Yu L, Gorovsky M A
Department of Biology, University of Rochester, New York 14627, USA.
Mol Cell Biol. 1997 Nov;17(11):6303-10. doi: 10.1128/MCB.17.11.6303.
Although quantitatively minor replication-independent (replacement) histone variants have been found in a wide variety of organisms, their functions remain unknown. Like the H3.3 replacement variants of vertebrates, hv2, an H3 variant in the ciliated protozoan Tetrahymena thermophila, is synthesized and deposited in nuclei of nongrowing cells. Although hv2 is clearly an H3.3-like replacement variant by its expression, sequence analysis indicates that it evolved independently of the H3.3 variants of multicellular eukaryotes. This suggested that it is the constitutive synthesis, not the particular protein sequence, of these variants that is important in the function of H3 replacement variants. Here, we demonstrate that the gene (HHT3) encoding hv2 or either gene (HHT1 or HHT2) encoding the abundant major H3 can be completely knocked out in Tetrahymena. Surprisingly, when cells lacking hv2 are starved, a major histone H3 mRNA transcribed by the HHT2 gene, which is synthesized little, if at all, in wild-type nongrowing cells, is easily detectable. Both HHT2 and HHT3 knockout strains show no obvious defect during vegetative growth. In addition, a mutant with the double knockout of HHT1 and HHT3 is viable while the HHT2 HHT3 double-knockout mutant is not. These results argue strongly that cells require a constitutively expressed H3 gene but that the particular sequence being expressed is not critical.
尽管在多种生物体中都发现了数量上较少的不依赖复制（替换）的组蛋白变体，但其功能仍然未知。与脊椎动物的H3.3替换变体一样，嗜热栖热四膜虫中的H3变体hv2在非生长细胞的细胞核中合成并沉积。尽管hv2通过其表达显然是一种类似H3.3的替换变体，但序列分析表明它是独立于多细胞真核生物的H3.3变体进化而来的。这表明这些变体的组成型合成而非特定的蛋白质序列在H3替换变体的功能中很重要。在这里，我们证明了编码hv2的基因（HHT3）或编码丰富的主要H3的任何一个基因（HHT1或HHT2）在四膜虫中都可以被完全敲除。令人惊讶的是，当缺乏hv2的细胞饥饿时，由HHT2基因转录的一种主要组蛋白H3 mRNA很容易被检测到，而在野生型非生长细胞中几乎不合成（如果有的话）。HHT2和HHT3敲除菌株在营养生长期间均未表现出明显缺陷。此外，HHT1和HHT3双敲除突变体是可行的，而HHT2 HHT3双敲除突变体则不可行。这些结果有力地表明，细胞需要一个组成型表达的H3基因，但所表达的特定序列并不关键。
