The gold standard for autophagy research

LC3 とLC3 結合反応系 The three proteins, LC3, GABARAP (GABAA receptor-associated protein), and GATE-16 (Golgi-associated ATPase enhancer), are mammalian homologues of yeast Atg8. Among them, LC3 has been studied most extensively and frequently used as an autophagy marker in mammals. Newly translated LC3 (proLC3) is immediately processed at the C-terminus by Atg4B or Atg4A, forming LC3-I. Upon induction of autophagy, LC3-I is sequentially transferred to E1 and E2, and conjugated to the substrate, PE (phosphatidylethanolamine). The resulting PE-conjugated LC3 is called LC3-II. Although LC3-II has a higher molecular weight than LC3-I, the mobility of LC3-II is greater than LC3-I on SDS-PAGE, due to higher hydrophobicity. GABARAP and GATE-16 are also conjugated to PE in a similar process.

CD1d is a membrane protein non-covalently bonded to β2-microglobulin (β2m) and shows high homology between human and mice. CD1d can present α-galactosylceramide (α-GalCer), a glycolipid extracted and isolated from the marine sponge, and this complex can activate human and murine CD1drestricted NKT cells. CD1d Tetramer-PE is a reagent prepared by tetramerization of complexes of CD1d and β2m by PE- or APC- labeled streptavidin. Binding this reagent to α-GalCer enables highly sensitive detection of CD1d-restricted NKT cells and can be combined with antibodies to study NKT cell function by flow cytometry.

Mitophagy is the selective degradation of old or depolarized mitochondria by autophagy and contributes maintaining a healthy population of mitochondria. Since damaged mitochondria lead to collapse cell homeostasis, mitophagy is believed to protect against diseases related to mitochondrial dysfunction such as neurodegenerative disorders.
Parkin, an ubiquitin ligase known as the gene responsible for Parkinson’s disease, plays an important role in autophagic elimination of mitophagy. When mitochondria are depolarized and dysfunctional, PTEN-induced putative kinase protein 1 (PINK1) accumulates on the outer membrane, and recruits Parkin on the damaged mitochondria. The outer membrane on the mitochondria is then ubiquitinated through the ubiquitin ligase activity of Parkin. Finally, the poly- ubiquitinated mitochondria are selectively recognized and executed by autophagic process.

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