Animal Bioscience

Asian Australasian Association of Animal Production Societies (아세아태평양축산학회)
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RNA helicase DEAD-box-5 is involved in R-loop dynamics of preimplantation embryos

  • Hyeonji Lee (Department of Stem Cell and Regenerative Biotechnology, Institute of Advanced Regenerative Science, Konkuk University) ;
  • Dong Wook Han (Guangdong Provincial Key Laboratory of Large Animal Models for Biomedicine, Wuyi University) ;
  • Seonho Yoo (Department of Stem Cell and Regenerative Biotechnology, Institute of Advanced Regenerative Science, Konkuk University) ;
  • Ohbeom Kwon (Department of Stem Cell and Regenerative Biotechnology, Institute of Advanced Regenerative Science, Konkuk University) ;
  • Hyeonwoo La (Department of Stem Cell and Regenerative Biotechnology, Institute of Advanced Regenerative Science, Konkuk University) ;
  • Chanhyeok Park (Department of Stem Cell and Regenerative Biotechnology, Institute of Advanced Regenerative Science, Konkuk University) ;
  • Heeji Lee (Department of Stem Cell and Regenerative Biotechnology, Institute of Advanced Regenerative Science, Konkuk University) ;
  • Kiye Kang (Department of Stem Cell and Regenerative Biotechnology, Institute of Advanced Regenerative Science, Konkuk University) ;
  • Sang Jun Uhm (Department of Animal Science, Sangji University) ;
  • Hyuk Song (Department of Stem Cell and Regenerative Biotechnology, Institute of Advanced Regenerative Science, Konkuk University) ;
  • Jeong Tae Do (Department of Stem Cell and Regenerative Biotechnology, Institute of Advanced Regenerative Science, Konkuk University) ;
  • Youngsok Choi (Department of Stem Cell and Regenerative Biotechnology, Institute of Advanced Regenerative Science, Konkuk University) ;
  • Kwonho Hong (Department of Stem Cell and Regenerative Biotechnology, Institute of Advanced Regenerative Science, Konkuk University)
  • Received : 2023.10.06
  • Accepted : 2023.12.07
  • Published : 2024.06.01
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Abstract

Objective: R-loops are DNA:RNA triplex hybrids, and their metabolism is tightly regulated by transcriptional regulation, DNA damage response, and chromatin structure dynamics. R-loop homeostasis is dynamically regulated and closely associated with gene transcription in mouse zygotes. However, the factors responsible for regulating these dynamic changes in the R-loops of fertilized mouse eggs have not yet been investigated. This study examined the functions of candidate factors that interact with R-loops during zygotic gene activation. Methods: In this study, we used publicly available next-generation sequencing datasets, including low-input ribosome profiling analysis and polymerase II chromatin immunoprecipitation-sequencing (ChIP-seq), to identify potential regulators of R-loop dynamics in zygotes. These datasets were downloaded, reanalyzed, and compared with mass spectrometry data to identify candidate factors involved in regulating R-loop dynamics. To validate the functions of these candidate factors, we treated mouse zygotes with chemical inhibitors using in vitro fertilization. Immunofluorescence with an anti-R-loop antibody was then performed to quantify changes in R-loop metabolism. Results: We identified DEAD-box-5 (DDX5) and histone deacetylase-2 (HDAC2) as candidates that potentially regulate R-loop metabolism in oocytes, zygotes and two-cell embryos based on change of their gene translation. Our analysis revealed that the DDX5 inhibition of activity led to decreased R-loop accumulation in pronuclei, indicating its involvement in regulating R-loop dynamics. However, the inhibition of histone deacetylase-2 activity did not significantly affect R-loop levels in pronuclei. Conclusion: These findings suggest that dynamic changes in R-loops during mouse zygote development are likely regulated by RNA helicases, particularly DDX5, in conjunction with transcriptional processes. Our study provides compelling evidence for the involvement of these factors in regulating R-loop dynamics during early embryonic development.

Keywords

Acknowledgement

The authors are indebted to all the members of the KH lab for helpful discussion.

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