Biology subjects

Schmitt-Ulms, G.

Publications and source records attributed to Schmitt-Ulms, G..

5 recordsLinked to original sources

bioRxiv2024

Transient receptor potential vanilloid channel 2 contributes to multi-modal endoplasmic reticulum and perinuclear space dilations that can also be observed in prion-infected mice

Our recent work on the prion protein and Na+,K+-ATPases (NKAs) led us to revisit data from over 50 years ago, which suggested a similarity 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 1 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 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 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 cryo-sectioned brains of prion-infected mice that can be filled with a fluorescent marker targeted to the ER and PNS. This raises the possibility that this vacuolation phenomenon contributes to spongiform degeneration seen in prion diseases.

neuroscience↗

bioRxiv2024

The Brain Interactome of a Permissive Prion Replication Substrate

Bank voles are susceptible to prion strains from many different species, yet the molecular mechanisms underlying the ability of bank vole prion protein (BVPrP) to function as a universal prion acceptor remain unclear. Potential differences in molecular environments and protein interaction networks on the cell surface of brain cells may contribute to BVPrPs unusual behavior. To test this hypothesis, we generated knock-in mice that express physiological levels of BVPrP (M109 isoform) and employed mass spectrometry to compare the interactomes of mouse (Mo) PrP and BVPrP following mild in vivo crosslinking of brain tissue. Substantial overlap was observed between the top interactors for BVPrP and MoPrP, with established PrP-interactors such as neural cell adhesion molecules, subunits of Na+/K+-ATPases, and contactin-1 being equally present in the two interactomes. We conclude that the molecular environments of BVPrP and MoPrP in the brains of mice are very similar. This suggests that the unorthodox properties of BVPrP are unlikely to be mediated by differential interactions with other proteins.

neuroscience↗

bioRxiv2024

The Molecular Determinants of a Universal Prion Acceptor

In prion diseases, the species barrier limits the transmission of prions from one species to another. However, cross-species prion transmission is remarkably efficient in bank voles, and this phenomenon can be recapitulated in mice by expression of the bank vole prion protein (BVPrP). The molecular determinants of BVPrPs ability to function as a universal or near-universal acceptor for prions remain incompletely defined. Building on our finding that cultured cells expressing BVPrP can replicate both mouse and hamster prion strains, we conducted a systematic analysis to identify key residues in BVPrP that permit cross-species prion replication. Consistent with previous findings, we demonstrate that residues N155 and N170 of BVPrP, which are absent in mouse PrP but present in hamster PrP, are critical for cross-species prion replication. Additionally, BVPrP residues V112, I139, and M205, which are absent in hamster PrP but present in mouse PrP, are also required to enable replication of both mouse and hamster prions. Unexpectedly, we found that residues E227 and S230 near the C-terminus of BVPrP severely restrict the accumulation of prions following cross-species prion challenge, suggesting that they may have evolved to counteract the inherent propensity of BVPrP to misfold. PrP variants with an enhanced ability to replicate both mouse and hamster prions displayed accelerated spontaneous aggregation kinetics in vitro. These findings suggest that BVPrPs unusual properties are governed by a key set of amino acids and that the enhanced misfolding propensity of BVPrP may enable cross-species prion replication.

neuroscience↗

bioRxiv2022

Somatostatin slows Aβ plaque deposition in aged APPNL-F/NL-F mice by blocking Aβ aggregation in a neprilysin-independent manner

The molecular underpinnings that govern the endoproteolytic release of the amyloid beta peptide (A{beta}) from the amyloid precursor protein (APP) are now quite well understood. The same cannot be said for the events that precipitate the aggregation and amyloid deposition of A{beta} in Alzheimers disease (AD). The 14-amino-acid cyclic neuroendocrine peptide somatostatin (SST-14) has long been thought of as playing a role, foremost by controlling the expression of the A{beta} clearing enzyme neprilysin, and more recently by directly interacting with A{beta} oligomers. Missing have been in vivo data in a relevant A{beta} amyloidosis model. Here we addressed this shortcoming by crossing AppNL-F/NL-F mice with Sst-deficient mice of identical genetic background to assess if and how the presence of Sst influences key pathological hallmarks of A{beta} amyloidosis that develop in AppNL-F/NL-F mice after 10 months of age. Surprisingly, we found that Sst had no influence on whole brain neprilysin transcript, protein or activity levels, an observation that cannot be accounted for by a compensatory upregulation of the Sst paralog, cortistatin (Cort), that we observed in 15-month-old Sst-deficient mice. The absence of Sst did lead to a subtle but significant increase in the density of cortical A{beta} amyloid plaques. Follow-on western blot analyses of whole brain extracts indicated that Sst interferes with early steps of A{beta} assembly that manifest in Sst null brains through the appearance of SDS-stable smears of 55- 150 kDa. As expected, no effect of Sst on tau steady-state levels or its phosphorylation were observed. Results from this study are easier reconciled with an emerging body of data that point toward Sst affecting A{beta} amyloid plaque formation through direct interference with A{beta} aggregation rather than through its effects on neprilysin expression.

neuroscience↗

bioRxiv2022

Identification of a cardiac glycoside exhibiting favorable brain bioavailability and potency for reducing levels of the cellular prion protein

Several strands of investigation have established that a reduction in the levels of the cellular prion protein (PrPC) is a promising avenue for the treatment of prion diseases. We recently described an indirect approach for reducing PrPC levels that targets Na,K-ATPases (NKAs) with cardiac glycosides (CGs), causing cells to respond with the degradation of these pumps and nearby molecules, including PrPC. Because the therapeutic window of widely used CGs is narrow and their brain bioavailability is low, we set out to identify a CG with improved pharmacological properties for this indication. Starting with the CG known as oleandrin, we combined in silico modeling of CG binding poses within human NKA folds, CG structure-activity relationship (SAR) data, and predicted blood-brain barrier (BBB) penetrance scores to identify CG derivatives with improved characteristics. Focusing on C4-dehydro-oleandrin as a chemically accessible shortlisted CG derivative, we show that it reaches four times higher levels in the brain than in the heart one day after subcutaneous administration, exhibits promising pharmacological properties, and suppresses steady-state PrPC levels by 84% in immortalized human cells that have been differentiated to acquire neural or astrocytic characteristics. Finally, we validate that the mechanism of action of this approach for reducing cell surface PrPC levels requires C4-dehydro-oleandrin to engage with its cognate binding pocket within the NKA subunit. The improved brain bioavailability of C4-dehydro-oleandrin, combined with its relatively low toxicity, make this compound an attractive lead for brain CG indications and recommends its further exploration for the treatment of prion diseases. AUTHOR SUMMARYPrion diseases are fatal neurodegenerative diseases for which there is no effective treatment. An abundance of data indicates that reducing the levels of a specific protein, termed the cellular prion protein (PrPC), would not only be safe but would delay disease onset and extend prion disease survival. This project builds on our recent discovery that PrPC binds to NKAs, specific cellular transport proteins that use energy to electrify cellular membranes by pumping charged potassium and sodium metals in and out of cells. We showed that targeting NKAs with their natural inhibitors, cardiac glycosides (CGs), causes brain cells to internalize and degrade NKAs, and that PrPC, on account of residing next to NKAs, gets co-degraded. Natural CGs act primarily on the heart. Here, we used computational modeling to identify a CG, termed KDC203, that is predicted to have favorable characteristics for brain applications. We show that KDC203 reduces PrPC levels by 84% in immortalized human brain-like cells grown in the dish. Moreover, we show that KDC203 exhibits relatively low toxicity, predominantly targets the brain when subcutaneously injected into mice, and has other promising pharmacological characteristics that recommend it for further evaluation for the treatment of prion diseases.

neuroscience↗