Automated CUT&Tag profiling of chromatin heterogeneity in mixed-lineage leukemia

Abstract

Acute myeloid and lymphoid leukemias often harbor chromosomal translocations involving the KMT2A gene, encoding the KMT2A lysine methyltransferase (also known as mixed-lineage leukemia-1), and produce in-frame fusions of KMT2A to other chromatin-regulatory proteins. Here we map fusion-specific targets across the genome for diverse KMT2A oncofusion proteins in cell lines and patient samples. By modifying CUT&Tag chromatin profiling for full automation, we identify common and tumor-subtype-specific sites of aberrant chromatin regulation induced by KMT2A oncofusion proteins. A subset of KMT2A oncofusion-binding sites are marked by bivalent (H3K4me3 and H3K27me3) chromatin signatures, and single-cell CUT&Tag profiling reveals that these sites display cell-to-cell heterogeneity suggestive of lineage plasticity. In addition, we find that aberrant enrichment of H3K4me3 in gene bodies is sensitive to Menin inhibitors, demonstrating the utility of automated chromatin profiling for identifying therapeutic vulnerabilities. Thus, integration of automated and single-cell CUT&Tag can uncover epigenomic heterogeneity within patient samples and predict sensitivity to therapeutic agents.

Glutamatergic and GABAergic neurons in the preoptic area of the hypothalamus play key roles in menopausal hot flashes

“…The CUT&Tag Assay Kit used in the experiment was purchased from Vazyme Biotech (Beijing) Co., Ltd. The experimental procedures, including the buffer preparation, ConA bead treatment, nuclear incubation with ConA beads, incubation with primary antibodies (mouse anti-ERα primary antibody, NB300-560, Novus, USA; mouse anti-ERβ primary antibody, NB200-305, Novus, USA), pA/G-Tnp incubation, fragmentation, DNA extraction, library amplification and purification of PCR products, were carried out according to the instructions and previous studies (Bartosovic et al, 2021;Janssens et al, 2021). The obtained library was submitted to Annoroad (Beijing) Co., Ltd., to be sequenced, and the bioinformatics analysis was carried out using the CUT_Tag_tool on the Vazymeyun platform.…”

Genomic Occupancy of the Bromodomain Protein Bdf3 Is Dynamic during Differentiation of African Trypanosomes from Bloodstream to Procyclic Forms

“…The originators of the protocol in mammalian cells claim that high-quality data can be obtained using 100 to 1,000 cells ( 43 ). The development of CUT&TAG (cleavage under targets and TAGmentation) has streamlined the process even further by eliminating some downstream steps associated with sequencing library preparation ( 43 ) and has further been refined to be fully automated ( 44 ). CUT&TAG has also been adapted for single-cell chromatin studies ( 45 ), which represents an exciting future application for the technique in T. brucei .…”

Profiling RNA at chromatin targets in situ by antibody-targeted tagmentation

“…This allows for seamless generation of Illumina sequencing libraries using a simple PCR reaction, without the need to purify RNA as in ChRIP-seq, APEX-seq and TRIBE. The lack of purification steps makes RT&Tag adaptable for automation as was done with AutoCUT&Tag 32 . Together with low cell number input, low sequencing depth, RT&Tag presents a high throughput method to study RNA metabolism by targeting chromatin factors and post-translational modifications.…”