Journal
DEVELOPMENT
Volume 145, Issue 20, Pages -Publisher
COMPANY BIOLOGISTS LTD
DOI: 10.1242/dev.170209
Keywords
ChIP-seq; Embryonic stem cells; Oct4; Prdm14; Primordial germ cells; Targeted DamID
Categories
Funding
- Wellcome Trust [092096, 103792, 092545, 105839, 096738]
- Herchel Smith Fund Research Studentship
- European Molecular Biology Organization Long-term Fellowship [ALTF 263_2014, ALTF 1600_2014]
- Uehara Memorial Foundation Research Fellowship
- Kanae Foundation for the Promotion of Medical Science
- Human Frontier Science Program [RGP0020/2012]
- Cancer Research UK [C6946/A14492]
- Royal Society Darwin Trust Research Professorship
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The precise control of gene expression by transcription factor networks is crucial to organismal development. The predominant approach for mapping transcription factor-chromatin interactions has been chromatin immunoprecipitation (ChIP). However, ChIP requires a large number of homogeneous cells and antisera with high specificity. A second approach, DamID, has the drawback that high levels of Dam methylase are toxic. Here, we modify our targeted DamID approach (TaDa) to enable cell type-specific expression in mammalian systems, generating an inducible system (mammalian TaDa or MaTaDa) to identify genome-wide protein/DNA interactions in 100 to 1000 times fewer cells than ChIP-based approaches. We mapped the binding sites of two key pluripotency factors, OCT4 and PRDM14, in mouse embryonic stem cells, epiblast-like cells and primordial germ cell-like cells (PGCLCs). PGCLCs are an important system for elucidating primordial germ cell development in mice. We monitored PRDM14 binding during the specification of PGCLCs, identifying direct targets of PRDM14 that are key to understanding its crucial role in PGCLC development. We show that MaTaDa is a sensitive and accurate method for assessing cell type-specific transcription factor binding in limited numbers of cells.
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