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Autophagy. 2009 Oct;5(7):1018-25. Epub 2009 Oct 6.

Identification of novel autophagy regulators by a luciferase-based assay for the kinetics of autophagic flux.

Author information

1
Apoptosis Department and Centre for Genotoxic Stress Research, Institute of Cancer Biology, Danish Cancer Society, Copenhagen, Denmark.

Abstract

Macroautophagy (hereafter referred to as autophagy) has recently emerged as an attractive target for the treatment of various degenerative diseases and cancer. The discovery of effective pharmaceutical regulators of autophagy has, however, been hindered by a lack of feasible assay systems for autophagic flux. Here, we present a luciferase-based reporter assay that measures autophagic flux in real time in living cells and demonstrate that this assay system is apt for the detection of dose- and stimulus-dependent differences in autophagy kinetics. Furthermore, by screening a small molecule kinase inhibitor library containing 80 compounds we identified 12 compounds as inducers of autophagic flux. Importantly, six inhibitors of the class I phosphoinositide 3-kinase -- protein kinase B -- mammalian target of rapamycin complex 1 axis, the central signaling pathway repressing autophagy, scored as autophagy inducers adequately validating the screen. We conclude that the assay system presented here allows easy and rapid monitoring of autophagy kinetics and is suitable for screening of small molecule libraries.

PMID:
19652534
DOI:
10.4161/auto.5.7.9443
[Indexed for MEDLINE]

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