Zhao Wenda, Eid Shehab, Sackmann Chris, Williams Declan, Wang Xinzhu, Ouyang Yunqing, Zerbes Thomas, Schmitt-Ulms Gerold
Tanz Centre for Research in Neurodegenerative Diseases, University of Toronto, Toronto, ON, Canada.
Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, ON, Canada.
Acta Neuropathol. 2025 Jun 23;149(1):68. doi: 10.1007/s00401-025-02906-2.
Our recent work on the prion protein and Na, K-ATPases (NKAs) led us to revisit data from over 50 years ago, which suggested similarities between vacuolation phenotypes in rodents poisoned with cardiac glycosides (CGs) and spongiform degeneration in prion disease. At that time, this hypothesis was dismissed because the vacuolation observed in prion diseases affects neurons, whereas CG poisoning in rodent brains led to swellings of the endoplasmic reticulum (ER) in astrocytes. We speculated that this difference might be specific to rodents and document here that the vacuolation shifts to neurons in mice expressing a humanized NKA α1 subunit. Next, we investigated the molecular mechanisms that could cause similar ER vacuolation in human cells in vitro. We found that certain stressors-such as overexpression of NKA α subunits and exposure to specific toxins known to trigger the unfolded protein response-can induce a phenotype characterized by profound ER dilation that is most strikingly observed for the perinuclear space (PNS). The ion imbalance typically caused by functional NKAs does not contribute to this phenotype. In fact, it can occur even with the overexpression of catalytically inactive NKAs. Several lines of evidence, generated with pharmacological agents, ion-specific dyes, antagonists, and truncated expression constructs, suggest that a calcium leak channel in the ER, known as transient receptor potential vanilloid 2 (TRPV2), plays a role in this ER and PNS dilation. Additionally, we observed that the formation of these vacuoles coincides with a decrease in steady-state levels of the lipid kinase PIKFYVE, which is recognized for its role in endolysosomal fission and fusion processes. Finally, we found evidence of vacuoles in cryosectioned brains of prion-infected mice that can be filled with a fluorescent marker targeted at the ER and PNS. This raises the possibility that this vacuolation phenomenon contributes to spongiform degeneration seen in prion diseases.
我们最近关于朊病毒蛋白和钠钾ATP酶（NKAs）的研究工作，促使我们重新审视50多年前的数据，这些数据表明，用强心苷（CGs）中毒的啮齿动物的空泡化表型与朊病毒病中的海绵状变性之间存在相似性。当时，这个假设被否定了，因为在朊病毒病中观察到的空泡化影响神经元，而啮齿动物大脑中的CG中毒导致星形胶质细胞内质网（ER）肿胀。我们推测这种差异可能是啮齿动物特有的，并在此记录，在表达人源化NKA α1亚基的小鼠中，空泡化转移到了神经元。接下来，我们研究了在体外人类细胞中可能导致类似ER空泡化的分子机制。我们发现，某些应激源，如NKA α亚基的过表达以及暴露于已知能触发未折叠蛋白反应的特定毒素，可诱导一种以ER显著扩张为特征的表型，这种扩张在核周间隙（PNS）最为明显。功能性NKAs通常引起的离子失衡对这种表型没有影响。事实上，即使是催化无活性的NKAs过表达也可能出现这种情况。用药物、离子特异性染料、拮抗剂和截短的表达构建体产生的几条证据表明，ER中的一种钙泄漏通道，即瞬时受体电位香草酸受体2（TRPV2），在这种ER和PNS扩张中起作用。此外，我们观察到这些空泡的形成与脂质激酶PIKFYVE稳态水平的降低同时发生，PIKFYVE因其在内溶酶体分裂和融合过程中的作用而被认可。最后，我们在朊病毒感染小鼠的冷冻切片大脑中发现了空泡的证据，这些空泡可以被一种针对ER和PNS的荧光标记物填充。这增加了这种空泡化现象导致朊病毒病中海绵状变性的可能性。
