Graphical summary of research aim and key findings.

Abstract

(A) To study the role of the GPI-anchor signal sequence (GPI-SS) in determining localization and biology of a protein, the GPI-SS of PrPC (dark green) was exchanged for the one of Thy-1 (red). Both proteins, WTPrPC and the PrPCGPIThy-1 mutant, are expressed at the cell surface and comprise a GPI-anchor. A sialic acid modification (green asterisk) typical for the GPI-anchor of PrPC is lacking in the GPI-anchor of PrPCGPIThy-1 (note its red GPI-anchor referring to the GPI-anchor of Thy-1 [in B]). This suggests that an altered GPI-SS in the mRNA results in a different GPI-anchor of the mature protein. (B) PrPC (green) and Thy-1 (blue) are both known residents of lipid rafts/DRMs. While PrPC is located in their periphery and able to leave and re-enter these membrane subdomains, Thy-1 has been shown to occupy more central regions therein. Despite several biochemical methods applied in our study, we were unable to demonstrate a relative re-distribution of PrPCGPIThy-1 towards Thy-1 compared to WTPrPC. (C) Nevertheless, the overall cellular sorting is altered with PrPCGPIThy-1 being relatively more transported towards the apical compartment compared to WTPrPC with its predominant basolateral sorting in a polarized epithelial cell model such as MDCK cells. This also holds true in our transgenic mice and translates to an increased axonal sorting of PrPCGPIThy-1 in primary neurons compared to a mainly somatodendritic presence of PrPC in wild-type neurons. (D) Altered GPI-anchor composition and sorting of PrPCGPIThy-1 results in different biological consequences (in comparison to WTPrPC): (i) Endogenous proteolytic shedding by the metalloprotease ADAM10 (orange) at the cell surface is reduced. (ii) Although PrPCGPIThy-1, in principle, is able to transduce PrPSc-associated toxic signaling (e.g. via cleaved caspase-3), signaling via the MAP kinase ERK1/2 is reduced upon prion infection. Though not investigated here, p38 signaling may be reduced at early time points, contributing to delay to terminal disease (showed as p38?). Key hallmarks of prion-associated neuropathology are also altered in the transgenic mice including (iii) decreased PrPSc production and deposition, (iv) reduced vacuolization (spongiosis) of the brain parenchyma, and (v) reduced induction of astrocytes and microglia (reactive gliosis). These changes are accompanied with prolonged survival of the PrPCGPIThy-1 mice and support a relevant impact of the GPI-SS on a GPI-AP`s biology.</p