Down-Regulation of Shadoo in Prion Infections Traces a Pre-Clinical Event Inversely Related to PrPSc Accumulation

Abstract

During prion infections of the central nervous system (CNS) the cellular prion protein, PrPC, is templated to a conformationally distinct form, PrPSc. Recent studies have demonstrated that the Sprn gene encodes a GPI-linked glycoprotein Shadoo (Sho), which localizes to a similar membrane environment as PrPC and is reduced in the brains of rodents with terminal prion disease. Here, analyses of prion-infected mice revealed that down-regulation of Sho protein was not related to Sprn mRNA abundance at any stage in prion infection. Down-regulation was robust upon propagation of a variety of prion strains in Prnp a and Prnp b mice, with the exception of the mouse-adapted BSE strain 301 V. In addition, Sho encoded by a TgSprn transgene was down-regulated to the same extent as endogenous Sho. Reduced Sho levels were not seen in a tauopathy, in chemically induced spongiform degeneration or in transgenic mice expressing the extracellular ADan amyloid peptide of familial Danish dementia. Insofar as prion-infected Prnp hemizygous mice exhibited accumulation of PrPSc and down-regulation of Sho hundreds of days prior to onset of neurologic symptoms, Sho depletion can be excluded as an important trigger for clinical disease or as a simple consequence of neuronal damage. These studies instead define a disease-specific effect, and we hypothesize that membrane-associated Sho comprises a bystander substrate for processes degrading PrPSc. Thus, while protease-resistant PrP detected by in vitro digestion allows post mortem diagnosis, decreased levels of endogenous Sho may trace an early response to PrPSc accumulation that operates in the CNS in vivo. This cellular response may offer new insights into the homeostatic mechanisms involved in detection and clearance of the misfolded proteins that drive prion disease pathogenesis.

Knockout of the prion protein (PrP)-like Sprn gene does not produce embryonic lethality in combination with PrP C -deficiency

“…Although Prnp and Sprn may encode proteins with overlapping activities (as discussed below), their activities are not interrelated to the extent that their expression levels are counterbalanced. The concept that lack of PrP C produces an increase in Sho expression has been excluded previously by protein analyses of the CNS in Prnp 0/0 mice (21,22,29), and here we can exclude that an alteration in Sprn gene dosage (and hence Sho protein level) affects CNS levels of PrP C (Fig. 4).…”

“…Prior studies have shown that overexpression of wild-type mouse Sho does not alter the outcome of infections with mouse-adapted prion isolates (21)(22)(23)29). On the other hand, a SPRN frame-shift mutation-a putative null allele-was identified in two of 107 vCJD patients but not in 861 controls (30), suggesting a role for Sho in disease susceptibility in humans.…”

“…In prior analyses and experiments presented in Fig. 3C, DEA extraction was used to separate membrane-tethered and extrinsic/secreted forms of APP (27,28) and Sho (22), but low M r Sho species were not scrutinized. Here, examination of DEA extracts from whole brain and from dissected olfactory bulb, pituitary, and cerebellum homogenates revealed a 6-kDa fragment of Sho in the pellet fraction of TgSprn mice (and in wild-type mice, but not in Sprn 0/0 mice, after longer autoradiographic exposures).…”

“…Interestingly, another biological context where expression of Sho protein is reduced in the instance of prion infections (21)(22)(23)(24). Although Sho down-regulation could be dissociated temporally from neurological signs used to diagnose end-stage disease (22,23), we note that prion disease can also be associated with changes outside of the CNS: specifically, a number of prion strains produce a loss in body mass, and less commonly, some strains produce an increase (32,33). Further studies will be needed to ascertain whether Sho levels are altered in hypothalamic neurons of mice infected with different prion strains and, if alterations are evident, whether or not there is a consistent relationship with perturbed body mass.…”

“…Sho, like PrP, is attached to the cell surface by a GPI anchor (21). In prion infections, levels of Sho protein are markedly reduced (21)(22)(23)(24). In terms of physiological action, Sho, like PrP C , can exhibit neuroprotective properties (21) and shares a number of binding partners in common with PrP C (25).…”