Apoptosis and Autophagy

Atg4C Antibody #5262

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Applications Key:   W = Western Blotting  IP = Immunoprecipitation
Reactivity Key: H = Human  M = Mouse  Mk = Monkey  
Species cross-reactivity is determined by Western blot.

Protocols

5262

Western Blotting, Immunoprecipitation,

Specificity / Sensitivity

Atg4C Antibody detects endogenous levels of total Atg4C protein. A band of unknown origin is detected at 23kDa. The intensity of this band is reduced by treatment with Atg4C siRNA.

Source / Purification

Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Ser430 of human Atg4C. Antibodies are purified by protein A and peptide affinity chromatography.

Western Blotting

Western blot analysis of extracts from various cell lines using Atg4C Antibody.

Western Blotting

Western blot analysis of extracts from HeLa cells, transfected with 100 nM SignalSilence® Control siRNA (Unconjugated) #6568 (-) or SignalSilence® Atg4C siRNA I (+), using Atg4C Antibody #5262 (upper) or β-Tubulin (9F3) Rabbit mAb #2128 (lower). The Atg4C Antibody confirms silencing of Atg4C expression, while the β-Tubulin (9F3) Rabbit mAb is used as a loading control.

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 and recycle the Atg8 homologue (6).

1. Reggiori, F. and Klionsky, D.J. et al. (2002) Eukaryot Cell1, 11 - 21.
2. Ohsumi, Y. et al. (2001) Nat Rev Mol Cell Biol2, 211 - 6.
3. Kabeya, Y. et al. (2000) EMBO J19, 5720 - 8.
4. Kabeya, Y. et al. (2004) J Cell Sci117, 2805 - 12.
5. Mariño, G. et al. (2003) J Biol Chem278, 3671 - 8.
6. Sou, Y.S. et al. (2008) Mol Biol Cell19, 4762 - 75.

Application References

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Companion Products

• 7071 Phototope®-HRP Western Blot Detection System, Anti-rabbit IgG, HRP-linked Antibody
• 5299 Atg4B Antibody
• 7727 Biotinylated Protein Ladder Detection Pack
• 7720 Prestained Protein Marker, Broad Range (Premixed Format)
• 6325 SignalSilence® Atg4C siRNA I

This product is for in vitro research use only and is not intended for use in humans or animals. This product is not intended for use as therapeutic or in diagnostic procedures.