Development of Site-Specific ChIP Technologies (Hodaka Fujii)

In this episode of the Epigenetics Podcast, we caught up with Dr. Hodaka Fujii, Professor of Biochemistry and Genome Biology at Hirosaki University Graduate School of Medicine and School of Medicine, to talk about his work on the development of locus-specific ChIP technologies.

The goal of conventional chromatin immunoprecipitation (ChIP) assays is to find genomic locations of transcription factor binding or genome-wide profiles of histone tail modifications.  In contrast to that, the guest of this episode, Dr. Fujii, has developed methods such as insertional chromatin immunoprecipitation (iChIP) and engineered DNA-binding molecule-mediated chromatin immunoprecipitation (enChIP) to identify the factors that are binding to specific sites on the genome.

In iChIP, LexA binding sites are inserted into the genomic region of interest. In parallel, the DNA-binding domain of LexA, fused with FLAG epitope tags and a nuclear localization signal, is expressed in the same cells. After crosslinking and chromatin preparation, the resulting chromatin is immunoprecipitated with an antibody against the tag. This allows proteins or RNA interacting with the region of interest to be analyzed with the appropriate downstream application. The enChIP takes a similar approach, but does not require insertion of the LexA binding sites. Instead, a FLAG-tagged dCas9 protein together with the respective guide RNA are used to target the region of the genome of interest. After the IP and the purification DNA, RNA, or proteins can be analyzed accordingly. The lack of the requirement of to insert the LexA binding sites into the genome makes enChIP much more straightforward than iChIP.

In this interview, we discuss the story behind how Dr. Fujii got into the field of epigenetics, how he developed iChIP, and how the method was improved over the years. Furthermore, we discuss the development of enChIP and how this can be used as an alternate method to Hi-C.

 

References

 

• Akemi Hoshino, Satoko Matsumura, … Hodaka Fujii (2004) Inducible Translocation Trap (Molecular Cell) DOI: 10.1016/j.molcel.2004.06.017
• Akemi Hoshino, Hodaka Fujii (2009) Insertional chromatin immunoprecipitation: a method for isolating specific genomic regions (Journal of Bioscience and Bioengineering) DOI: 10.1016/j.jbiosc.2009.05.005
• Toshitsugu Fujita, Hodaka Fujii (2013) Efficient isolation of specific genomic regions and identification of associated proteins by engineered DNA-binding molecule-mediated chromatin immunoprecipitation (enChIP) using CRISPR (Biochemical and Biophysical Research Communications) DOI: 10.1016/j.bbrc.2013.08.013
• Toshitsugu Fujita, Miyuki Yuno, … Hodaka Fujii (2015) Identification of Non-Coding RNAs Associated with Telomeres Using a Combination of enChIP and RNA Sequencing (PLOS ONE) DOI: 10.1371/journal.pone.0123387
• Toshitsugu Fujita, Miyuki Yuno, Hodaka Fujii (2016) Efficient sequence-specific isolation of DNA fragments and chromatin by in vitro enChIP technology using recombinant CRISPR ribonucleoproteins (Genes to Cells) DOI: 10.1111/gtc.12341
• Toshitsugu Fujita, Miyuki Yuno, … Hodaka Fujii (2017) Identification of physical interactions between genomic regions by enChIP-Seq (Genes to Cells) DOI: 10.1111/gtc.12492
• Toshitsugu Fujita, Fusako Kitaura, … Hodaka Fujii (2017) Locus-specific ChIP combined with NGS analysis reveals genomic regulatory regions that physically interact with the Pax5 promoter in a chicken B cell line (DNA Research) DOI: 10.1093/dnares/dsx023

 

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