title: Inhibition of autophagic-lysosomal delivery and autophagic lactolysis by asparagine.
abstract: Overall autophagy was measured in isolated hepatocytes as the sequestration and lysosomal hydrolysis of electroinjected [14C]lactose, using HPLC to separate the degradation product [14C]glucose from undegraded lactose. In addition, the sequestration step was measured separately as the transfer from cytosol to sedimentable cell structures of electroinjected [3H]raffinose or endogenous lactate dehydrogenase (LDH; in the presence of leupeptin to inhibit lysosomal proteolysis). Inhibitor effects at postsequestrational steps could be detected as the accumulation of autophaged lactose (which otherwise is degraded intralysosomally), or of LDH in the absence of leupeptin. Asparagine, previously shown to inhibit autophagic but not endocytic protein breakdown, strongly suppressed the autophagic hydrolysis of electroinjected lactose. Vinblastine, which inhibits both types of degradation, likewise suppressed lactose hydrolysis. Asparagine had little or no effect on sequestration, but caused an accumulation of autophaged LDH and lactose, indicating inhibition at a postsequestrational step. Neither asparagine nor vinblastine affected the degradation of intralysosomal lactose preaccumulated in the presence of the reversible lysosome inhibitor propylamine. However, if lactose was preaccumulated in the presence of asparagine, both asparagine and vinblastine suppressed its subsequent degradation. The data thus indicate that autophagic-lysosomal delivery, i.e., the transfer of autophaged material from prelysosomal vacuoles to lysosomes, is inhibited selectively by asparagine and non-selectively by vinblastine.
authors.affiliation: Department of Tissue Culture, Norwegian Radium Hospital, Oslo, Norway.
authors.forename: H
authors.initials: H
authors.lastname: Høyvik
authors.forename: P B
authors.initials: PB
authors.lastname: Gordon
authors.forename: T O
authors.initials: TO
authors.lastname: Berg
authors.forename: P E
authors.initials: PE
authors.lastname: Strømhaug
authors.forename: P O
authors.initials: PO
authors.lastname: Seglen
mesh_terms: D000818:Animals; D001216:Asparagine / Q000494:pharmacology*; D002851:Chromatography, High Pressure Liquid; D003593:Cytoplasm / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D007785:Lactose / Q000378:metabolism*; D008247:Lysosomes / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D051381:Rats; D011919:Rats, Inbred Strains; D014747:Vinblastine / Q000494:pharmacology; D001616:beta-Galactosidase / Q000378:metabolism
chemical_list: D014747:Vinblastine; D001216:Asparagine; D007770:L-Lactate Dehydrogenase; D001616:beta-Galactosidase; D007785:Lactose