Abstract
Apart from their roles as chaperones, heat shock proteins are involved in other vital activities including apoptosis with mammalian Hsp60 being ascribed proapoptotic as well as antiapoptotic roles. Using conditional RNAi or overexpression of Hsp60D, a member of the Hsp60 family in Drosophila melanogaster, we show that the downregulation of this protein blocks caspase-dependent induced apoptosis. GMR-Gal4-driven RNAi for Hsp60D in developing eyes dominantly suppressed cell death caused by expression of Reaper, Hid, or Grim (RHG), the key activators of canonical cell death pathway. Likewise, Hsp60D-RNAi rescued cell death induced by GMR-Gal4-directed expression of full-length and activated DRONC. Overexpression of Hsp60D enhanced cell death induced either by directed expression of RHG or DRONC. However, the downregulation of Hsp60D failed to suppress apoptosis caused by unguarded caspases in DIAP1-RNAi flies. Furthermore, in DIAP1-RNAi background, Hsp60D-RNAi also failed to inhibit apoptosis induced by RHG expression. The Hsp60 and DIAP1 show diffuse and distinct granular overlapping distributions in the photoreceptor cells with the bulk of both proteins being outside the mitochondria. Depletion of either of these proteins disrupts the granular distribution of the other. We suggest that in the absence of Hsp60D, DIAP1 is unable to dissociate from effecter and executioner caspases, which thus remain inactive.
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Acknowledgements
We thank Drs. M. Freeman (Cambridge, UK), B. Hay, and I. Muro (California Institute of Technology, USA), P. Meier and G.I. Evan (Breakthrough Toby Robins Breast Cancer Research Centre, UK), M. Miura (Brain Science Institute, RIKEN), Dr. V. Sriram (National Centre for Biological Sciences, Bangalore, India) and the Bloomington Fly Stock Centre (Indiana, USA) for providing different fly stocks used in this study. We also thank Dr. K. White (CBRC, Massachusetts General Hospital) and Dr. S. Ganesh (IIT Kanpur, India) for providing DIAP1 and Grp75 antibodies, respectively. We thank Dr. L. S. Shashidhara for his help in the generation of the Hsp60D transgenic flies at the Centre for Cellular and Molecular Biology (Hyderabad). This work was supported by a research grant from the Department of Biotechnology, Government of India, N. Delhi, to SCL. The Laser Scanning Confocal Microscope Facility is supported by the Department of Science and Technology, Government of India, N. Delhi. RA is supported by a research fellowship from the Council of Scientific and Industrial Research, N. Delhi.
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Arya, R., Lakhotia, S.C. Hsp60D is essential for caspase-mediated induced apoptosis in Drosophila melanogaster . Cell Stress and Chaperones 13, 509–526 (2008). https://doi.org/10.1007/s12192-008-0051-3
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DOI: https://doi.org/10.1007/s12192-008-0051-3