Kokity Lilla, Czimmerer Zsolt, Benyhe-Kis Bernadett, Poscher Anna, Belai Emese, Steinbach Gábor, Lipinszki Zoltan, Pirity Melinda Katalin
Biological Research Centre, Institute of Genetics, Hungarian Research Network, Szeged, Hungary.
Faculty of Science and Informatics, Doctoral School in Biology, University of Szeged, Szeged, Hungary.
Front Cell Dev Biol. 2024 Nov 29;12:1498346. doi: 10.3389/fcell.2024.1498346. eCollection 2024.
Early embryonic development is a complex process where undifferentiated cells lose their pluripotency and start to gastrulate. During gastrulation, three germ layers form, giving rise to different cell lineages and organs. This process is regulated by transcription factors and epigenetic regulators, including non-canonical polycomb repressive complex 1s (ncPRC1s). Previously, we reported that ncPRC1-member RYBP (RING1 and YY1 binding protein) is crucial for embryonic implantation and cardiac lineage commitment in mice. However, the role of RYBP in gastrulation and mesoderm formation has not yet been defined. In this study, we used 2D and 3D model systems, to analyze the role of RYBP in mesoderm formation. First, we showed that cardiac and endothelial progenitors-both derived from mesoderm-are underrepresented in the cardiac colonies. In the absence of RYBP, the formation of major germ layers was also disrupted, and the expression of mesoderm- ( and ) and endoderm-specific (, ) genes was significantly downregulated. Using 3D embryoid bodies as gastrulation models, we showed that RYBP can co-localize with mesoderm lineage marker protein BRACHYURY and endoderm marker protein GATA4 and both proteins. In mutants, both proteins were detected at low levels and showed altered distribution. Additionally, we compared our results to available single-cell transcriptomes and showed that and co-expressed in the primitive streak and six mesodermal clusters. Since caudal mesoderm exhibited one of the strongest co-expressions, we tested axial elongation in and gastruloids. In the absence of RYBP, gastruloids exhibited shortened tails and low BRACHYURY levels in the tailbud. Finally, we identified BRACHYURY as a novel binding partner of RYBP and presented evidence of possible cooperative function during mesoderm formation and axial elongation. Together, our results demonstrate the previously unknown role of RYBP in mesoderm formation. We believe our findings will contribute to better understanding of the highly conserved process of gastrulation.
早期胚胎发育是一个复杂的过程，在此过程中未分化的细胞失去其多能性并开始原肠胚形成。在原肠胚形成期间，三个胚层形成，产生不同的细胞谱系和器官。这个过程由转录因子和表观遗传调节因子调控，包括非经典多梳抑制复合物1（ncPRC1）。此前，我们报道ncPRC1成员RYBP（RING1和YY1结合蛋白）对小鼠胚胎着床和心脏谱系定向分化至关重要。然而，RYBP在原肠胚形成和中胚层形成中的作用尚未明确。在本研究中，我们使用二维和三维模型系统来分析RYBP在中胚层形成中的作用。首先，我们发现均源自中胚层的心脏和内皮祖细胞在心脏集落中的占比不足。在缺乏RYBP的情况下，主要胚层的形成也受到破坏，中胚层特异性（和）以及内胚层特异性（、）基因的表达显著下调。使用三维胚状体作为原肠胚形成模型，我们发现RYBP可与中胚层谱系标记蛋白短尾相关转录因子和内胚层标记蛋白GATA4以及这两种蛋白共定位。在突变体中，这两种蛋白的检测水平较低且分布改变。此外，我们将我们的结果与现有的单细胞转录组进行比较，发现和在原条和六个中胚层簇中共表达。由于尾侧中胚层表现出最强的共表达之一，我们在和类原肠胚中测试了轴向伸长。在缺乏RYBP的情况下，类原肠胚的尾巴缩短且尾芽中的短尾相关转录因子水平较低。最后，我们确定短尾相关转录因子是RYBP的一种新的结合伙伴，并提供了在中胚层形成和轴向伸长过程中可能存在协同功能的证据。总之，我们的结果证明了RYBP在中胚层形成中此前未知的作用。我们相信我们的发现将有助于更好地理解高度保守的原肠胚形成过程。
