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Inducible Calling Cards: Developing Mouse Reagents for Experimentally Controlled Transposon Insertion in vivo.

Simona Sarafinovska1,2, Arthi Venkatesan1,2, Titilope Akinwe1,2

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We developed inducible Calling Cards (iCC) in mice for precise control of DNA recording. Jun-iCC captured neural activity, while Sp1-iCC showed higher insertions but caused developmental issues.

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Area of Science:

  • Genetics
  • Neuroscience
  • Molecular Biology

Background:

  • The piggyBac transposase facilitates genetic screens in mice.
  • Fusions of piggyBac with transcription factors (TFs) create Calling Cards to mark DNA binding sites.
  • Inducible transposases are needed for precise temporal control of recording in vivo.

Purpose of the Study:

  • To engineer and test inducible Calling Cards (iCC) in the murine brain for drug-inducible DNA recording.
  • To assess the efficacy and temporal control of Jun-iCC and Sp1-iCC systems.
  • To evaluate potential developmental impacts of inducible TF-transposase fusions.

Main Methods:

  • Engineered knock-in mice with tamoxifen-inducible TF-transposase fusions (Jun-iCC and Sp1-iCC).
  • Tested iCC systems in vitro and in vivo for insertion efficacy and temporal control.
  • Assessed mouse development, behavior, and neural activity recording.

Main Results:

  • Jun-iCC mice showed tamoxifen-dependent recording and captured neural activity during seizures, but with low insertion numbers.
  • Sp1-iCC resulted in higher insertion numbers but caused developmental abnormalities like reduced viability and anophthalmia.
  • The ERT2 domain's interaction with Sp1 may underlie observed developmental defects.

Conclusions:

  • Inducible Calling Cards enable drug-inducible transposon integration in vivo.
  • Jun-iCC is suitable for recording transient neural activity, despite low insertion rates.
  • Sp1-iCC requires further optimization to mitigate developmental side effects while leveraging its high insertion capacity.