Mapping Chromatin Occupancy of Ppp1r1b-lncRNA Genome-Wide Using Chromatin Isolation by RNA Purification (ChIRP)-seq

• Published in: Cells (Basel, Switzerland) Vol. 12; no. 24; p. 2805
• Main Authors: Hwang, John, Kang, Xuedong, Wolf, Charlotte, Touma, Marlin
• Format: Journal Article
• Language: English
• Published: Switzerland MDPI AG 08.12.2023
• Subjects: Animals; Binding sites; Cardiac muscle; Chromatin; Chromatin - genetics; chromatin isolation by RNA purification; chromatin occupancy; DNA-Binding Proteins - metabolism; Enhancers; Epigenetics; Fetuses; Gene Expression Regulation; Gene mapping; Gene regulation; Genes; Genomes; Homeobox; Humans; Introns; long non-coding RNA; Mice; Muscles; Myoblasts; Myocytes; Next-generation sequencing; Non-coding RNA; Polycomb group proteins; Polycomb Repressive Complex 2 - metabolism; Ppp1r1b-lncRNA; Purification; RNA polymerase; RNA, Long Noncoding - genetics; RNA, Long Noncoding - metabolism; RNA-protein interactions; Software; Transcription factors; Transcription initiation; chromatin occupancy; long non-coding RNA; chromatin isolation by RNA purification; Ppp1r1b-lncRNA; epigenetics
• ISSN: 2073-4409; 2073-4409
• DOI: 10.3390/cells12242805

Long non-coding RNA (lncRNA) mediated transcriptional regulation is increasingly recognized as an important gene regulatory mechanism during development and disease. LncRNAs are emerging as critical regulators of chromatin state; yet the nature and the extent of their interactions with chromatin remain to be fully revealed. We have previously identified as an essential epigenetic regulator of myogenic differentiation in cardiac and skeletal myocytes in mice and humans. We further demonstrated that function is mediated by the interaction with the chromatin-modifying complex polycomb repressive complex 2 (PRC2) at the promoter of myogenic differentiation transcription factors, and . Herein, we employed unbiased chromatin isolation by RNA purification (ChIRP) and high throughput sequencing to map the repertoire of chromatin occupancy genome-wide in the mouse muscle myoblast cell line. We uncovered a total of 99732 true peaks corresponding to binding sites at high confidence ( -value < 1E-5) and enrichment score ≥ 10). The -binding sites averaged 558 bp in length and were distributed widely within the coding and non-coding regions of the genome. Approximately 46% of these true peaks were mapped to gene elements, of which 1180 were mapped to experimentally validated promoter sequences. Importantly, the promoter-mapped binding sites were enriched in myogenic transcription factors and heart development while exhibiting focal interactions with known motifs of proximal promoters and transcription initiation by RNA Pol-II, including TATA-box, transcription initiator motif, CCAAT-box, and GC-box, supporting role in transcription initiation of myogenic regulators. Remarkably, nearly 40% of -binding sites mapped to gene introns were enriched with the Homeobox family of transcription factors and exhibited TA-rich motif sequences, suggesting potential motif-specific -bound introns. Lastly, more than 136521 enhancer sequences were detected in -occupancy sites at high confidence. Among these enhancers, 3390 (12%) exhibited cell type/tissue-specific enrichment in fetal heart and muscles. Together, our findings provide further insights into the genome-wide Chromatin interactome that may dictate its function in myogenic differentiation and potentially other cellular and biological processes.