Factors that modulate cellular homeostasis to overcome replicative stress in aging

我们已经研究了不同化合物对单个复制叉子在强阻塞时停滞的频率的活性。下面总结了一些最新的结果： 我们发现，用于癌症治疗的聚腺苷二磷酸核糖合成的抑制剂会增加单叉失速的频率。相反，抑制聚ADP核糖的降解减少了单失速叉子的频率。 一种化合物可以增强蛋白质伴侣的活性，从而改善蛋白质的正确折叠，还可以减少单叉失速事件的频率。 我们的结果表明，一些小分子效应器可以在基因组水平上影响事件，从而降低基因组不稳定和应激诱导事件的频率。我们计划测试更多的化合物，并将跟踪那些在测试中显示活性的化合物。 我们还表明，该试验可以在小鼠造血干细胞(HSCs)中进行。这使得研究“抗衰老”化合物对来自年轻和老年捐赠者的干细胞抵抗复制压力的能力的影响成为可能。我们发现，与年轻捐献者相比，老年造血干细胞中单叉失速事件的频率上升。此外，我们还研究了单叉口失速事件增加的年龄。值得注意的是，这出现在相当于小鼠的中年。因此，在这个小鼠干细胞模型中，细胞克服复制障碍的能力在它们达到“老化”状态之前就已经下降了。 最近，我们发现有两种形式的复制体遇到了障碍。一种在S期早期和常染色质中活跃，另一种在S晚期和异染色质中活跃。我们现在的问题是，单叉失速事件的增加是发生在基因组的一个区域还是另一个区域，还是两个区域。

We have examined the activity of different compounds on the frequency of single replication forks stalled at a strong block. Some recent results are summarized here: We have found that inhibitors of poly ADP ribose synthesis, which are used in cancer therapy, increase the frequency of single fork stalling. In contrast, inhibition of the degradation of poly ADP ribose reduces the frequency of single stalled forks. A compound that enhances the activity of a protein chaperone that improves correct protein folding also reduces the frequency of single fork stalling events. Our results suggest that some small molecule effectors can influence events at the level of the genome, such that the frequency of a genome destabilizing and stress inducing event is reduced. We plan to test additional compounds, and will follow those that show activity in the assay. We have also shown that assay can be performed in murine hematopoietic stem cells (HSCs). This makes it possible to examine the influence of "anti aging" compounds on the competence of stem cells, from young and old donors, to resist replication stress. We find that the frequency of single fork stalling events rises in HSCs from old as compared to young donors. Furthermore, we have examined the age at which the increase in single fork stalling events can be observed. Remarkably this appears at an equivalent to middle age in mice. Consequently it appears that in this mouse stem cell model cellular competence to overcome replication blocks declines well before they reach an "aged" status. Recently, we found that there are two forms of the replisomes that encounter the block. One is active in early S phase and in euchromatin, the other in late S phase and in heterochromatin. We are now asking if the increase in single fork stalling events occurs in one or the other or both regions of the genome.