Prion Protein Fragment PrP-(106–126) Induces Apoptosis via Mitochondrial Disruption in Human Neuronal SH-SY5Y Cells

Abstract

The synthetic peptide PrP-(106 -126) has previously been shown to be neurotoxic. Here, for the first time, we report that it induces apoptosis in the human neuroblastoma cell line SH-SY5Y. The earliest detectable apoptotic event in this system is the rapid depolarization of mitochondrial membranes, occurring immediately upon treatment of cells with PrP-(106 -126). Subsequent to this, cytochrome c release and caspase activation were observed. Caspase inhibitors demonstrated that while the peptide activates caspases they are not an absolute requirement for apoptosis. Parallel to caspase activation, PrP-(106 -126) was also observed to trigger a rise in intracellular calcium through release of mitochondrial calcium stores. This leads to the activation of calpains, another family of proteases. A calpain inhibitor demonstrated that while calpains are activated by the peptide they also are not an absolute requirement for apoptosis. Interestingly a combination of caspase and calpain inhibitors significantly inhibited apoptosis. This illustrates alternative pathways leading to apoptosis via caspases and calpains and that blocking both pathways is required to inhibit apoptosis. These results implicate the mitochondrion as a primary site of action of PrP-(106 -126).

Accumulation of Citrullinated Proteins by Up-Regulated Peptidylarginine Deiminase 2 in Brains of Scrapie-Infected Mice

“…27 PrP 106-126 peptide that shares several characteristics with PrP Sc induces Ca 2ϩ release from endoplasmic reticulum and mitochondria. 28,29 These reports suggest that intracellular Ca 2ϩ is increased in a variety of cells in the brains of scrapie-infected mice. The increase in intracellular Ca 2ϩ during scrapie infection would augment the activity of PAD2, which, in turn, would lead to citrullination of various proteins.…”

Induction of ligand-specific PrPCsignaling in human neuronal cells

“…3 Previous studies have purported to describe the mechanism by which prion, and specifically PrP 106-126, regulates apoptosis. 22,29 Prion related apoptosis studies have proposed a multitude of mechanisms, which are often contradictory, for the induction of apoptosis following prion protein stimulation. 3 The data reported here seems to indicate an apoptotic response via Caspase 3 (Fig.…”

“…A central challenge to understanding the molecular basis of TSEs is determining whether the The PrP 106-126 peptide fragment has been widely utilized as an experimental ligand to model prion effects and this made it an ideal stimulant of PrP C signaling. 17,[20][21][22][23][24] In addition, 6H4 is a well-characterized, commercially-available monoclonal antibody for PrP C . Prion protein specific antibody has been utilized to "activate" PrP C through direct binding for investigations of PrP C function.…”

Calpain-dependent Endoproteolytic Cleavage of PrPSc Modulates Scrapie Prion Propagation

“…Since Ca 2ϩ modulates calpain activity, the observation that scrapie infection induces abnormalities in Ca 2ϩ homeostasis (34) may be significant. Also relevant in this regard are the findings that treatment of human SH-SY5Y neuroblastoma cells with the neurotoxic PrP-(106 -126) peptide resulted in a rapid rise in intracellular calcium and a concomitant increase in calpain activity (35). Interestingly, quinacrine, the most potent substituted tricyclic inhibitor of PrP Sc accumulation (36), blocks Ca 2ϩ channels (37) raising the intriguing possibility that its mode of action may, at least in part, be related to reducing intracellular Ca 2ϩ resulting in lower calpain activity.…”