RGD Reference Report - In vitro reconstitution of fusion between immature autophagosomes and endosomes. - Rat Genome Database

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In vitro reconstitution of fusion between immature autophagosomes and endosomes.

Authors: Morvan, J  Kochl, R  Watson, R  Collinson, LM  Jefferies, HB  Tooze, SA 
Citation: Morvan J, etal., Autophagy. 2009 Jul;5(5):676-89. Epub 2009 Jul 6.
RGD ID: 4892338
Pubmed: PMID:19337031   (View Abstract at PubMed)

Autophagy is a highly conserved degradative pathway whereby a double membrane engulfs cytoplasmic constituents to form an autophagic vacuole or autophagosome. An essential requirement for efficient autophagy is the acquisition of an adequate degradative capacity by the autophagosomes. To acquire this capacity the immature autophagic vacuoles (AVis) obtain lysosomal hydrolases by fusion with endosomes. The current models suggest that at least two types of endosomes, early and late, fuse with AVis to form mature, degradative AVds. This fusion and maturation requires proteins also involved in endosome maturation such as Rab7. However, it is not known if there are molecular requirements unique to AVi-endosome fusion. To identify and investigate the molecular requirements of this fusion we developed a cell-free fusion assay based on content mixing, which occurs after fusion of isolated AVis and different endosomal fractions. Our assay shows that isolated AVis can fuse to a similar extent in vitro with both early and late endosomes. Furthermore, fusion between autophagosomes and endosomes requires cytosolic and endosomal proteins, but does not show a nucleotide-dependence, and is partially N-ethylmaleimide sensitive. We also demonstrate that the lipidated form of the autophagosomal protein LC3 is dispensable for this fusion event.



Gene Ontology Annotations    Click to see Annotation Detail View

Cellular Component

  
Object SymbolSpeciesTermQualifierEvidenceWithNotesSourceOriginal Reference(s)
Map1lc3aRatautophagosome  IDA  RGD 
Map1lc3aRatautophagosome membrane  IDA  RGD 
Igf2rRatlate endosome  IDA  RGD 
Map1lc3aRatlate endosome  IDA  RGD 
Rab7aRatlate endosome  IDA  RGD 

Objects Annotated

Genes (Rattus norvegicus)
Igf2r  (insulin-like growth factor 2 receptor)
Map1lc3a  (microtubule-associated protein 1 light chain 3 alpha)
Rab7a  (RAB7A, member RAS oncogene family)

Objects referenced in this article
Gene Lamp2 lysosomal-associated membrane protein 2 Rattus norvegicus

Additional Information