RME-8, a Conserved J-Domain Protein, Is Required for Endocytosis inCaenorhabditis elegans

Abstract

By genetic analysis of Caenorhabditis elegans mutants defective in yolk uptake, we have identified new molecules functioning in the endocytosis pathway. Here we describe a novel J-domaincontaining protein, RME-8, identified by such genetic analysis. RME-8 is required for receptormediated endocytosis and fluid-phase endocytosis in various cell types and is essential for C. elegans development and viability. In the macrophage-like coelomocytes, RME-8 localizes to the limiting membrane of large endosomes. Endocytosis markers taken up by the coelomocytes rapidly accumulate in these large RME-8 -positive endosomes, concentrate in internal subendosomal structures, and later appear in RME-8 -negative lysosomes. rme-8 mutant coelomocytes fail to accumulate visible quantities of endocytosis markers. These observations show that RME-8 functions in endosomal trafficking before the lysosome. RME-8 homologues are found in multicellular organisms from plants to humans but not in the yeast Saccharomyces cerevisiae. These sequence homologies suggest that RME-8 fulfills a conserved function in multicellular organisms. INTRODUCTIONIntracellular trafficking of endocytosed molecules requires an elaborate network of organelles (Mellman, 1996;Mukherjee et al., 1997). The compartments within the endocytic network were established in mammalian cells by studying the trafficking kinetics of ligands and receptors (such as transferrin and transferrin receptor, epidermal growth factor [EGF], and EGF-receptor, and low-density lipoprotein [LDL] and LDL receptor) and of fluid-phase markers. Ligands and receptors are often internalized through clathrin-coated pits and vesicles and then are targeted to early or sorting endosomes by vesicle fusion. In the sorting endosome, many ligands can dissociate from their receptors because of a low pH environment. The ligands bound for degradation are then sorted and targeted to the late endosome (Dunn and Maxfield, 1992). Recycling receptors are sorted to an endocytic recycling compartment, from which they recycle back to the plasma membrane (van der Sluijs et al., 1992). In the late endosome, ligands and other lysosome targeted molecules are further sorted, concentrated, and delivered to the lysosome for enzymatic degradation (Futter et al., 1996).Each of these trafficking steps occurs in a specific vesicular compartment. Despite the continuous flow of endocytosed molecules, these compartments are characterized by unique morphologies, sizes, intravesicular pH levels, membrane proteins, and lipid compositions, although these characteristics can vary with different cell types (Mellman, 1996;Mukherjee et al., 1997;Odorizzi et al., 2000). In mammalian cells, sorting endosomes have a tubular-vesicular morphology with an internal pH ϳ6.0 (Mukherjee et al., 1997). Late endosomes are similar in size to sorting endosomes but have characteristic internal vesicles formed by the invagination of late endosomal membranes (Hirsch et al., 1968;Futter et al., 1996). Two alternative models were proposed to describe the t...

Role of the Caenorhabditis elegans Multidrug Resistance Gene, mrp-4, in Gut Granule Differentiation

“…We compared the distribution of MRP-4 relative to GFP fusion proteins that in wild-type are localized to early endosomes (RAB-5), recycling endosomes (RME- 1), and late endosomes/lysosomes (LMP-1, RAB-7, RME-8) Zhang et al 2001;Chen et al 2006). MRP-4 showed little colocalization with any of the markers in wild type consistent with its localization to gut granules (supplemental Table II at http:// www.genetics.org/supplemental/).…”

Membrane Trafficking Illuminates a Path to Parkinson’s Disease

“…DNAJC13 (Dnaj (Hsp40) homolog, subfamily C, member 13) is a responsible gene that is responsible for the autosomal-dominant PARK21-linked familial form of PD (Vilarino-Guell et al 2014). DNAJC13 was originally identified as the mammalian homolog of receptor mediated endocytosis 8 (RME-8) in a screen for the endocytic defect phenotype in Caenorhabditis elegans (Zhang et al 2001). As the name implies, several lines of evidence indicate that RME-8 may serve as the regulator of endocytosis.…”

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

“…Endosomes-It has been demonstrated that RME-8 displays a punctate localization pattern due to its association with early endosomal structures (4,6,7,18,24,37) that can be disrupted by the PI(3)-kinase inhibitor wortmannin and expression of the PI(3)P phosphatase MTMR2 (24). Therefore, to complement the above biochemical assays, we screened the various RME-8 mutants for subcellular localization changes from punctate to a diffused cytoplasmic pattern.…”

“…Further structure-function analysis will be required to experimentally determine whether the N-terminal region of RME-8 truly adopts a PH-like fold and to examine whether RME-8's association with PI(3,5)P 2 and PI(3,4,5)P 3 is biologically relevant. However, it is interesting to note that in addition to early endosomes, RME-8 has been reported to localize to late endosomes and recycling endosomes, which are known to contain PI(3,5)P 2 and PI(3,4,5)P 3 , respectively (4,7,45). Thus, it will be interesting to assess whether these PIP isoforms localized on distinct endosome subtypes regulate subcellular localization of RME-8.…”

“…Receptor-mediated endocytosis 8 (RME-8) 3 was first discovered utilizing genetic screens in Caenorhabditis elegans, where rme-8 mutants exhibited defects in receptor-mediated yolk endocytosis in the oocyte and fluid phase endocytosis in the coelomocyte (4). 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).…”