Abstract
ABSTRACTThe Sec61-complex as a dynamic polypeptide-conducting channel mediates protein transport into the human endoplasmic reticulum (ER) with the help of additional components. ER membrane resident Hsp40-type co-chaperone Sec63 as well as the ER lumenal Hsp70-type chaperone BiP were proposed to facilitate channel opening in a precursor-specific fashion. Here, we report on their rules of engagement in ER import of the prion protein (PrP) by addressing sixteen PrP-related variants which differ in their signal peptides and mature parts, respectively. Transport into the ER of semi-permeabilized human cells was analyzed upon depletion of the components by siRNA- or toxin-treatment. The results are consistent with the view of separate functions of BiP and Sec63 and strongly suggest that the co-chaperone/chaperone-pair facilitates Sec61 channel gating to the open state when precursor polypeptides with weak signal peptides in combination with detrimental features in the adjacent mature part were targeted. Thus, we expand the view of chaperone-mediated Sec61 channel gating by providing a novel example of a polybasic motif that interferes with signal peptide-mediated Sec61 channel gating. This article has an associated First Person interview with the first author of the paper.
Highlights
The endoplasmic reticulum (ER) represents the major site of membrane and secretory protein biogenesis of the mammalian cell
Depletion of BiP inhibits ER import of prion protein due to the signal peptide The PrP-derived signal peptide is believed to have a weak Sec61 channel gating capacity compared to the signal peptides (SP) of ppl (Rutkowski et al, 2001)
Attributing its weak capacity for ER import to the causes, we investigated the requirements for targeting and translocation of the PrP precursor to the human ER
Summary
The endoplasmic reticulum (ER) represents the major site of membrane and secretory protein biogenesis of the mammalian cell. Failed ER import and subsequent cytosolic aggregation or integration into the ER membrane with an unusual topology is, in the case of the prion protein (PrP), associated with neurotoxicity The normally GPI-anchored plasma membrane protein shows an unusual complex topology with a soluble and two membrane integrated forms driven by various signals in the PrP sequence
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