Abstract
In the post-genomic perspective, the quest of programmed cell death (PCD) mechanisms in kinetoplastid parasites lies in the identification and characterization of cell death executer proteins. Here, we show that baicalein (BLN), a potent topoisomerase IB inhibitor, generates an oxidative stress in the parasites leading to altered physiological and morphological parameters, which are characteristic of PCD. For the first time we elucidate that, caspase-independent activation of a novel effector molecule, endonuclease G (LdEndoG), mediates BLN-induced cell death. Functional characterization of LdEndoG identifies Flap endonuclease-1 (LdFEN-1) and LdTatD-like nuclease as other effector molecules. BLN treatment translocates LdEndoG from mitochondria to nucleus, where it forms separate complexes with LdFEN-1 and LdTatD to constitute a DNA ‘degradesome’ unique to these parasites. Conditional antisense knockdown of LdEndoG provides protection against PCD. This knowledge paves the path toward a better understanding of the PCD pathway in simpler systems, which could be exploited in anti-leishmanial chemotherapy.
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Abbreviations
- PCD:
-
programmed cell death
- BLN:
-
baicalein
- PI:
-
propidium iodide
- ROS:
-
reactive oxygen species
- CPT:
-
camptothecin
- NAC:
-
N-acetyl cysteine
- TSH:
-
trypanothione
- BAPTA:
-
1,2-bis(o-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid
- ATA:
-
aurin tricarboxylic acid
- Δψm:
-
mitochondrial transmembrane potential
- FITC:
-
fluorescein thiocyanate
- H2DCFDA:
-
5-(and -6)-chloromethyl-2′,7′-dichlorodihydrofluorescein diacetate acetyl ester
- FEN-1:
-
flap endonuclease-1
- CCCP:
-
carbonyl cyanide m-chlorophenyl hydrazone
- zVAD-fmk:
-
Val-Ala-Asp-fluoromethyl ketone
- TUNEL:
-
terminal deoxynucleotidyltransferase enzyme-mediated dUTP end labeling
- DMSO:
-
dimethyl sulfoxide
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Acknowledgements
We are grateful to Professor S Roy, Director, IICB, for his interest in this work. We thank Professor GAM Cross for gifting the Leismania transfection vectors. We thank Arunima Biswas for her help in flow cytometry. This study was supported by the grant from Department of Biotechnology, Government of India (BT/PR6399/BRB/10/434/05) to HKM. SBD was supported by a Senior Research Fellowship from CSIR, Government of India.
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BoseDasgupta, S., Das, B., Sengupta, S. et al. The caspase-independent algorithm of programmed cell death in Leishmania induced by baicalein: the role of LdEndoG, LdFEN-1 and LdTatD as a DNA ‘degradesome’. Cell Death Differ 15, 1629–1640 (2008). https://doi.org/10.1038/cdd.2008.85
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DOI: https://doi.org/10.1038/cdd.2008.85
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