The DnaJ-domain Protein RME-8 Functions in Endosomal Trafficking

Abstract

Through a proteomic analysis of clathrin-coated vesicles from rat liver we identified the mammalian homolog of receptor-mediated endocytosis 8 (RME-8), a DnaJ domain-containing protein originally identified in a screen for endocytic defects in Caenorhabditis elegans. Mammalian RME-8 has a broad tissue distribution, and affinity selection assays reveal the ubiquitous chaperone Hsc70, which regulates protein conformation at diverse membrane sites as the major binding partner for its DnaJ domain. RME-8 is tightly associated with microsomal membranes and co-localizes with markers of the endosomal system. Small interfering RNA-mediated knock down of RME-8 has no influence on transferrin endocytosis but causes a reduction in epidermal growth factor internalization. Interestingly, and consistent with a localization to endosomes, knock down of RME-8 also leads to alterations in the trafficking of the cation-independent mannose 6-phosphate receptor and improper sorting of the lysosomal hydrolase cathepsin D. Our data demonstrate that RME-8 functions in intracellular trafficking and provides the first evidence of a functional role for a DnaJ domainbearing co-chaperone on endosomes.

DNAJC13 mutations in Parkinson disease

“…Preliminary functional analysis indicates that the expression of DNAJC13 p.Asn855Ser in COS7 cells leads to accumulation of TF in endosomal compartments, suggesting interference with endosomal compartmentalization or trafficking as the pathological mechanism of disease. The importance of DNAJC13 function in endosomal trafficking is evident from several studies showing alterations in trafficking of well-known retromer cargos when its function is perturbed (13,20). The function of retromer itself is linked to PD through the observation that mutations in another retromer subunit, VPS35, can cause familial parkinsonism (6,7).…”

“…To investigate the physiological consequence of the DNAJC13 p.Asn855Ser mutation, we assessed the uptake of transferrin (TF), a membrane receptor ligand that is constitutively internalized and recycled through the endosome pathway (12,13). COS7 cells expressing a GFP control, wild-type (WT) or mutant DNAJC13 were exposed to 568-conjugated TF for 1 h at 48C and either fixed immediately (T0) or after a 40 min incubation at 378C (T40).…”

“…Despite this effort, none of the variants identified could account for disease in these patents; coding variants with a minor allele frequency under 5% are presented in Supplementary Material, Table S4. DNAJC13, also known as RME-8, is a DNAJ-domain-bearing protein that localizes to the membranes of the endosomal system where it binds through its DNAJ-domain to the molecular chaperone heat shock cognate 70 (Hsc70) (HSPA8) (13). HSPA8 is a clathrin uncoating ATPase that has been most extensively studied for its role in the dissociation of clathrin from clathrincoated vesicles (CCVs) (17).…”

How Ricin and Shiga Toxin Reach the Cytosol of Target Cells: Retrotranslocation from the Endoplasmic Reticulum

“…Furthermore, the clathrin associated Hcs70 cochaperone RME-8 regulates endosomal trafficking of STx (Popoff et al 2009) whilst having no effect on the transport of cholera toxin (Girard et al 2005). …”

Receptor-mediated Endocytosis 8 Utilizes an N-terminal Phosphoinositide-binding Motif to Regulate Endosomal Clathrin Dynamics

“…Likewise, studies of rme-8 mutants in Drosophila displayed blockage in the internalization of the Bride of Sevenless receptor causing the formation of the rough eye phenotype (5). RME-8 has not only been shown to be highly conserved in the animal kingdom (6,7), but it is also present in plants, where studies in Arabidopsis thaliana have demonstrated that RME-8 mutants exhibit gravitropism defects and associate with endosomal structures (8). Finally, a recent study has determined that a gain of function mutation in RME-8 correlates with Parkinson disease (9), highlighting the importance of studying RME-8 biology in more detail.…”