Molecular Chaperone Hsp70 and Its Constitutively Active Form Hsc70 Play an Indispensable Role During Eye Development of Drosophila melanogaster
- 8.2k Downloads
In the present study, we demonstrate that molecular chaperone Hsp70 and Hsc70 is essential for normal organization and development of ommatidial cells in Drosophila melanogaster eye. An exogenously expressed dominant negative mutant of Hsp70 (K71E) and Hsc70.4 (K71S and D206S) in an eye-specific manner resulted in eye degeneration that includes loss of eye pigment, disorganized ommatidia, abnormality in bristle cell arrangement and reduction in the eye size. The developmental organization of ommatidial cells (cone, photoreceptor, pigment, and bristle cell complex) was disturbed in Hsp70 and Hsc70 mutants. Acridine orange (AO) and caspase 3 staining showed an increased cell death in Hsp70 and Hsc70 mutant eyes. Genetic interaction study of Hsp70 and Hsc70 mutants with candidate genes of JNK signaling pathway and immunocytochemistry study using phospho-JNK antibody suggested that mutation in Hsp70 and Hsc70 results in ectopic activation of JNK signaling in fly eye. Further, anti-PH3 staining in Hsp70 and Hsc70 mutant eyes revealed a reduced number of mitotic cells in second mitotic wave (SMW) of developing eye and anti-Rh1 staining showed reduced Rh1 expression, accumulation of Rh1 in the cytoplasm, and rhabdomere degeneration. Thus, on the basis of results, it was concluded that molecular chaperone Hsp70 and Hsc70 play an indispensable role during Drosophila eye development.
KeywordsDominant negative mutation Hsp70 (K71E) Hsc70 (K71S) Drosophila eye F-actin accumulation Excessive cell death Caspase activation JNK signaling
We thank Prof. J. K. Roy (Banaras Hindu University, Varanasi, India), Prof. Nancy M. Bonini (Howard Hughes Medical Institute) for UAS-Hsp70 K71E , Prof. Drick Bohman (University of Rochester Medical Center, Rochester, NY USA) for UAS-Bsk DN , Prof. Martinez-Arias (University of Cambridge, UK) for puc E69 , Prof. M. Miura (Graduate Institute of Pharmaceuticals Sciences, University of Tokyo) for UAS-eiger, and Bloomington Drosophila stock Centre (Indiana University, 1001 E. Third St. Bloomington, USA) for fly stocks. We also thank Prof. Patric Dolph (Dart Morth College Medical School, USA) for anti-Rh1 antibody. The financial assistance from Department of Science & Technology (DST), New Delhi, India, to AKT (SR/FT/LS-1/2010), Laser Scanning Confocal Microscope facility supported by the Department of Biotechnology (DBT), India, and support from The Puri Foundation for Education in India is duly acknowledged.
Conceived and designed the experiments: AK and AKT. Performed the experiments: AK. Analyzed the data: AK and AKT. Wrote the paper: AK and AKT. All authors reviewed the manuscript.
Compliance with Ethical Standards
This work was supported by the Department of Science and Technology (DST), New Delhi, India, to AKT (SR/FT/LS-1/2010).
Conflict of Interest
The authors declare that they have no conflicts of interest.
- 12.Shukla AK, Pragya P, Chaouhan HS, Tiwari AK, Patel DK, Abdin MZ, Chowdhuri DK Heat shock protein-70 (Hsp-70) suppresses paraquat-induced neurodegeneration by inhibiting JNK and caspase-3 activation in Drosophila model of Parkinson's disease. PLoS One 9(6):e98886. doi: 10.1371/journal.pone.0098886
- 16.Shukla AK, Pragya P, Chaouhan HS, Tiwari AK, Patel DK, Abdin MZ, Chowdhuri DK (2014) Heat shock protein-70 (Hsp-70) suppresses paraquat-induced neurodegeneration by inhibiting JNK and caspase-3 activation in Drosophila model of Parkinson’s disease. PLoS One 9(6):e98886. doi: 10.1371/journal.pone.0098886 CrossRefPubMedPubMedCentralGoogle Scholar
- 31.Gorbatyuk MS, Knox T, LaVail MM, Gorbatyuk OS, Noorwez SM, Hauswirth WW, Lin JH, Muzyczka N, Lewin AS (2010) Restoration of visual function in P23H rhodopsin transgenic rats by gene delivery of BiP/Grp78. Proc Natl Acad Sci U S A 107(13):5961–5966. doi: 10.1073/pnas.0911991107
- 32.Athanasiou D, Kosmaoglou M, Kanuga N, Novoselov SS, Paton AW, Paton JC, Chapple JP, Cheetham ME (2012) BiP prevents rod opsin aggregation. Mol Biol Cell 23(18):3522–3531. doi: 10.1091/mbc.E12-02-0168
- 40.Ouweneel WJ (1970) Normal and abnormal determination in the imaginal discs of Drosophila, with special reference to the eye discs. Acta Embryol Exp (Palermo) 1:95–119Google Scholar
- 58.Arya R, Lakhotia SC (2006) A simple nail polish imprint technique for examination of external morphology of Drosophila eyes. Curr Sci 90(9):1179–1180Google Scholar
- 74.Riggs B, Rothwell W, Mische S, Hickson GR, Matheson J, Hays TS, Gould GW, Sullivan W (2003) Actin cytoskeleton remodeling during early Drosophila furrow formation requires recycling endosomal components nuclear-fallout and Rab11. J Cell Biol 163(1):143–154. doi: 10.1083/jcb.200305115 CrossRefPubMedPubMedCentralGoogle Scholar
- 77.Mitra A, Chinchore Y, Kinser R, Dolph PJ Characterization of two dominant alleles of the major rhodopsin-encoding gene ninaE in Drosophila. Mol Vis 17:3224–3233Google Scholar