Regulation of protamine incorporation and nuclear reorganization during Drosophila spermatogenesis

果蝇精子发生过程中鱼精蛋白掺入和核重组的调节

• 批准号: 10462097
• 负责人: Jun Il Park
• 金额: $ 5.18万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-05-01 至 2023-10-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10462097
• 关键词: Alleles; Architecture; Attention; Biological Models; Biological Process; Cell Nucleolus; Cell Nucleus; Cell Proliferation; Chromatin; Cytology; DNA; DNA-Directed RNA Polymerase; Data; Defect; Development; Dominant-Negative Mutation; Drosophila genus; Drosophila melanogaster; Failure; Fertility; Fertilization; Gene Expression; Genes; Genetic Transcription; Genome; Growth; Histones; Homologous Gene; Human; Lead; Literature; Male Infertility; Mediating; Molecular Biology; Morphogenesis; Nuclear; Ontology; Phenotype; Process; Protamines; Protein Isoforms; Protein Sequence Analysis; Protein Splicing; Proteins; RNA; RNA Splicing; Regulation; Role; Somatic Cell; Sperm Maturation; Spermatids; Spermatocytes; Spermatogenesis; Spermiogenesis; Sterility; Testing; Testis; Y Chromosome; base; chromatin remodeling; clinically relevant; genome integrity; insight; interest; mutant; novel; nucleolin; protein expression; protein protein interaction; sperm cell; transcription factor; transcriptome sequencing

ABSTRACT In many species, sperm maturation requires a process of nuclear compaction, which occurs via dramatic chromatin reorganization from histone to protamine-based architecture. Nuclear compaction is essential for sperm fertility, functioning to enhance hydrodynamics and genomic integrity. While several genes have been identified to be involved in this process, it is unknown how these genes are regulated. Here, we have utilized the Drosophila testis model system to identify Modulo, the Drosophila homolog of Nucleolin, as a key player in sperm nuclear reorganization. In somatic cells, modulo has been shown to be involved in chromatin remodeling, cell proliferation, and morphogenesis. Viable modulo mutant allele that specifically influences testis-specific isoform has been known to be sterile, but the reason underlying this sterility remained unknown. We find that modulo mutants display nuclear compaction defects during late spermatogenesis alongside decreased protamine protein incorporation. Importantly, modulo’s unique phenotype cannot be explained simply by lack of protamine incorporation: compound mutant that lacks protamines Prot A/B and Mst77F causes only mild nuclear deformation rather than complete nuclear decompaction as observed in modulo mutant. Indeed, in addition to decreased incorporation of main protamines in mutant, we have detected increased expression of Mst77Y, a lesser-studied protamine gene with numerous duplications on the Y-chromosome. Thus, we hypothesize that modulo mediates sperm nuclei compaction by coordinating expression of multiple genes that are specifically involved in nuclear compaction during spermiogenesis. To test this hypothesis, we will conclusively establish the cytological defects of modulo mutant by inducing expression of mst77Y in wild-type nuclei and assessing protamine protein expression in whole testis vs. in spermatid nuclei. Additionally, we will investigate transcription and/or splicing as modulo’s mechanism for regulating its target genes. Previous literature has identified modulo to interact with testis-specific transcription factors while protein sequence analysis and gene ontology have predicted modulo as being involved in splicing. Therefore, we further hypothesize that modulo regulates its target genes at the RNA level either by functioning during transcription or splicing. If our hypothesis is true, it would result in the identification of a previously uncharacterized regulatory network governing nuclear reorganization in late stage spermatogenesis and have implications for male infertility.

抽象的 在许多物种中，精子成熟需要一个核压实过程，这是通过 从组蛋白到基于蛋白质的结构的戏剧性染色质重组。核 压实对于精子生育力至关重要，可增强氢动力学和基因组 正直。尽管已经确定了几个基因参与此过程，但尚不清楚 这些基因的调节方式。在这里，我们利用果蝇睾丸模型系统 识别Modulo，果蝇的核苷同系物，是精子核的关键参与者 重组。在体细胞中，Modulo已显示与染色质重塑有关， 细胞增殖和形态发生。特别影响的可行模量突变等位基因 已知睾丸特异性的同工型是无菌的，但是这种不育的原因 仍然未知。我们发现Modulo突变体在最近显示出核压实缺陷 与蛋白质蛋白掺入的降低的精子发生。重要的是，Modulo的 独特的表型不能简单地通过缺乏蛋白质感染来解释：化合物 缺乏protanes蛋白A/B和MST77F的突变体仅引起轻度核变形 如Modulo突变体中观察到的完全核分解。确实，除了 突变体中主prot型的掺入减少，我们检测到表达增加 MST77Y是一种较少研究的蛋白质基因，在Y染色体上具有许多重复。 这是我们假设Modulo通过配位表达介导精子核压实 多种基因在精子发生过程中特别参与核压实的基因。到 检验这一假设，我们将最终确定Modulo突变体的细胞学缺陷 MST77Y在野生型核中诱导的表达并评估蛋白蛋白在 整个睾丸与精子核里的睾丸。此外，我们将研究转录和/或剪接 作为调节其靶基因的机制。以前的文献已经确定了Modulo 与睾丸特异性转录因子相互作用，而蛋白质序列分析和基因 本体学预测Modulo参与了剪接。因此，我们进一步假设 通过在转录过程中发挥作用，可以调节其在RNA水平的靶基因 或剪接。如果我们的假设是正确的，那将导致以前的识别 在晚期核重组的未表征的监管网络 精子发生，对男性不育症有影响。