Toxoplasma H2A Variants Reveal Novel Insights into Nucleosome Composition and Functions for this Histone Family
Toxoplasma H2A Variants Reveal Novel Insights into Nucleosome Composition and Functions for this Histone Family
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DOI：
10.1016/j.jmb.2009.07.017
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发表时间：
2009-09-11
影响因子：
5.6
通讯作者：
Angel，Sergio O.
中科院分区：
文献类型：
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作者：
Dalmasso，Maria C.;Onyango，David O.;Angel，Sergio O.
弓形虫是一种专性胞内寄生虫。弓形虫病是不治之症，因为它有能力从快速复制的速殖子阶段分化为潜伏的包囊形式(缓殖子阶段)。与弓形虫分化相关的基因调控涉及组蛋白修饰，但对这种早期分枝的真核生物中的组蛋白知之甚少。在这里，我们报告了三个H_2A组蛋白，变异体H_2AX和H_2AZ，以及一个典型的H_2A1的特征。H_2AZ是次要寄生虫H_2A成员。H_2A1和H_2AX都有一个SQ基序，但只有H_2AX有一个完整的SQ(E/D)ϕ(其中ϕ表示疏水残基)，该基序在DNA损伤时被磷酸化。我们发现一种新的H_2B变异体与H_2AZ和H_2A1相互作用，但不与H_2AX相互作用。染色质免疫沉淀法(ChIP)显示，H_2AZ和H_2Bv在活性基因上富含，而H_2AX在抑制基因和沉默的TgIRE重复元件上富含。在DNA损伤过程中，我们检测到H_2AX磷酸化增加以及H_2a1和H_2AX转录增加。我们发现，在体外产生的缓殖子中，h2AX的表达增加，而h2a1或h2az的表达没有增加。对体内产生的成熟缓殖子进行的类似分析表明，h2az和h2ax表达，但h2a1不表达，这与h2a1是寄生虫中典型的组蛋白同源基因的想法一致。在缓殖子分化过程中，H_2AX的增加局限于沉默的区域，这与生命周期这个阶段的静止性质是一致的。我们的结果表明，早期分枝真核寄生虫弓形虫含有新组成的核小体，这可能会影响寄生虫生物学的多个方面，包括临床上重要的缓殖子分化过程。
Toxoplasma gondii is an obligate intracellular parasite. Toxoplasmosis is incurable because of its ability to differentiate from the rapidly replicating tachyzoite stage into a latent cyst form (bradyzoite stage). Gene regulation pertinent to Toxoplasma differentiation involves histone modification, but very little is known about the histone proteins in this early branching eukaryote. Here, we report the characterization of three H2A histones, variants H2AX and H2AZ, and a canonical H2A1. H2AZ is the minor parasite H2A member. H2A1 and H2AX both have an SQ motif, but only H2AX has a complete SQ(E/D)ϕ (where ϕ denotes a hydrophobic residue) known to be phosphorylated in response to DNA damage. We show that a novel H2B variant interacts with H2AZ and H2A1 but not with H2AX. Chromatin immunoprecipitation (ChIP) revealed that H2AZ and H2Bv are enriched at active genes while H2AX is enriched at repressed genes as well as the silent TgIRE repeat element. During DNA damage, we detected an increase in H2AX phosphorylation as well as increases in h2a1 and h2ax transcription. We found that expression of h2ax, but not h2a1 or h2az, increases in bradyzoites generated in vitro. Similar analysis performed on mature bradyzoites generated in vivo, which are arrested in G0, showed that h2az and h2ax are expressed but h2a1 is not, consistent with the idea that h2a1 is the canonical histone orthologue in the parasite. The increase of H2AX, which localizes to silenced areas during bradyzoite differentiation, is consistent with the quiescent nature of this stage of the life cycle. Our results indicate that the early-branching eukaryotic parasite Toxoplasma contains nucleosomes of novel composition, which is likely to impact multiple facets of parasite biology, including the clinically important process of bradyzoite differentiation.