Ascorbate peroxidase from Leishmania major controls the virulence of infective stage of promastigotes by regulating oxidative stress.
BACKGROUND
Peroxidase represents a heterogeneous group of distinct enzyme family that plays extremely diverse biological functions. Ascorbate peroxidase from Leishmania major (LmAPX) has been shown to be central to the redox defense system of Leishmania. To investigate further its exact physiological role in Leishmania, we attempted to create LmAPX -knockout mutants by gene replacement in L. major strains.


RESULTS
The null mutant cell culture contains a higher percentage of metacyclic and apoptotic cells compared to both wild type and LmAPX overexpressing cells. Flowcytometric analysis reveals the presence of a higher concentration of intracellular H(2)O(2), indicative of increased oxidative stress in parasites lacking LmAPX. IC(50) value for exogenously added H(2)O(2) shows that deletion of LmAPX in L. major renders the cell more susceptible to H(2)O(2). Real time PCR studies demonstrate an elevated mRNA level of non-selenium glutathione peroxidase in LmAPX null mutant cell line, suggesting that these enzymes were induced to compensate the LmAPX enzyme. The null mutant cells exhibit hypervirulence after infection with macrophages as well as inoculation into BALB/c mice; in contrast, overexpressing cells show avirulence.


CONCLUSIONS
Collectively, these data provide strong evidence that LmAPX is an important factor for controlling parasite differentiation and survival within macrophages.
Pal
Swati
S
Division of Structural Biology & Bio-Informatics, Indian Institute of Chemical Biology, Kolkata, West Bengal, India.
Dolai
Subhankar
S
Yadav
Rajesh K
RK
Adak
Subrata
S
D000483:Alleles; D000818:Animals; D060387:Ascorbate Peroxidases; D018320:Leishmania major / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development / Q000472:pathogenicity; D009154:Mutation; D018384:Oxidative Stress; D010544:Peroxidases / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics; D017382:Reactive Oxygen Species / Q000378:metabolism; D014774:Virulence
D012333:RNA, Messenger; D017382:Reactive Oxygen Species; D010544:Peroxidases; D060387:Ascorbate Peroxidases