Cell Signaling Technology

Product Pathways - Apoptosis

Atg4A (D62C10) Rabbit mAb #7613

Applications Reactivity Sensitivity MW (kDa) Isotype
W IP H R Endogenous 48-60 Rabbit IgG

Applications Key:  W=Western Blotting  IP=Immunoprecipitation
Reactivity Key:  H=Human  R=Rat
Species cross-reactivity is determined by western blot. Species enclosed in parentheses are predicted to react based on 100% sequence homology.

Protocols

Specificity / Sensitivity

Atg4A (D62C10) Rabbit mAb recognizes endogenous levels of total Atg4A protein. This antibody recognizes unidentified bands within the molecular weight range of 48-60 kDa which decrease with silencing of Atg4A expression.

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues near the carboxy terminus of human Atg4A protein.

Western Blotting

Western Blotting

Western blot analysis of extracts from HeLa cells, transfected with 100 nM SignalSilence® Control siRNA (Unconjugated) #6568 (-) or SignalSilence® Atg4A siRNA I #6427 (+), using Atg4A (D62C10) Rabbit mAb (upper) or α-Tubulin (11H10) Rabbit mAb #2125 (lower). The Atg4A (D62C10) Rabbit mAb confirms silencing of Atg4A expression, while the α-Tubulin (11H10) Rabbit mAb is used as a loading control.

Western Blotting

Western Blotting

Western blot analysis of extracts from various cell lines using Atg4A (D62C10) Rabbit mAb.

Western Blotting

Western Blotting

Western blot analysis of extracts from 293T cells, mock transfected (-) or transfected with a DYKDDDDK-tagged human Atg4A construct (+), using Atg4A (D62C10) Rabbit mAb (upper) and DYKDDDDK Tag Antibody #2368 (lower).


Background

Autophagy is a catabolic process for the autophagosomic-lysosomal degradation of bulk cytoplasmic contents. Control of autophagy was largely discovered in yeast and involves proteins encoded by a set of autophagy-related genes (Atg) (1). Formation of autophagic vesicles requires a pair of essential ubiquitin-like conjugation systems, Atg12-Atg5 and Atg8-phosphatidylethanolamine (Atg8-PE), which are widely conserved in eukaryotes (2). Numerous mammalian counterparts to yeast Atg proteins have been described, including three Atg8 proteins (GATE-16, GABARAP, and LC3) and four Atg4 homologues (Atg4A/autophagin-2, Atg4B/autophagin-1, Atg4C/autophagin-3, and Atg4D/autophagin-4) (3-5). The cysteine protease Atg4 is pivotal to autophagosome membrane generation and regulation. Atg4 primes the Atg8 homologue for lipidation by cleaving its carboxy terminus and exposing its glycine residue for E1-like enzyme Atg7. The Atg8 homologue is transferred to the E2-like enzyme Atg3 before forming the Atg8-PE conjugate. During later stages of autophagy, Atg4 can reverse this lipidation event by cleaving PE, thereby recycling the Atg8 homologue (6).

Atg4A predominately cleaves GATE-16, athough it can cleave the other mammalian Atg8 homologues with lesser efficiencies (4,7,8). Mutation in the corresponding Atg4A gene is critical for redox regulation and inhibits autophagosome formation. Expression of this Atg4A mutation demonstrates a role for reactive oxygen species in nutrient-deprived autophagy (9).

  1. Reggiori, F. and Klionsky, D.J. (2002) Eukaryot Cell 1, 11-21.
  2. Ohsumi, Y. (2001) Nat Rev Mol Cell Biol 2, 211-6.
  3. Kabeya, Y. et al. (2000) EMBO J 19, 5720-8.
  4. Kabeya, Y. et al. (2004) J Cell Sci 117, 2805-12.
  5. Mariño, G. et al. (2003) J Biol Chem 278, 3671-8.
  6. Sou, Y.S. et al. (2008) Mol Biol Cell 19, 4762-75.
  7. Scherz-Shouval, R. et al. (2003) J Biol Chem 278, 14053-8.
  8. Li, M. et al. (2011) J Biol Chem 286, 7327-38.
  9. Scherz-Shouval, R. et al. (2007) EMBO J 26, 1749-60.

Application References

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For Research Use Only. Not For Use In Diagnostic Procedures.

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