Objective

The pioneer transcription factor DUXC is essential for early bovine embryonic development, where it triggers embryonic genome activation (EGA) and initiates the first wave of transcription. Despite its crucial role, the molecular mechanisms by which DUXC remodels chromatin and whether it is sufficient to reprogram the epigenome independently remain largely unclear.

Our project aims to elucidate these mechanisms using bovine embryonic stem cells (bESCs) as an in vitro model. Specifically, we seek to decipher how DUXC activates early embryonic transcriptional programs and whether it can redirect bESCs toward a totipotent-like state, thereby mimicking the transcriptional and epigenetic landscape of preimplantation embryos.

Methods

An inducible DUXC expression system is being engineered in bovine embryonic stem cells (iDUXC-bESCs). We will combine ATAC-Seq to assess histone modifications and chromatin accessibility, along with RNA-Seq to quantify the transcriptional responses of molecular targets. This approach will also enable us to assess the reliability of using the identified DNA-binding motif of DUX4, the human ortholog of DUXC, for predicting DUXC targets. This complements the identification of potential DUXC targets, which have been identified by RNA-Seq following the inhibition of DUXC translation using interfering RNA.

Results

We anticipate that DUXC induction will elicit a robust activation of cleavage-stage genes and retrotransposons, mirroring epigenomic features of early embryos, and reprogram ESCs into a transcriptional and epigenetic state resembling totipotency. We anticipate identifying a defined set of direct DUXC targets, along with enriched histone acetylation at key regulatory loci.

Conclusion

This study will provide direct mechanistic evidence that DUXC governs chromatin accessibility and initiates embryonic genome activation. It will establish bovine iDUXC-bESCs as a tractable model for studying totipotency and early lineage programming. These findings will advance our understanding of genome reprogramming and pioneer factor function in mammals.