JPDI, a Novel Endoplasmic Reticulum-resident Protein Containing Both a BiP-interacting J-domain and Thioredoxin-like Motifs

Abstract

Several endoplasmic reticulum (ER)-resident luminalproteins have a characteristic ER retrieval signal, KDEL, or its variants at their C terminus. Our previous work searching EST databases for proteins containing the C-terminal KDEL motif predicted some novel murine proteins, one of which designated JPDI (J-domaincontaining protein disulfide isomerase-like protein) is characterized in this study. The primary structure of JPDI is unique, because in addition to a J-domain motif adjacent to the N-terminal translocation signal sequence, four thioredoxin-like motifs were found in a single polypeptide. As examined by Northern blotting, the expression of JPDI was essentially ubiquitous in tissues and almost independent of ER stress. A computational prediction that JPDI is an ER-resident luminal protein was experimentally supported by immunofluorescent staining of epitope-tagged JPDI-expressing cells together with glycosylation and protease protection studies of this protein. JPDI probably acts as a DnaJlike partner of BiP, because a recombinant protein carrying the J-domain of JPDI associated with BiP in an ATP-dependent manner and enhanced its ATPase activity. We speculate that for the folding of some proteins in the ER, chaperoning by BiP and formation of proper disulfide bonds may synchronously occur in a JPDI-dependent manner.

ERdj4 and ERdj5 Are Required for Endoplasmic Reticulum-associated Protein Degradation of Misfolded Surfactant Protein C

“…Surprisingly, mutation of the HPD motif significantly enhanced association of ERdj5 with misfolded SP-C (Supplemental Figure S2A). Further analyses revealed that, in contrast to results of a previous study (Hosoda et al, 2003), the HPD mutation did not alter association of ERdj5 (or ERdj4) with BiP (Supplemental Figure S2C). Therefore, stable complexes of misfolded SP-C, BiP, and ERdj5 formed in the absence of the catalytic activity of ERdj5.…”

“…ERdj5 (DNAJC10/JPDI) is an ER-resident lumenal protein with a canonical C-terminal KDEL retrieval motif, a J domain, and four thioredoxin-like domains (Cunnea et al, 2003;Hosoda et al, 2003). Deletion of the J domain prevented interaction of ERdj5 with both BiP and misfolded SP-C, suggesting that association of ERdj5 with substrate was mediated by BiP.…”

“…vitro (Cunnea et al, 2003;Hosoda et al, 2003) may reflect the substrate specificity of ERdj5 or the requirement of a partner protein(s), such as BiP, for unfoldase activity. Alternatively, ERdj5 may act as a redox sensor to identify proteins with unpaired/mispaired cysteines and target them for elimination via ERAD.…”

“…ERdj4 and ERdj5 were inserted into pIRES2-EGFP vector (Clontech), BiP was inserted into pcDNA3.1 (Invitrogen, Carlsbad, CA), and ERdj3 was inserted into pCMV6-XL5 (OriGene Technologies, Rockville, MD). The hemagglutinin (HA) tag was added at a position before the C-terminal KDEL in ERdj5 as described previously (Hosoda et al, 2003). Mouse insulin 2-Flag was cloned from mouse pancreatic ␤-cell cDNA using PCR with reverse transcription and inserted into the pIRES2-EGFP vector.…”

The Role of p58IPK in Protecting the Stressed Endoplasmic Reticulum

“…These results point to a new functional assignment for p58 IPK in serving as a BiP cochaperone to optimize protein folding homeostasis in the ER. Because p58 IPK represents one among several (at least five others) ER localized Jdomain proteins that interface with BiP (Feldheim et al, 1992;Brightman et al, 1995;Shen et al, 2002;Hosoda et al, 2003;Shen and Hendershot, 2005), its absence would result in an ER lumen that is only modestly compromised in overall folding capacity. The functional consequences of this slightly lower protein maturation capacity would be obscured under all but the most taxing conditions and can explain each of the following phenotypic observations in cells and animals lacking p58 IPK .…”