Rucli Sofia, Descostes Nicolas, Ermakova Yulia, Chitnavis Urvashi, Couturier Jeanne, Boskovic Ana, Boulard Matthieu
Epigenetics & Neurobiology Unit, European Molecular Biology Laboratory, EMBL Rome, Rome, Italy.
Collaboration for a joint PhD degree between EMBL and Heidelberg University, Heidelberg, Germany.
Genome Biol. 2025 Mar 24;26(1):69. doi: 10.1186/s13059-025-03537-2.
How reversible glycosylation of DNA-bound proteins acts on transcription remains scarcely understood. O-linked β-N-acetylglucosamine (O-GlcNAc) is the only known form of glycosylation modifying nuclear proteins, including RNA polymerase II (RNA Pol II) and many transcription factors. Yet, the regulatory function of the O-GlcNAc modification in mammalian chromatin remains unclear.
Here, we combine genome-wide profiling of O-GlcNAc-modified proteins with perturbations of intracellular glycosylation, RNA Pol II-degron, and super-resolution microscopy. Genomic profiling of O-GlcNAc-modified proteins shows a non-random distribution across the genome, with high densities in heterochromatin regions as well as on actively transcribed gene promoters. Large-scale intersection of the O-GlcNAc signal at promoters with public ChIP-seq datasets identifies a high overlap with RNA Pol II and specific cofactors. Knockdown of O-GlcNAc Transferase (Ogt) shows that most direct target genes are downregulated, supporting a global positive role of O-GlcNAc on the transcription of cellular genes. Rapid degradation of RNA Pol II results in the decrease of the O-GlcNAc levels at promoters encoding transcription factors and DNA modifying enzymes. RNA Pol II depletion also unexpectedly causes an increase of O-GlcNAc levels at a set of promoters encoding for the transcription machinery.
This study provides a deconvoluted genomic profiling of O-GlcNAc-modified proteins in murine and human cells. Perturbations of O-GlcNAc or RNA Pol II uncover a context-specific reciprocal functional interplay between the transcription machinery and the O-GlcNAc modification.
DNA结合蛋白的可逆糖基化如何作用于转录仍鲜为人知。O-连接的β-N-乙酰葡糖胺（O-GlcNAc）是已知的唯一修饰核蛋白的糖基化形式，包括RNA聚合酶II（RNA Pol II）和许多转录因子。然而，O-GlcNAc修饰在哺乳动物染色质中的调节功能仍不清楚。
在这里，我们将O-GlcNAc修饰蛋白的全基因组分析与细胞内糖基化、RNA Pol II降解子和超分辨率显微镜的扰动相结合。O-GlcNAc修饰蛋白的基因组分析显示其在全基因组中呈非随机分布，在异染色质区域以及活跃转录的基因启动子上密度较高。启动子处O-GlcNAc信号与公共ChIP-seq数据集的大规模交叉分析表明，它与RNA Pol II和特定辅因子高度重叠。敲低O-连接N-乙酰葡糖胺转移酶（Ogt）表明，大多数直接靶基因被下调，这支持了O-GlcNAc对细胞基因转录的整体正向作用。RNA Pol II的快速降解导致编码转录因子和DNA修饰酶的启动子处O-GlcNAc水平降低。RNA Pol II的缺失还意外地导致一组编码转录机制的启动子处O-GlcNAc水平升高。
本研究提供了小鼠和人类细胞中O-GlcNAc修饰蛋白的详细基因组分析。O-GlcNAc或RNA Pol II的扰动揭示了转录机制与O-GlcNAc修饰之间特定背景下的相互功能作用。
