Ubiquitin-activating enzyme, E1, is associated with maturation of autophagic vacuoles.

Abstract

Abstract. The ubiquitin-activating enzyme, El, is required for initiating a multi-step pathway for the covalent linkage of ubiquitin to target proteins. A CHO cell line containing a mutant thermolabile El, ts20, has been shown to be defective in stress-induced degradation of proteins at restrictive temperature (Gropper et al., 1991. J. Biol. Chem. 266:3602-3610). Parental E36 cells responded to restrictive temperature by stimulating lysosome-mediated protein degradation twofold. Such a response was not observed in ts20 cells. The absence of accelerated degradation in these cells at 39.5~ was accompanied by an accumulation of autolysosomes. The fractional volume of these degradative autophagic vacuoles was at least sixfold greater than that observed for either E36 cells at 30.5 ~ or 39.5~ or ts20 cells at 30.5~ These vacuoles were acidic and contained both acid phosphatase and cathepsin L, but, unlike the autolysosomes observed in E36 cells, ubiquitin-conjugated proteins were conspicuously absent. Combined, our results suggest that in ts20 cells, which are unable to generate ubiquitin-protein conjugates due to heat inactivation of El, the formation and maturation of autophagosomes into autolysosomes is normal, but the conversion of autolysosomes into residual bodies is disrupted.

Suppressors of clathrin deficiency: overexpression of ubiquitin rescues lethal strains of clathrin-deficient Saccharomyces cerevisiae.

“…Recently, the ubiquitin-activating enzyme (El) was localized to the cytoplasmic face of autolysosomes (63), and mammalian cells harboring a temperature-sensitive mutation in El were found to be defective in the autophagic response upon exposure to high temperatures (12,25,61). Late-stage degradative vacuoles containing acid hydrolases accumulated, but conversion to residual bodies was blocked and luminal ubiquitin conjugates, which are normally found in autolysosomes, were absent (39). In S. cerevisiae, ubiquitin conjugates have also been localized to the lysosome-like vacuole and accumulate in strains which lack the key vacuolar peptidases, proteinases A and B (69).…”

The formation of peripheral myelin protein 22 aggregates is hindered by the enhancement of autophagy and expression of cytoplasmic chaperones

“…The specimens were then treated with 2% OsO 4 and uranyl acetate, and dehydrated and embedded in Epon resin. For localization of acid phosphatase (AcP) activity, the samples were treated with cytidine 5′-monophosphate and cerium chloride after fixation (Lenk et al, 1992). Thin sections were examined on a JEOL 100CX transmission electron microscope.…”

“…Therefore, the presence of the lysosomal enzyme, AcP, in aggresome-containing cells was examined by monitoring enzymatic activity (Lenk et al, 1992) (Fig. 4A).…”

Autophagy: Eating for Good Health

“…Whether ubiquitination itself is sufficient to target soluble proteins for autophagic degradation remains an open question, although a number of interactions between the two pathways have been documented. Free ubiquitin and ubiquitin conjugants are found within autophagosomes (47,48), predominantly in autophagolysosomes (49). The E1 ubiquitin-conjugating enzyme equips cells to accelerate autophagosomal degradation of proteins in response to heat-stress (50,51), apparently by promoting proteolysis by autophagolysosomes, rather than their biogenesis (49).…”

“…Free ubiquitin and ubiquitin conjugants are found within autophagosomes (47,48), predominantly in autophagolysosomes (49). The E1 ubiquitin-conjugating enzyme equips cells to accelerate autophagosomal degradation of proteins in response to heat-stress (50,51), apparently by promoting proteolysis by autophagolysosomes, rather than their biogenesis (49). Although their proteasomes function normally, mice that lack the autophagy enzyme Atg7 amass aggregates of ubiquitinated proteins within their liver cells (52).…”