Biotinylated Tn5 transposase‐mediated CUT&Tag efficiently profiles transcription factor‐DNA interactions in plants

Abstract

Summary In contrast to CUT&Tag approaches for profiling bulk histone modifications, current CUT&Tag methods for analysing specific transcription factor (TF)‐DNA interactions remain technically challenging due to TFs having relatively low abundance. Moreover, an efficient CUT&Tag strategy for plant TFs is not yet available. Here, we first applied biotinylated Tn5 transposase‐mediated CUT&Tag (B‐CUT&Tag) to produce high‐quality libraries for interrogating TF‐DNA interactions. B‐CUT&Tag combines streptavidin‐biotin‐based DNA purification with routine CUT&Tag, optimizing the removal of large amounts of intact chromatin not targeted by specific TFs. The biotinylated chromatin fragments are then purified for construction of deep sequencing libraries or qPCR analysis. We applied B‐CUT&Tag to probe genome‐wide DNA targets of Squamosa promoter‐binding‐like protein 9 (SPL9), a well‐established TF in Arabidopsis; the resulting profiles were efficient and consistent in demonstrating its well‐established target genes in juvenile‐adult transition/flowering, trichome development, flavonoid biosynthesis, wax synthesis and branching. Interestingly, our results indicate functions of AtSPL9 in modulating growth‐defence trade‐offs. In addition, we established a method for applying qPCR after CUT&Tag (B‐CUT&Tag‐qPCR) and successfully validated the binding of SPL9 in Arabidopsis and PHR2 in rice. Our study thus provides a convenient and highly efficient CUT&Tag strategy for profiling TF‐chromatin interactions that is widely applicable to the annotation of cis‐regulatory elements for crop improvement.

CUT&Tag and DiBioCUT&Tag enable investigation of the AT-rich and dynamic epigenome ofPlasmodium falciparumfrom low input samples

“…histones) in the vicinity of the binding site are biotinylated and later profiled by anti-biotin CUT&Tag (DiBioCUT&Tag). CUT&Tag of the biotinylated chromatin enables the use of a standard α-biotin antibody [35], but requires genetic modification of the target proteins. Furthermore, direct coupling of the target protein and the biotin ligase can lead to background biotinylation and could prevent temporal resolution of dynamic binding patterns.…”

Krüppel-like factor 12 decreases progestin sensitivity in endometrial cancer by inhibiting the progesterone receptor signaling pathway

“…CUT&Tag was performed according to the published procedure [ 20 ] with modifications using the Hyperactive Universal CUT&Tag Assay Kit for Illumina (Vazyme). In brief, 6×10 6 Ishikawa cells in the logarithmic growth phase were collected and washed with 500 μL wash buffer and centrifuged at 2500 rpm for 5 min at room temperature.…”

Post-Translational Modifications in Histones and Their Role in Abiotic Stress Tolerance in Plants

“…CUT & Tag is designed to target specific genomic regions or proteins, including histones with specific modifications. For instance, researchers can use antibodies against histone modifications of interest (e.g., H3K4me3 for active promoters) to selectively profile these modifications in response to abiotic stress [141]. It can also generates high-resolution maps of histone modifications, allowing researchers to pinpoint precisely where these modifications occur in the genome.…”