Muhammad Abubakar, Sarkadi Zsuzsa, van Emden Thomas, Mazumder Agnisrota, Capella Matias, Fekete Gergely, Sreechakram Vishnu N Suma, Al-Sady Bassem, Papp Balázs, Barrales Ramón Ramos, Braun Sigurd
Institute for Genetics, Justus-Liebig-University Giessen, Giessen, Germany.
BioMedical Center (BMC), Division of Physiological Chemistry, Faculty of Medicine, LMU Munich, Planegg-Martinsried, Germany.
bioRxiv. 2024 Feb 15:2024.02.13.579970. doi: 10.1101/2024.02.13.579970.
Heterochromatin plays a critical role in regulating gene expression and maintaining genome integrity. While structural and enzymatic components have been linked to heterochromatin establishment, a comprehensive view of the underlying pathways at diverse heterochromatin domains remains elusive. Here, we developed a systematic approach to identify factors involved in heterochromatin silencing at pericentromeres, subtelomeres, and the silent mating type locus in . Using quantitative measures, iterative genetic screening, and domain-specific heterochromatin reporters, we identified 369 mutants with different degrees of reduced or enhanced silencing. As expected, mutations in the core heterochromatin machinery globally decreased silencing. However, most other mutants exhibited distinct qualitative and quantitative profiles that indicate domain-specific functions. For example, decreased mating type silencing was linked to mutations in heterochromatin maintenance genes, while compromised subtelomere silencing was associated with metabolic pathways. Furthermore, similar phenotypic profiles revealed shared functions for subunits within complexes. We also discovered that the uncharacterized protein Dhm2 plays a crucial role in maintaining constitutive and facultative heterochromatin, while its absence caused phenotypes akin to DNA replication-deficient mutants. Collectively, our systematic approach unveiled a landscape of domain-specific heterochromatin regulators controlling distinct states and identified Dhm2 as a previously unknown factor linked to heterochromatin inheritance and replication fidelity.
异染色质在调节基因表达和维持基因组完整性方面发挥着关键作用。虽然结构和酶成分已与异染色质的建立相关联，但对于不同异染色质结构域潜在途径的全面认识仍然难以捉摸。在这里，我们开发了一种系统方法，以鉴定参与着丝粒周围、亚端粒以及酿酒酵母中沉默交配型位点的异染色质沉默的因子。通过定量测量、迭代遗传筛选和结构域特异性异染色质报告基因，我们鉴定出369个具有不同程度沉默降低或增强的突变体。正如预期的那样，核心异染色质机制中的突变会导致整体沉默降低。然而，大多数其他突变体表现出不同的定性和定量特征，表明其具有结构域特异性功能。例如，交配型沉默的降低与异染色质维持基因中的突变有关，而亚端粒沉默的受损与代谢途径相关。此外，相似的表型特征揭示了复合物内亚基的共同功能。我们还发现，未表征的蛋白质Dhm2在维持组成型和兼性异染色质方面起着关键作用，而其缺失会导致类似于DNA复制缺陷型突变体的表型。总体而言，我们的系统方法揭示了控制不同状态的结构域特异性异染色质调节因子的全貌，并确定Dhm2是一个与异染色质遗传和复制保真度相关的先前未知的因子。
