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NextPBM - A platform to study cell-specific transcription factor binding and cooperativity

ABSTRACT: This SuperSeries is composed of the SubSeries listed below.

ORGANISM(S): Homo sapiens

PROVIDER: GSE123947 | GEO | 2018/12/18

REPOSITORIES: GEO

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NextPBM: a platform to study cell-specific transcription factor binding and cooperativity.

Project description:High-throughput (HT) in vitro methods for measuring protein-DNA binding have become invaluable for characterizing transcription factor (TF) complexes and modeling gene regulation. However, current methods do not utilize endogenous proteins and, therefore, do not quantify the impact of cell-specific post-translational modifications (PTMs) and cooperative cofactors. We introduce the HT nextPBM (nuclear extract protein-binding microarray) approach to study DNA binding of native cellular TFs that accounts for PTMs and cell-specific cofactors. We integrate immune-depletion and phosphatase treatment steps into our nextPBM pipeline to characterize the impact of cofactors and phosphorylation on TF binding. We analyze binding of PU.1/SPI1 and IRF8 from human monocytes, delineate DNA-sequence determinants for their cooperativity, and show how PU.1 affinity correlates with enhancer status and the presence of cooperative and collaborative cofactors. We describe how nextPBMs, and our accompanying computational framework, can be used to discover cell-specific cofactors, screen for synthetic cooperative DNA elements, and characterize TF cooperativity.

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NextPBM - A platform to study cell-specific transcription factor binding and cooperativity [array]

Project description:High-throughput (HT) in vitro methods for measuring protein-DNA binding have become invaluable for characterizing transcription factor (TF) complexes and modeling gene regulation. However, current methods do not utilize endogenous proteins and, therefore, do not quantify the impact of cell-specific post-translational modifications (PTMs) and cooperative cofactors. We introduce the HT nextPBM (nuclear extract protein-binding microarray) approach to study DNA binding of native cellular TFs that accounts for PTMs and cell-specific cofactors. We integrate immune-depletion and phosphatase-treatment steps into our nextPBM pipeline to characterize the impact of cofactors and phosphorylation on TF binding. We analyze binding of PU.1/SPI1 and IRF8 from human monocytes, delineate DNA-sequence determinants for their cooperativity, and show how PU.1 affinity correlates with enhancer status and the presence of cooperative and collaborative cofactors. We describe how nextPBMs, and our accompanying computational framework, can be used to discover cell-specific cofactors, screen for synthetic cooperative DNA elements, and characterize TF cooperativity at single-nucleotide resolution.

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NextPBM - A platform to study cell-specific transcription factor binding and cooperativity [ChIP-Seq]

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