Transposable element mobilization during spermatogenesis in Drosophila

果蝇精子发生过程中转座元件的动员

基本信息

批准号：
10747733
负责人：
Lauren Ann Tracy
金额：
$ 4.03万
依托单位：
DUKE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-09-01 至 2025-08-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10747733
关键词：
Binding Sites Cells Circular DNA Code Complementary DNA DNA DNA Ligases DNA Repair Pathway DNA Transposable Elements DNA-Directed DNA Polymerase Development Disease Drosophila genus Drosophila melanogaster Elements Event Factor VIII Female Future Generations Genes Genetic Code Genome Germ Cells Hemophilia A Human Genome Infertility Integration Host Factors Knowledge Length Life Cycle Stages Location Modeling Monitor Mutation Nomads Oocytes Oogenesis Organism Pathway interactions Placentation Process Production RNA RNA Interference RNA-Directed DNA Polymerase Regulatory Element Reporter Repression Retrotransposon Single-Stranded DNA Small RNA Sperm Maturation Spermatocytes Spermatogenesis Sterility Study models Technology Testing Testis Time Writing XRCC1 gene Y Chromosome cell type cost egg experimental study fly gene regulatory network improved male member nanopore piRNA preference recruit sperm cell syncytin transcription factor

项目摘要

Abstract Transposons are mobile pieces of DNA that comprise significant proportions of eukaryotic genomes, including around 45% of the human genome and 30% of the fly genome. While most transposons have lost the ability to jump into new locations in the genome, several in each organism maintain the ability to mobilize. New transposon insertions are particularly consequential if they are generated in germline cells. Germline insertions are subsequently present in every cell of the organism they develop into and can be passed on to future generations. Thus, germline cells employ multilayered mechanisms to repress transposons. These repressive mechanisms are essential to largely suppress transposon for development of germline cells into mature eggs and sperm because transposon activity can contribute to sterility. Since these repressive mechanisms like the piRNA pathway render transposition events so rare, they are difficult to detect and our understanding of how transposons mobilize in the germline remains incomplete. Namely, the host factors that transposons employ to generate new insertions and the time point when they make new insertions during sperm maturation are undefined. To find potentially active transposons in the male germline, I have sequenced circular DNA from Drosophila testes with the piRNA pathway depleted. Disrupting the piRNA pathway allowed for transposon activation and revealed that the nomad transposon generates the most circular DNA in the testes. Circular DNA is a transposition intermediate of LTR retrotransposons and its presence can indicate the likelihood of a transposon being able to still make new insertions. With the knowledge that nomad is the most active transposon in the male germline, I propose to use this transposon to study how transposons mobilize in the germline. I hypothesize that transposons utilize host factors from the alt-EJ DNA repair pathway to achieve stage specific mobilization during spermatogenesis. I will utilize circular nomad DNA as a readout to find host factors required for its production and transposition reporters for nomad to precisely identify when it generates new insertions.

抽象的转座子是移动DNA的移动片段，包括大量的真核基因组，包括大约45％的人类基因组和30％的蝇基因组。虽然大多数转座都失去了跳入基因组的新位置，每个生物体中的几个都保持动员的能力。新的转座子如果插入是在种系细胞中产生的，则尤其是结果。种系插入是随后，它们会出现在它们发展到的生物的每个细胞中，并可以传递给子孙后代。因此，种系细胞采用多层机制来抑制转座子。这些压制机制对于在很大程度上抑制转座子以将种系细胞发展成成熟的卵和精子至关重要因为转座子活性会导致不育。由于这些抑制性机制，例如pirna 途径使转置事件如此罕见，它们很难检测到我们对转座动员在种系中仍然不完整。也就是说，转座子采用的宿主因素生成新的插入和在精子成熟期间进行新插入的时间点是不明确的。为了在雄性种系中找到潜在的活性转座子，我从果蝇睾丸与piRNA途径枯竭。破坏piRNA途径允许转座子激活并揭示了Nomad Transposon在睾丸中产生最多的圆形DNA。圆形DNA 是LTR逆转座子的中间体的换位，其存在可能表明A的可能性转座子仍然能够进行新的插入。知道游牧是最活跃的转座子的知识在男性种系中，我建议使用该转座子研究转座子如何动员种系。我假设转座子利用来自Alt-EJ DNA修复途径的宿主因子来实现特定阶段精子发生过程中的动员。我将利用圆形游牧DNA作为读数来查找所需的主机因素为了生产和换位记者，以供游牧民族确切确定何时生成新的插入。