Toxoplasma gondii metacaspase 2 is an important factor that influences bradyzoite formation in the Pru strain

Abstract

Toxoplasma gondii is an important zoonotic protozoan of the phylum Apicomplexa that can infect nearly all warm-blooded animals. The parasite can exist as the interconvertible tachyzoite or bradyzoite forms, leading to acute or latent infection, respectively. No drug has been reported to penetrate the cyst wall and reduce bradyzoite survival and proliferation till now. The transcriptional level of metacaspases 2 (TgMCA2) in T. gondii is significantly upregulated during the formation of bradyzoites in the Pru strain, indicating that it may play an important role in the formation of bradyzoites. To further explore the function of TgMCA2, we constructed a TgMCA2 gene-knockout variant of the Pru strain (Δmca2). Comparative analysis revealed that the proliferative capacity of Pru Δmca2 increased, while the invasion and egressing properties were not affected by the knockout. Further data shows that the tachyzoites of Δmca2 failed to induce differentiation and form bradyzoites in vitro, and the transcriptional levels of some of the bradyzoite-specific genes (such as BAG1, LDH2, and SAG4A) in Δmca2 were significantly lower compared with that in the Pru strain at the bradyzoite stage. In vivo, no cysts were detected in Δmca2-infected mice. Further determination of parasite burden in Δmca2- and Pru-infected mice brain tissue at the genetic level showed that the gene load was significantly lower than that in Pru. In summary, we confirmed that TgMCA2 contributes to the formation of bradyzoites, and could provide an important foundation for the development of attenuated vaccines for the prevention of T. gondii infection.

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Data availability

All datasets generated for this study are included in the manuscript/supplementary files.

Abbreviations

Pru:

T. gondii type II Prugniuad (Pru) strains

MCA:

metacaspases

Δmca2 :

TgMCA2 gene-knockout variant of the Pru strain

BAG1:

bradyzoite-specific gene 1

LDH2:

lactate dehydrogenase 2

SAG4A:

surface antigen protein 4A

HFFs:

human foreskin fibroblasts

FBS:

fetal bovine serum

DMEM:

Dulbecco’s modified Eagle’s medium

DHFR:

dihydrofolate reductase

IFA:

immunofluorescence assay

CST1:

cyst wall glycoprotein

BPK1:

bradyzoite pseudokinase

NST1:

nucleotide sugar transporter

GRA:

dense granular proteins

ROP:

rhoptry proteins

AP2:

transcription factors

BFD1:

Myb-like transcription factor

ENO:

enolase

CDPK2:

calcium-dependent protein kinase

eIF2:

eukaryotic initiation factor

PKAc3:

protein kinase A catalytic subunit 3

VAC:

vacuolar compartment

CPL:

cathepsin

CRT:

chloroquine resistance transporter

IMC1:

inner membrane complex 1

GAP45:

glideosome-associated protein 45

PV:

parasitophorous vacuole

SAG1:

surface antigen 1

DBA:

Dolichos biflorus agglutinin

HA:

hemagglutinin

Δmca2-cm:

TgMCA2-deficient complementary strain

UPRT:

uracil phosphoribosyltransferase

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Acknowledgments

We would like to show our appreciation to Yong Fu, Xiao Zhang, and Ying Xu (China Agricultural University, China) for their help and suggestions.

Code availability

Not applicable.

Funding

This study was supported by the National Key Research and Development Program of China (2017YFD0501304) and the National Natural Science Foundation of China (31672544, 31730096,81971964).

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Contributions

LQ, LJ, and LMY conceived and designed the study. LMY and LMZ performed the experiments. SXJ analyzed the data and drafted the manuscript. YX and LJ helped in manuscript writing. All authors read and approved the final manuscript.

Corresponding authors

Correspondence to Jing Liu or Qun Liu.

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Ethics statement

Our research with all animals were approved by the Beijing Laboratory Administration Committee in accordance with the recommendations in the Guide for the Care and Use of Laboratory Animals of the Ministry of Science and Technology of China (Approval No.: 18049). The mice were humanely euthanized when they reached the end of the experiment.

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Song, X., Lin, M., Li, M. et al. Toxoplasma gondii metacaspase 2 is an important factor that influences bradyzoite formation in the Pru strain. Parasitol Res 119, 2287–2298 (2020). https://doi.org/10.1007/s00436-020-06722-3

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Keywords

  • Toxoplasma gondii
  • Differentiation
  • Bradyzoite
  • Cyst
  • Metacaspase 2
  • Bradyzoite-specific genes