Cell Signaling Technology

Product Pathways - Autophagy Signaling

LC3 Control Cell Extracts #11972

Description

Total cell extracts from HeLa cells were untreated or treated with 50 μM chloroquine overnight. Supplied in SDS Sample Buffer. Boil for 2 minutes in the original tube, then load 10 μl per mini-gel lane.

Western Blotting

Western Blotting

Western blot analysis of LC3 Control Cell Extracts from HeLa cells, untreated (-) or chloroquine-treated (50 μM, overnight; +), using LC3B (D11) XP® Rabbit mAb #3868 (upper), LC3A/B Antibody #4108 (middle), or β-Tubulin (9F3) Rabbit mAb #2128 (lower).

Applications

Western Blots: CST recommends boiling control cell extracts for two minutes prior to loading. Load 10 µl of each cell extract per lane.

Background

Autophagy is a catabolic process for the autophagosomic-lysosomal degradation of bulk cytoplasmic contents (1,2). Autophagy is generally activated by conditions of nutrient deprivation, but it has also been associated with a number of physiological processes including development, differentiation, neurodegenerative diseases, infection, and cancer (3). Autophagy marker Light Chain 3 (LC3) was originally identified as a subunit of microtubule-associated proteins 1A and 1B (termed MAP1LC3) (4) and subsequently found to contain similarity to the yeast protein Apg8/Aut7/Cvt5 critical for autophagy (5). Three human LC3 isoforms (LC3A, LC3B, and LC3C) undergo post-translational modifications during autophagy (6-9). Cleavage of LC3 at the carboxy terminus immediately following synthesis yields the cytosolic LC3-I form. During autophagy, LC3-I is converted to LC3-II through lipidation by a ubiquitin-like system involving Atg7 and Atg3 that allows for LC3 to become associated with autophagic vesicles (6-10). The presence of LC3 in autophagosomes and the conversion of LC3 to the lower migrating form, LC3-II, have been used as indicators of autophagy (11).

  1. Reggiori, F. and Klionsky, D.J. (2002) Eukaryot. Cell 1, 11-21.
  2. Codogno, P. and Meijer, A.J. (2005) Cell Death Differ. 12 Suppl 2, 1509-1518.
  3. Levine, B. and Yuan, J. (2005) J. Clin. Invest. 115, 2679-2688.
  4. Mann, S.S. and Hammarback, J.A. (1994) J. Biol. Chem. 269, 11492-11497.
  5. Lang, T. et al. (1998) EMBO J. 17, 3597-3607.
  6. Kabeya, Y. et al. (2000) EMBO J. 19, 5720-5728.
  7. He, H. et al. (2003) J. Biol. Chem. 278, 29278-29287.
  8. Tanida, I. et al. (2004) J. Biol. Chem. 279, 47704-47710.
  9. Wu, J. et al. (2006) Biochem. Biophys. Res. Commun. 339, 437-442.
  10. Ichimura, Y. et al. (2000) Nature 408, 488-492.
  11. Kabeya, Y. et al. (2004) J. Cell Sci. 117, 2805-2812.

Application References

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Companion Products

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

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