Disruption of a mitochondrial protease machinery in Plasmodium falciparum is an intrinsic signal for parasite cell death

S Rathore,A Srivastava,V S Chauhan,D Sinha,M Gupta,M S Narayanan,S Jain,M Asad,D Gupta,G Ramasamy,A Mohmmed

doi:10.1038/cddis.2011.118

Abstract

The ATP-dependent ClpQY protease system in Plasmodium falciparum is a prokaryotic machinery in the parasite. In the present study, we have identified the complete ClpQY system in P. falciparum and elucidated its functional importance in survival and growth of asexual stage parasites. We characterized the interaction of P. falciparum ClpQ protease (PfClpQ) and PfClpY ATPase components, and showed that a short stretch of residues at the C terminus of PfClpY has an important role in this interaction; a synthetic peptide corresponding to this region antagonizes this interaction and interferes with the functioning of this machinery in the parasite. Disruption of ClpQY function by this peptide caused hindrance in the parasite growth and maturation of asexual stages of parasites. Detailed analyses of cellular effects in these parasites showed features of apoptosis-like cell death. The peptide-treated parasites showed mitochondrial dysfunction and loss of mitochondrial membrane potential. Dysfunctioning of mitochondria initiated a cascade of reactions in parasites, including activation of VAD–FMK-binding proteases and nucleases, which resulted in apoptosis-like cell death. These results show functional importance of mitochondrial proteases in the parasite and involvement of mitochondria in programmed cell death in the malaria parasites.

Highlights

ATP-dependent protease machineries including the eukaryotic 26S proteasome and the prokaryotic casenolytic proteases (Clp) systems are large protein-degradation complexes that have an essential role in cell-cycle regulation.[6,7] The ClpQY machinery is a multimeric ATP-dependent protease system in prokaryotes that resembles the eukaryotic 26S proteasome; it consists of two stacked hexameric rings of ClpQ protease that are capped on one of both sides with a hexameric ring of AAA-type ATPase, ClpY.[8]
The westernblot analysis using anti-P. falciparum ClpQ (PfClpQ) antibodies detected the pro-PfClpQ (B22 kDa) and mature-PfClpQ (B18 kDa) in trophozoite- and schizont- stage parasites with maximum expression in schizont stage. These results suggest that both PfClpQ and P. falciparum ClpY orthologue (PfClpY) are expressed in the blood-stage parasites in the late-trophozoite and schizont stages
Our results show that the ClpQ and ClpY protein in P. falciparum are part of a protein complex as shown in prokaryotes.[6,7,8]

Summary

Introduction

ATP-dependent protease machineries including the eukaryotic 26S proteasome and the prokaryotic casenolytic proteases (Clp) systems are large protein-degradation complexes that have an essential role in cell-cycle regulation.[6,7] The ClpQY machinery is a multimeric ATP-dependent protease system in prokaryotes that resembles the eukaryotic 26S proteasome; it consists of two stacked hexameric rings of ClpQ protease that are capped on one of both sides with a hexameric ring of AAA-type ATPase, ClpY.[8]. Disruption of ClpQY activity using a small peptide that interferes with functioning of this machinery, caused dysregulation of mitochondrial functions, which in turn activated cascade of reactions leading to apoptosis-like cell death of the parasite. These results suggest role of mitochondrial proteases in maintenance of functional mitochondria and points towards involvement of mitochondria in apoptosis-like cell death in parasite

Methods

Results

Conclusion