Epigenomic Profiling of B Cell Subsets by CUT&Tag.

Epigenetic programs play a key role in regulating the development and function of immune cells. However, conventional methods for profiling epigenetic mechanisms, such as the post-translational modifications to histones, present several technical challenges that prevent a complete understanding of gene regulation. Here, we provide a detailed protocol of the Cleavage Under Targets and Tagmentation (CUT&Tag) chromatin profiling technique for identifying histone modifications in human and mouse lymphocytes.

Distinct transcriptomic and epigenomic modalities underpin human memory T cell subsets and their activation potential.

Human memory T cells (MTC) are poised to rapidly respond to antigen re-exposure. Here, we derived the transcriptional and epigenetic programs of resting and ex vivo activated, circulating CD4+ and CD8+ MTC subsets. A progressive gradient of gene expression from naïve to TCM to TEM is observed, which is accompanied by corresponding changes in chromatin accessibility. Transcriptional changes suggest adaptations of metabolism that are reflected in altered metabolic capacity. Other differences involve regulatory modalities comprised of discrete accessible chromatin patterns, transcription factor binding motif enrichment, and evidence of epigenetic priming. Basic-helix-loop-helix factor motifs for AHR and HIF1A distinguish subsets and predict transcription networks to sense environmental changes. Following stimulation, primed accessible chromatin correlate with an augmentation of MTC gene expression as well as effector transcription factor gene expression. These results identify coordinated epigenetic remodeling, metabolic, and transcriptional changes that enable MTC subsets to ultimately respond to antigen re-encounters more efficiently.

Identification of chromatin states during zebrafish gastrulation using CUT&RUN and CUT&Tag.

BACKGROUND: Cell fate decisions are governed by interactions between sequence-specific transcription factors and a dynamic chromatin landscape. Zebrafish offer a powerful system for probing the mechanisms that drive these cell fate choices, especially in the context of early embryogenesis. However, technical challenges associated with conventional methods for chromatin profiling have slowed progress toward understanding the exact relationships between chromatin changes, transcription factor binding, and cellular differentiation during zebrafish embryogenesis. RESULTS: To overcome these challenges, we adapted the chromatin profiling methods Cleavage Under Targets and Release Using Nuclease (CUT&RUN) and CUT&Tag for use in zebrafish and applied these methods to generate high-resolution enrichment maps for H3K4me3, H3K27me3, H3K9me3, RNA polymerase II, and the histone variant H2A.Z using tissue isolated from whole, mid-gastrula stage embryos. Using this data, we identify a subset of genes that may be bivalently regulated during both zebrafish and mouse gastrulation, provide evidence for an evolving H2A.Z landscape during embryo development, and demonstrate the effectiveness of CUT&RUN for detecting H3K9me3 enrichment at repetitive sequences. CONCLUSIONS: Our results demonstrate the power of combining CUT&RUN and CUT&Tag methods with the strengths of the zebrafish system to define emerging chromatin landscapes in the context of vertebrate embryogenesis.

Chromatin dynamics at the maternal to zygotic transition: recent advances from the zebrafish model.

Early animal development is characterized by intense reorganization of the embryonic genome, including large-scale changes in chromatin structure and in the DNA and histone modifications that help shape this structure. Particularly profound shifts in the chromatin landscape are associated with the maternal-to-zygotic transition, when the zygotic genome is first transcribed and maternally loaded transcripts are degraded. The accessibility of the early zebrafish embryo facilitates the interrogation of chromatin during this critical window of development, making it an important model for early chromatin regulation. Here, we review our current understanding of chromatin dynamics during early zebrafish development, highlighting new advances as well as similarities and differences between early chromatin regulation in zebrafish and other species.