ApiAP2 transcription factor restricts development of the Toxoplasma tissue cyst

Joshua B. Radke, Olivier Lucas, Erandi K. De Silva, Yanfen Ma, William Sullivan, Louis M. Weiss, Manuel Llinas, Michael W. White

Research output: Contribution to journalArticle

44 Citations (Scopus)

Abstract

Cellular differentiation leading to formation of the bradyzoite tissue cyst stage is the underlying cause of chronic toxoplasmosis. Consequently, mechanisms responsible for controlling development in the Toxoplasma intermediate life cycle have long been sought. Here, we identified 15 Toxoplasma mRNAs induced in early bradyzoite development that encode proteins with apicomplexan AP2 (ApiAP2) DNA binding domains. Of these 15 mRNAs, the AP2IX-9 mRNA demonstrated the largest expression increase during alkaline-induced differentiation. At the protein level, we found that AP2IX-9 was restricted to the early bradyzoite nucleus and is repressed in tachy-zoites and in mature bradyzoites from 30-d infected animals. Conditional overexpression of AP2IX-9 significantly reduced tissue cyst formation and conferred alkaline pH-resistant growth, whereas disruption of the AP2IX-9 gene increased tissue cyst formation, indicating AP2IX-9 operates as a repressor of bradyzoite development. Consistent with a role as a repressor, AP2IX-9 specifically inhibited the expression of bradyzoite mRNAs, including the canonical brady-zoite marker, bradyzoite antigen 1 (BAG1). Using protein binding microarrays, we established the AP2 domain of AP2IX-9 binds a CAGTGT DNA sequence motif and is capable of binding cis-regulatory elements controlling the BAG1 and bradyzoite-specific nucle-oside triphosphatase (B-NTPase) promoters. The effect of AP2IX-9 on BAG1 expression was direct because this factor inhibits expression of a firefly luciferase reporter under the control of the BAG1 promoter in vivo, and epitope-tagged AP2IX-9 can be immunopre-cipitated with the BAG1 promoter in parasite chromatin. Altogether, these results indicate AP2IX-9 restricts Toxoplasma commitment to develop the mature bradyzoite tissue cyst.

Original languageEnglish
Pages (from-to)6871-6876
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume110
Issue number17
DOIs
StatePublished - Apr 23 2013

Fingerprint

Transcription Factor AP-2
Toxoplasma
Cysts
Antigens
Messenger RNA
Firefly Luciferases
Protein Array Analysis
Nucleotide Motifs
Toxoplasmosis
Differentiation Antigens
Life Cycle Stages
Protein Binding
Chromatin
Epitopes
Parasites
Proteins
DNA
Growth
Genes

Keywords

  • Apicomplexa
  • Gene expression
  • Gene regulation

ASJC Scopus subject areas

  • General

Cite this

ApiAP2 transcription factor restricts development of the Toxoplasma tissue cyst. / Radke, Joshua B.; Lucas, Olivier; De Silva, Erandi K.; Ma, Yanfen; Sullivan, William; Weiss, Louis M.; Llinas, Manuel; White, Michael W.

In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 110, No. 17, 23.04.2013, p. 6871-6876.

Research output: Contribution to journalArticle

Radke, Joshua B. ; Lucas, Olivier ; De Silva, Erandi K. ; Ma, Yanfen ; Sullivan, William ; Weiss, Louis M. ; Llinas, Manuel ; White, Michael W. / ApiAP2 transcription factor restricts development of the Toxoplasma tissue cyst. In: Proceedings of the National Academy of Sciences of the United States of America. 2013 ; Vol. 110, No. 17. pp. 6871-6876.
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T1 - ApiAP2 transcription factor restricts development of the Toxoplasma tissue cyst

AU - Radke, Joshua B.

AU - Lucas, Olivier

AU - De Silva, Erandi K.

AU - Ma, Yanfen

AU - Sullivan, William

AU - Weiss, Louis M.

AU - Llinas, Manuel

AU - White, Michael W.

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N2 - Cellular differentiation leading to formation of the bradyzoite tissue cyst stage is the underlying cause of chronic toxoplasmosis. Consequently, mechanisms responsible for controlling development in the Toxoplasma intermediate life cycle have long been sought. Here, we identified 15 Toxoplasma mRNAs induced in early bradyzoite development that encode proteins with apicomplexan AP2 (ApiAP2) DNA binding domains. Of these 15 mRNAs, the AP2IX-9 mRNA demonstrated the largest expression increase during alkaline-induced differentiation. At the protein level, we found that AP2IX-9 was restricted to the early bradyzoite nucleus and is repressed in tachy-zoites and in mature bradyzoites from 30-d infected animals. Conditional overexpression of AP2IX-9 significantly reduced tissue cyst formation and conferred alkaline pH-resistant growth, whereas disruption of the AP2IX-9 gene increased tissue cyst formation, indicating AP2IX-9 operates as a repressor of bradyzoite development. Consistent with a role as a repressor, AP2IX-9 specifically inhibited the expression of bradyzoite mRNAs, including the canonical brady-zoite marker, bradyzoite antigen 1 (BAG1). Using protein binding microarrays, we established the AP2 domain of AP2IX-9 binds a CAGTGT DNA sequence motif and is capable of binding cis-regulatory elements controlling the BAG1 and bradyzoite-specific nucle-oside triphosphatase (B-NTPase) promoters. The effect of AP2IX-9 on BAG1 expression was direct because this factor inhibits expression of a firefly luciferase reporter under the control of the BAG1 promoter in vivo, and epitope-tagged AP2IX-9 can be immunopre-cipitated with the BAG1 promoter in parasite chromatin. Altogether, these results indicate AP2IX-9 restricts Toxoplasma commitment to develop the mature bradyzoite tissue cyst.

AB - Cellular differentiation leading to formation of the bradyzoite tissue cyst stage is the underlying cause of chronic toxoplasmosis. Consequently, mechanisms responsible for controlling development in the Toxoplasma intermediate life cycle have long been sought. Here, we identified 15 Toxoplasma mRNAs induced in early bradyzoite development that encode proteins with apicomplexan AP2 (ApiAP2) DNA binding domains. Of these 15 mRNAs, the AP2IX-9 mRNA demonstrated the largest expression increase during alkaline-induced differentiation. At the protein level, we found that AP2IX-9 was restricted to the early bradyzoite nucleus and is repressed in tachy-zoites and in mature bradyzoites from 30-d infected animals. Conditional overexpression of AP2IX-9 significantly reduced tissue cyst formation and conferred alkaline pH-resistant growth, whereas disruption of the AP2IX-9 gene increased tissue cyst formation, indicating AP2IX-9 operates as a repressor of bradyzoite development. Consistent with a role as a repressor, AP2IX-9 specifically inhibited the expression of bradyzoite mRNAs, including the canonical brady-zoite marker, bradyzoite antigen 1 (BAG1). Using protein binding microarrays, we established the AP2 domain of AP2IX-9 binds a CAGTGT DNA sequence motif and is capable of binding cis-regulatory elements controlling the BAG1 and bradyzoite-specific nucle-oside triphosphatase (B-NTPase) promoters. The effect of AP2IX-9 on BAG1 expression was direct because this factor inhibits expression of a firefly luciferase reporter under the control of the BAG1 promoter in vivo, and epitope-tagged AP2IX-9 can be immunopre-cipitated with the BAG1 promoter in parasite chromatin. Altogether, these results indicate AP2IX-9 restricts Toxoplasma commitment to develop the mature bradyzoite tissue cyst.

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KW - Gene expression

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