Optimized CUT&Tag enables robust epigenome profiling in Schizosaccharomyces pombe

Area of Science:

• Epigenetics
• Molecular Biology
• Microbiology

Background:

• Chromatin immunoprecipitation sequencing (ChIP-seq) is limited by crosslinking artifacts and high cell input requirements.
• Schizosaccharomyces pombe (S. pombe) epigenome profiling requires optimized methodologies for high-resolution analysis.

Purpose of the Study:

• To optimize permeabilization for CUT&Tag in S. pombe for robust H3K9me3 profiling.
• To establish a high-resolution epigenome profiling protocol with reduced cell input.

Main Methods:

• Optimized permeabilization using Lywallzyme for S. pombe spheroplast generation (>95% efficiency).
• Systematic parameter optimization of CUT&Tag, including enzyme concentration and cell load.
• H3K9me3 mapping in wild-type and clr4Δ strains using the optimized protocol.

Main Results:

• Lywallzyme outperformed other enzymes for spheroplast generation.
• Identified optimal digestion kinetics and an inverse cell load-efficiency relationship.
• Captured specific heterochromatic enrichment at centromeres/telomeres with high signal-to-noise ratio.

Conclusions:

• The optimized CUT&Tag protocol enables robust epigenomic analysis in S. pombe with minimal cell input.
• This method overcomes limitations of ChIP-seq, offering enhanced resolution for epigenomic studies.