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The study of nascent RNA transcripts in asynchronous cells and cells released from mitosis at the indicated timepoints between Smarce1- MD (R42A) and Smarce1- MD mESCs.

ABSTRACT: The study of nascent RNA transcripts in asynchronous cells and cells released from mitosis at the indicated timepoints between Smarce1- MD (R42A) and Smarce1- MD mESCs.

PROVIDER: PRJNA783564 | ENA |

REPOSITORIES: ENA

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Project description:During mitosis, RNA polymerase and most transcription factors are excluded from the chromosomes and transcription ceases. The transcriptional re-activation of the genome, following mitosis, requires the re-setting of cell-type specific programs that were initially established during development. However, only about one-fifth of transcription factors are retained on chromosomes throughout mitosis and a subset of these have been shown to facilitate target gene reactivation during mitotic exit. How such “bookmarking” factors bind to chromatin in mitosis and re-activate transcription is central to the stability of transcriptional programs across multiple cell cycles. We compared a diverse set of transcription factors involved in liver differentiation and found different modes of mitotic chromosome binding. The pioneer transcription factor FoxA1, which is among the first to bind liver genes in development, exhibits virtually complete mitotic chromosome binding, whereas other liver factors bind with a range of efficiencies. Yet genome-wide analysis shows that only about 15% of the FoxA1 interphase target sites are bound in mitosis; the latter include sites at genes for maintaining cell differentiation. FoxA1 mutants that perturb specific and nonspecific DNA binding reveal a significant contribution of nonspecific binding events in mitotic chromatin. Such nonspecific binding appears to spread from interphase FoxA1 targets and may serve as storage sites. The hierarchy of specific binding, nonspecific binding, partial chromatin binding, and failure to bind mitotic chromosomes reflects the temporal sequence of the factors’ developmental roles in gene activation. Gene expression was assessed in asynchronously cycling cells in order to determine whether FoxA1 is disposed to bind to high- or low-expressed genes during mitosis. Heatmap analysis shows that FoxA1's mitotic targets are among the highest-expressed genes in asynchronous cells. Total RNA was collected from three different plates with asynchronous HUH7 cells using the RNeasy mini kit (Qiagen Valencia CA). Expression microarrays were performed with a Human Gene 1.OST array (Affymetrix) at the UPenn Microarray Core Facility and assessed using Partek.

Dataset Information

The study of nascent RNA transcripts in asynchronous cells and cells released from mitosis at the indicated timepoints between Smarce1- MD (R42A) and Smarce1- MD mESCs.

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