Defining the regulation of repressive epigenetic marks by the ubiquitin-conjugating enzyme UBE2K and its impact on cell fate decisions and organismal aging

生物体的发育和生存与其维持细胞蛋白质组完整性的能力有关。人胚胎干细胞（hESCs）在培养中是不朽的。这种能力要求避免任何蛋白质稳态失衡（proteostasis），否则会损害hESC的身份。我们发现hESCs表现出UBE2K水平的增加，UBE2K是一种调节蛋白质泛素化的E2酶。值得注意的是，UBE2K是hESCs神经元分化所必需的。有趣的是，我们发现UBE2K与组蛋白H3.3结合并调节其泛素化。此外，UBE2K缺失通过诱导H3K9的三甲基化改变hESCs的表观遗传格局。尽管组蛋白编码在染色质和基因调控中起着关键作用，但调节hESCs独特染色质模式的分子机制在很大程度上仍然未知。在这里，我们将试图定义UBE2K对H3K9三甲基化的调节如何影响hESCs的神经发生。由于细胞重编程涉及染色质的显著重组，我们将研究UBE2K水平的变化是否会改变体细胞的H3K9me3景观并促进细胞重编程。鉴于组蛋白修饰在细胞功能中的重要性，我们将确定UBE2K调节H3K9三甲基化的分子机制。表观遗传变化不仅是发育的决定因素，也是衰老的标志。H3K9me3表观遗传修饰在衰老过程中下调。最重要的是，表观遗传景观可以通过调节来延长秀丽隐杆线虫等模式生物的寿命。这些发现突出了表观遗传失调作为哺乳动物衰老驱动因素的作用。因此，定义与年龄相关的表观遗传标记的新调节剂可能为衰老研究提供进一步的见解。有趣的是，我们观察到UBE2K的缺失会触发秀丽隐杆线虫的H3K9me3水平。我们将研究UBE2K的调节是否足以延长寿命并改善与生物衰老相关的H3K9me3的下调。最后，我们将进行RNAi筛选，以确定UBE2K如何在体内调节H3K9的三甲基化。因此，我们的实验可以定义衰老的两个初步标志（即蛋白质平衡的丧失，表观遗传变化）之间的相互联系，并揭示减缓衰老过程的新机制。综上所述，我们的研究计划可以在干细胞研究、细胞治疗、衰老和与年龄相关的疾病等几个领域产生重大影响。

The development and survival of an organism are linked to its ability to maintain the integrity of the cellular proteome. Human embryonic stem cells (hESCs) are immortal in culture. This capacity demands avoidance of any imbalance in protein homeostasis (proteostasis) that would otherwise compromise hESC identity. We have found that hESCs exhibit increase levels of UBE2K, an E2 enzyme that regulates protein ubiquitination. Notably, UBE2K is required for neuronal differentiation of hESCs. Interestingly, we found that UBE2K binds to histone H3.3 and regulates its ubiquitination. Moreover, loss of UBE2K changes the epigenetic landscape of hESCs by inducing trimethylation of H3K9. Although the histone code has a critical role in chromatin and gene regulation, the molecular mechanisms that modulate the unique chromatin pattern of hESCs remain largely unknown. Here we will seek to define how modulation of H3K9 trimethylation by UBE2K impinges upon neurogenesis of hESCs. Since cell reprogramming involves a marked reorganization of chromatin, we will examine whether changes in UBE2K levels alter the H3K9me3 landscape of somatic cells and facilitate cell reprogramming. Given the importance of histone modifications in cell function, we will define the molecular mechanisms by which UBE2K regulates H3K9 trimethylation. Epigenetic changes are not only a determinant of development but also a hallmark of aging. H3K9me3 epigenetic modifications are downregulated during the aging process. Most importantly, the epigenetic landscape can be modulated to extend lifespan in model organisms such as C. elegans. These findings highlight the role of epigenetic dysregulation as a driver of mammalian aging. Thus, defining novel modulators of age-associated epigenetic marks may provide further insights into aging research. Interestingly, we have observed that loss of UBE2K triggers H3K9me3 levels in C. elegans. We will examine whether modulation of UBE2K is sufficient to extend longevity and ameliorate the downregulation of H3K9me3 associated to biological aging. Finally, we will perform a RNAi screen to define how UBE2K modulates trimethylation of H3K9 in vivo. Thus, our experiments can define the interconnectedness between two tentative hallmarks of aging (i.e., loss of proteostasis, epigenetic changes) and uncover novel mechanisms to slow down the aging process. Taken together, our research proposal can have a big impact in several fields such as stem cell research, cell therapy, aging and age-related diseases.