Catalogo Articoli (Spogli Riviste)

OPAC HELP

Titolo:
A novel assay to study autophagy: regulation of autophagosome vacuole sizeby amino acid deprivation
Autore:
Munafo, DB; Colombo, MI;
Indirizzi:
Univ Nacl Cuyo, CONICET, Fac Ciencias Med, Inst Histol & Embriol,Lab Biol Celular & Mol, RA-5500 Mendoza, Argentina Univ Nacl Cuyo Mendoza Argentina RA-5500 Mol, RA-5500 Mendoza, Argentina
Titolo Testata:
JOURNAL OF CELL SCIENCE
fascicolo: 20, volume: 114, anno: 2001,
pagine: 3619 - 3629
SICI:
0021-9533(200110)114:20<3619:ANATSA>2.0.ZU;2-0
Fonte:
ISI
Lingua:
ENG
Soggetto:
ISOLATED RAT HEPATOCYTES; PROTEIN CONJUGATION SYSTEM; SACCHAROMYCES-CEREVISIAE; VESICULAR TRANSPORT; MEMBRANE-FUSION; HT-29 CELLS; YEAST; DEGRADATION; PATHWAY; SEQUESTRATION;
Keywords:
autophagy; vinblastine; starvation; NEM-sensitive protein; PI3-kinase; LC3;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Life Sciences
Citazioni:
52
Recensione:
Indirizzi per estratti:
Indirizzo: Colombo, MI Univ Nacl Cuyo, CONICET, Fac Ciencias Med, Inst Histol & Embriol,Lab Biol Celular & Mol, RA-5500 Mendoza, Argentina Univ Nacl Cuyo Mendoza Argentina RA-5500 Mendoza, Argentina
Citazione:
D.B. Munafo e M.I. Colombo, "A novel assay to study autophagy: regulation of autophagosome vacuole sizeby amino acid deprivation", J CELL SCI, 114(20), 2001, pp. 3619-3629

Abstract

Autophagy is a normal degradative pathway that involves the sequestration of cytoplasmic portions and intracellular organelles in a membrane vacuole called the autophagosome. These vesicles fuse with lysosomes and the sequestered material is degraded. Owing to the complexity of the autophagic pathway and to its inaccessibility to external probes, little is known about themolecular mechanisms that regulate autophagy in higher eukaryotic cells. We used the autofluorescent drug monodansylcadaverine (MDC), a specific autophagolysosome marker to analyze at the molecular level the machinery involved in the autophagic process. We have developed a morphological and biochemical assay to study authophagy in living cells based on the incorporation of MDC. With this assay we observed that the accumulation of MDC was specifically induced by amino acid deprivation and was inhibited by 3-methlyadenine, a classical inhibitor of the autophagic pathway. Additionally, wortmannin, an inhibitor of PI3-kinases that blocks autophagy at an early stage, inhibited the accumulation of MDC in autophagic vacuoles. We also found that treatment of the cells with N-ethylmaleimide (NEM), an agent known to inhibit several vesicular transport events, completely blocked the incorporation of MDC, suggesting that an NEM-sensitive protein is required for the formation of autophagic vacuoles. Conversely, vinblastine, a microtubule depolymerizing agent that induces the accumulation of autophagic vacuoles by preventing their degradation, increased the accumulation of MDC and altered the distribution and size of the autophagic vacuoles. Our results indicate that in the presence of vinblastine very large MDC-vacuoles accumulated mainly under starvation conditions, indicating that the expansion of autophagosomesis upregulated by amino acid deprivation. Furthermore, these MDC-vacuoles were labeled with LC3, one of the mammalian homologues of the yeast proteinApg8/Aut7 that plays an important role in autophagosome formation.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 26/01/21 alle ore 04:12:26