Basis of lethality in C. elegans lacking CUP-5, the Mucolipidosis Type IV orthologue

Abstract

Mutations in MCOLN1, which encodes the protein h-mucolipin-1, result in the lysosomal storage disease Mucolipidosis Type IV. Studies on CUP-5, the human orthologue of h-mucolipin-1 in Caenorhabditis elegans, have shown that these proteins are required for lysosome biogenesis. We show here that the lethality in cup-5 mutant worms is due to two defects, starvation of embryonic cells and general developmental defects. Starvation leads to apoptosis through a CED-3-mediated pathway. We also show that providing worms with a lipid-soluble metabolite partially rescues the embryonic lethality but has no effect on the developmental defects, the major cause of the lethality. These results indicate that supplementing the metabolic deficiency of Mucolipidosis Type IV patients mat not be sufficient to alleviate the symptoms due to tissue degeneration.

Suppression of thecup-5mucolipidosis type IV-related lysosomal dysfunction by the inactivation of an ABC transporter inC. elegans

“…RNAi was done by the feeding method as previously described (Timmons and Fire, 1998). Markers used were: pgp-2(gk114) I (Nunes et al, 2005); cup-5(ar465) III (Fares and Greenwald, 2001b); cup-5(zu223) III (Hersh et al, 2002);unc-36(e251) III (closely linked to cup-5) (Brenner, 1974;Fares and Greenwald, 2001a;Fares and Greenwald, 2001b);unc-69(e587) III (Brenner, 1974); ced-9(n1950n2161) III (Gumienny et al, 1999); rme-2((b1008) IV (Grant and Hirsh, 1999); cpl-1(ok360) V (Britton and Murray, 2004); rme-1(b1045) V (Grant et al, 2001); cup-10(ar479) V (Dang et al, 2003); rme-6(b1018) X (Sato et al, 2005); bIs1 pRF4] (which expresses a fusion of the worm YP170 protein VIT-2 to GFP) (Grant and Hirsh, 1999);cdIs77[GFP::LGG-1;unc-119(+)-pmyo-2::GFP] X (which expresses GFP-LGG-1 in all cells and GFP in the pharynx) (Schaheen et al, 2006);nT1[qIs51] IV, V (is a dominant balancer) (Siegfried et al, 2004); and qC1 (a balancer chromosome that suppresses recombination between cup-5 and unc-36) (Graham and Kimble, 1993). cup-5(zu223) unc-36(e251) worms bearing various transgenes were isolated from qC1-balanced parent heterozygotes; the eggs from these homozygous progeny were analyzed in the various assays.…”

“…We used quantitative fluorescence measurements instead of western blots for this analysis because the defect in cup-5(zu223) embryos is confined to specific stages of embryonic development (Schaheen et al, 2006). We also focused on developing intestinal cells at these embryonic stages because we can unambiguously identify intestinal (and pharyngeal) cells in cup-5(zu223) embryos based on position and morphology in the embryos.…”

“…The large vacuoles are hybrids of late endosomes and lysosomes and represent the terminal endocytic compartments, at least in scavenger cells called coelomocytes (Treusch et al, 2004). In cup-5 null embryos, the degradation of yolk and of membrane proteins is significantly delayed, particularly in developing intestinal cells (Schaheen et al, 2006). As in other tissues, this defect is accompanied by an expansion in the size of the terminal compartment.…”

“…As in other tissues, this defect is accompanied by an expansion in the size of the terminal compartment. Furthermore, this leads to ectopic apoptosis, to developmental abnormalities and ultimately to organism death primarily because of the developmental defects (Hersh et al, 2002;Schaheen et al, 2006).…”

Characterization and expression analysis of mcoln1.1 and mcoln1.2, the putative zebrafish co-orthologs of the gene responsible for human mucolipidosis type IV

“…Studies on CUP-5, the Caenorhabditis elegans ortholog of MCOLN1, have shown that this protein is involved in endocytic processes and lysosome biogenesis and function (Fares and Greenwald, 2001;Schaheen et al, 2006). Interestingly, the endocytosis defect observed in cup-5 mutants could be rescued by the transgenic expression of human MCOLN1 or MCOLN3 (Treusch et al, 2004).…”