Prion Protein Protects Human Neurons against Bax-mediated Apoptosis

Abstract

The function of the cellular prion protein (PrP) is still poorly understood. We present here an unprecedented role for PrP against Bax-mediated neuronal apoptosis and show that PrP potently inhibits Bax-induced cell death in human primary neurons. Deletion of four octapeptide repeats of PrP (PrP⌬OR) and familial D178N and T183A PrP mutations completely or partially eliminate the neuroprotective effect of PrP. PrP remains anti-apoptotic despite truncation of the glycosylphosphatidylinositol (GPI) anchor signal peptide, indicating that the neuroprotective form of PrP does not require the abundant cell surface GPI-anchored PrP. Our results implicate PrP as a potent and novel anti-apoptotic protein against Bax-mediated cell death.Prion protein (PrP) 1 is a sialoglycoprotein that is highly expressed in brain, heart, lungs, and lymphoid system and at lower levels in several other tissues such as muscle (1, 2). Mature PrP contains two N-linked glycans and a disulfide bond (reviewed in Ref.3). PrP possesses a C-terminal GPI-anchoring signal and a transmembrane domain that can generate type I ( Ctm PrP) or type II ( Ntm PrP) transmembrane-spanning isoforms in isolated endoplasmic reticulum microsomes or phospholiposomes (4 -7). In most cells, the majority of the PrP localizes to the cell surface as a GPI-anchored protein (8). The complete translocation of PrP is dependent on translocation accessory factors (TrAF). In the absence of TrAF, PrP is exclusively synthesized in a transmembrane topology (7).Whereas the role of the infectious form of PrP in a number of human and animal neurodegenerative diseases has been extensively studied, the normal function of PrP is still poorly understood. PrP-null mice display no dramatic phenotype (9). However, evidence indicates that PrP may promote sleep continuity (10). PrP is involved in the regulation of presynaptic copper concentration, intracellular calcium concentration, activation of lymphocytes, astrocyte proliferation, and signal transduction and has antioxidant properties (11-16). Although controversial, PrP-null mice are also found to be impaired in long term potentiation (17)(18)(19). In addition, it has been shown that PrP-null neuronal cell lines are more susceptible to serum deprivation-induced cell death and that Bcl-2 overexpression can attenuate the sensitivity of PrP-null neuronal cell lines to serum deprivation (20). Kurschner and Morgan (21, 22) reported that yeast PrP fusion proteins interact with Bcl-2. Furthermore, four identical N-terminal PrP octapeptide repeats (OR) that are highly conserved in evolution share limited similarity with the Bcl-2 homology domain 2 (BH2) of Bcl-2 proteins (23, 24). Bcl-2 proteins are central to the regulation of cell death, and the BH2 domain is crucial to the anti-apoptotic function of Bcl-2 and its interaction with the pro-apoptotic Bax protein (24,25). Based on these features of PrP, we hypothesized that similar to Bcl-2 family members, PrP may play a role in the regulation of neuronal apoptosis. In the present study, we have...

Genetic Mapping of Activity Determinants within Cellular Prion Proteins

“…Not all mutant PrP alleles scrutinized here lead to loss of protective activity, however, arguing that PrP is not indiscriminately sensitive to perturbation. For example, a Prnp allele impaired in its ability to undergo GPI anchor addition exhibited protective activity comparable with a wt allele, as was also the case for a similar allele tested in the context Bax-initiated apoptosis (26). Although the high frequency at which granule cells are co-transfected when exposed to mixtures of two plasmids indicates that transgene-encoded Dpl and PrP C proteins likely co-exist within the same cells in our paradigm, it is of interest to note that effects in trans (in a cellular sense) have been noted in other paradigms.…”

“…The first is defined by the charged motif KKRPKP at the N terminus of mature PrP C (PrP⌬23-28), and the second corresponds to the octarepeat region (PrP⌬51-90). Of note, a similar octarepeat deletion allele failed to protect against Bax-mediated cell death of human neurons (26) and cell death induced by serum deprivation of a Prnp 0/0 neuronal cell line (57,58). Missense mutations substituting octarepeat histidine residues for glycine (PHGG(G/ S)WGQ 4 3 PGGG(G/S)WGQ 4 ) also inactivated the protective effect of PrP C (Fig.…”

“…Whether cell-surface shed or donated PrP C (59 -61), or other GPI anchor-less forms of PrP C (which may nonetheless retain membrane association) (62) similar to the protein encoded by the S232ter and PrP⌬GPI alleles (this paper and Ref. 26), contribute to neuroprotective effects discerned in Tg mice remains to be established.…”

Molecular biology of prion protein and its first homologous protein

“…Kuwahara, et al previously reported that hippocampal neuronal cells from PrP-/-mice easily undergo apoptosis after withdrawal of serum, which can be prevented by either re-introduction of PrP C or by expressing the anti-apoptotic protein, Bcl-2 (46). Bounhar, et al also showed that PrP C protected human primary neurons from the apoptosis induced by the pro-apoptotic protein Bax (47). It is therefore suggested that PrP C might be an anti-apoptotic protein while PrPLP/Dpl might be pro-apoptotic.…”

Cell Surface Expression of the Prion Protein in Yeast Does Not Alter Copper Utilization Phenotypes

“…Identifying the function of PrP C may provide important clues to pathogenic mechanisms, because there is evidence that PrP C , in addition to serving as a precursor of PrP Sc , acts as a mediator of PrP Sc -induced neurotoxicity (3)(4)(5). A variety of functions have been proposed for PrP C , including roles in metal ion trafficking (6), protection from oxidative stress and apoptosis (7,8), cell adhesion (9), and transmembrane signaling (10). However, the evidence in favor of each of these hypothesized functions is not definitive.…”