Understanding genomic-context specific deposition and function of H3.V and H4.V histone variants in Trypanosoma brucei

在许多生物体中的研究表明，用组蛋白变体取代规范的组蛋白可以影响DNA的可及性。因此，组蛋白变异体沉积代表了一种调节DNA模板过程的手段，如转录、复制或DNA修复。大量不同的组蛋白变体和复杂的全基因组染色质模式减缓了对导致组蛋白变体在特定基因组位点沉积的机制以及基因组环境在确定组蛋白变体功能中的作用的全面了解的进展。为了克服这些障碍，我们建议研究组蛋白变体的沉积和功能。组蛋白变体是一种单细胞真核生物，具有许多特征，使其非常适合于研究组蛋白变体生物学的基本机制。布氏毛滴虫的每个典型组蛋白只有一个变异体，并且由于基因排列成多顺反子转录单位(PTU)，只有相对较少的~200个转录起始位点(TSS)和转录终止位点(TTS)。此外，我们还发现，在布氏锥虫中，四种不同的组蛋白变异体，即H_2A.Z、H_2.V、H_3.V和H_4V，在全基因组中表现出非常不同的分布，因此很容易发现变异分布中的微小变化。最近，我们发现H3.V的缺失导致转录终止缺陷和基因组结构的改变。此外，我们观察到H3.V和H4.V的同时缺失强烈地影响了特定基因组位点的染色质紧致，并通过重组增加了抗原的切换。根据这些发现，我们推测H3.V具有不同的生物学功能，取决于其基因组背景。我们怀疑它沉积在双链断裂的位置，以帮助DNA修复，并在PTU的末端，以确保适当的转录终止。为了验证这些假设，我们将调查H3.V是如何以及为什么针对特定的基因组座位，并确定基因组环境是否影响它在局部染色质结构和更高级别基因组组织中所起的作用。

Studies in numerous organisms have shown that the replacement of canonical histones with histone variants can affect DNA accessibility. Thus, histone variant deposition represents a means to regulate any DNA-templates process, such as transcription, replication or DNA repair. A large number of different histone variants and complex genome-wide chromatin patterns have slowed progress towards a comprehensive understanding of the mechanisms leading to the deposition of histone variants at specific genomic loci and the role of the genomic context in determining the function of histone variants. Thus, for most histone variants it is not understood how and why they are targeted to specific genomic locus and whether they can have different functions depending on their genomic context.To overcome these hurdles, we propose to study histone variant deposition and function in T. brucei, a unicellular eukaryote with many features that make it ideally-suited to investigate basic mechanisms of histone variant biology. T. brucei possess only one variant for each canonical histone and, due to the arrangement of genes into polycistronic transcription units (PTUs), a relatively small number of only ~200 transcription start sites (TSSs) and transcription termination sites (TTSs). In addition, we have shown that in T. brucei the four different histone variants, H2A.Z, H2.V, H3.V and H4.V, exhibit a very distinct genome-wide distribution, making it easy to detect small changes in variant distribution.Recently, we have found that loss of H3.V leads to a defect in transcription termination and changes in genome organization. In addition, we have observed that concurrent deletion of H3.V and H4.V strongly affects chromatin compaction at specific genomic loci and increases antigen switching by recombination.Based on these findings we hypothesize that, depending on its genomic context, H3.V has different biological functions. We suspect that it is deposited at sites of double stranded break to aid in DNA repair and at the end of PTUs to ensure proper transcription termination. To test these hypotheses, we will investigate how and why H3.V is targeted to specific genomic loci and determine whether the genomic context affects the role it plays in local chromatin structure and higher order genome organization.