Neurobiological Basis of Neuronal Survival

在本财政年度中，我们发现ER-线粒体界面对于通过利用抗凋亡蛋白Bcl-xL增强ER和线粒体之间的Ca 2+信号传导来增加细胞生物能量是重要的。摘要如下。Bcl-2家族与细胞定位相关的非凋亡功能正在出现。 Bcl-xL，特别是，已被假设为刺激生物能量通过调节钙离子信号在ER。 然而，Bcl-xL通过3型肌醇1，4，5-三磷酸受体（IP 3R 3）特异性地在直接ER到线粒体Ca 2+转移中的功能先前尚未被研究。 在这里，我们发现CHO细胞中的Bcl-xL的显著部分以簇状形式存在于ER-E2-on界面或MAM（ER-associated ER membrane）处，在那里它与IP 3R 3相互作用以增强线粒体Ca 2+信号传导并改变生物能量学。 特别是，Bcl-xL增强线粒体Ca 2+瞬变毒胡萝卜素（Tg）诱导的ER Ca 2+耗竭，同时伴随着减少胞质Ca 2+释放。 因此，在Bcl-xL过表达细胞中观察到增强的TCA循环活性，其在长时间Tg损伤后进一步增加。 这种作用是Bcl-xL易位到MAM的结果，这是在非凋亡应激事件期间首次观察到Bcl-xL易位。 我们的研究结果表明，MAM是一个关键的细胞信号连接，Bcl-xL可能已经进化到包括多个功能。 一个作用是促进细胞代谢以促进正常细胞功能，第二个任务是通过增加细胞的生物能量学能力来防止细胞凋亡的启动。

In this fiscal year we have found that the ER-mitochondrion interface is important for increasing cellular bioenergetics by utilizing an anti-apoptotic protein Bcl-xL to enhancing the Ca2+ signaling between the ER and mitochondria. The synopsis is as follows. Non-apoptotic functions related to cellular localization are emerging for the Bcl-2 family. Bcl-xL, in particular, has been hypothesized to stimulate bioenergetics by regulating Ca2+ signaling at the ER. However, the functioning of Bcl-xL specifically in direct ER to mitochondrial Ca2+ transfer, through type 3 inositol 1,4,5-trisphosphate receptors (IP3R3), has not previously been investigated. Here we found a significant fraction of Bcl-xL in CHO cells exists in clustered form at the ER-mitochondrion interface or MAM (mitochondria-associated ER membrane) where it interacts with IP3R3 to enhance mitochondrial Ca2+ signaling and alter bioenergetics. Particularly, Bcl-xL enhanced mitochondrial Ca2+ transients upon thapsigargin (Tg)-induced ER Ca2+ depletion, while concomitantly reducing cytosolic Ca2+ release. Consequently, enhanced TCA cycle activity was observed in Bcl-xL overexpressing cells, which was further increased after prolonged Tg insult. This action is the result of Bcl-xL translocation to the MAM, which is the first known observation of Bcl-xL translocating during a non-apoptotic stress event. Our results suggest the MAM is a critical cell-signaling junction whereby Bcl-xL might have evolved to include multiple functionalities. One role being to facilitate cellular metabolism to promote normal cell functioning, and the second task being to prevent the initiation of apoptosis by increasing the bioenergetics capacity of the cell.