Producción de anticuerpos monoclonales específicos para la proteína LC3

LUJAN HUGO; ROPOLO ANDREA; COLOMBO MARISA; MILISTELLO RODRIGO

2006-02-02

2006-05-06

Biológica

Otros campos

The microtubule=associated protein 1 light chain 3 (MAP1-LC3) was originally discovered as a protein associated to microtubules, that co-purifies with both MAP1A and MAP1B. More recently, it was demonstrated that LC3 is a mammalian homologue of the product of the yeast gene Atg8, a protein required for the formation of autophagosomes. LC3 is recruited to the sequestering membrane after a pos-traslational processing that involves conjugation toa a lipid molecule. Thus, LC3 exists in two forms, the cytosolic form LC3-I, and the processed lipidated form (i.e. LC3-II) located on the autophagosomal membrane. When autophagy is induced LC3 expression and processing are upregulated. This stravation-dependent increment in LC3-II level is blocked by treatment with the autophagy inhibitors 3 mehtiladenine or wortmannin. In contrast, treatment with vinblastine resulted in an additional large increase in LC3-II and causes the accumulation of larger autophagosomes. Rescently it has been suggested that MAP=LC3 can act as an adaptor protein between microtubules and autophagosomes. To date, LC3 is one of the best available markers for autophagosomes. SOURCE The mouse monoclonal antibody 5D3 was raised using as immunogen the recombinant protein GST-LC3. APPLICATIONS The mAb 5D3 is recommended for detection of endogenous LC3 in HeLa cells by immunofluorescence. By Western blot we have been able to detect the recombinant protein and in K562 cells the mAb 5D3 recognizes a band of 16 kDa in whole cell extracts.