, respectively, lacking particular Hl and H2B variants. The variable proteins are specific for the corresponding mutants, suggesting that the H1 and core variants should be individually involved in regulation of the expression of specific genes, through alterations in the chrornatin structure localized in specific genomic regions.