Plasmodium Ape1 is a multifunctional enzyme in mitochondrial base excision repair and is required for efficient transition from liver to blood stage infection

DNA Repair (Amst). 2021 May:101:103098. doi: 10.1016/j.dnarep.2021.103098. Epub 2021 Mar 14.

Abstract

The malaria parasite has a single mitochondrion which carries multiple tandem repeats of its 6 kb genome encoding three proteins of the electron transport chain. There is little information about DNA repair mechanisms for mitochondrial genome maintenance in Plasmodium spp. Of the two AP-endonucleases of the BER pathway encoded in the parasite nuclear genome, the EndoIV homolog PfApn1 has been identified as a mitochondrial protein with restricted functions. We explored the targeting and biochemical properties of the ExoIII homolog PfApe1. PfApe1 localized in the mitochondrion and exhibited AP-site cleavage, 3'-5' exonuclease, 3'-phosphatase, nucleotide incision repair (NIR) and RNA cleavage activities indicating a wider functional role than PfApn1. The parasite enzyme differed from human APE1 in possessing a large, disordered N-terminal extension. Molecular modelling revealed conservation of structural domains but variations in DNA-interacting residues and an insertion in the α-8 loop suggested differences with APE1. Unlike APE1, where AP-site cleavage and NIR activities could be mutually exclusive based on pH and Mg2+ ion concentration, PfApe1 was optimally active under similar conditions suggesting that it can function both as an AP-endonuclease in BER and directly cleave damaged bases in NIR under similar physiological conditions. To investigate the role of Ape1 in malaria life cycle, we disrupted the gene by double-cross-over homologous recombination. Ape1 knockout (KO) P. berghei parasites showed normal development of blood and mosquito stages. However, inoculation of mice with Ape1 KO salivary gland sporozoites revealed a reduced capacity to initiate blood stage infection. Ape1 KO parasites underwent normal liver stage development until merozoites egressed from hepatocytes. Our results indicated that the delay in pre-patent period was due to the inability of Ape1 KO merosomes to infect erythrocytes efficiently.

Keywords: AP-endonuclease; Ape1; BER; DNA repair; Mitochondrion; Plasmodium.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • DNA / metabolism
  • DNA Damage*
  • DNA Repair*
  • DNA-(Apurinic or Apyrimidinic Site) Lyase / metabolism*
  • Humans
  • Kinetics
  • Life Cycle Stages
  • Malaria, Falciparum
  • Mice
  • Mitochondria / enzymology*
  • Mitochondria / genetics
  • Models, Molecular
  • Multifunctional Enzymes
  • Plasmodium berghei
  • Plasmodium falciparum / enzymology*
  • Plasmodium falciparum / genetics
  • Plasmodium falciparum / growth & development
  • Protein Conformation
  • Substrate Specificity

Substances

  • Multifunctional Enzymes
  • DNA
  • DNA-(Apurinic or Apyrimidinic Site) Lyase