Abstract
At zygotic genome activation (ZGA), changes in chromatin structure are associated with new transcription immediately following the maternal-to-zygotic transition (MZT). The nuclear architectural proteins, cohesin and CCCTC-binding factor (CTCF), contribute to chromatin structure and gene regulation. We show here that normal cohesin function is important for ZGA in zebrafish. Depletion of cohesin subunit Rad21 delays ZGA without affecting cell cycle progression. In contrast, CTCF depletion has little effect on ZGA whereas complete abrogation is lethal. Genome wide analysis of Rad21 binding reveals a change in distribution from pericentromeric satellite DNA, and few locations including the miR-430 locus (whose products are responsible for maternal transcript degradation), to genes, as embryos progress through the MZT. After MZT, a subset of Rad21 binding occurs at genes dysregulated upon Rad21 depletion and overlaps pioneer factor Pou5f3, which activates early expressed genes. Rad21 depletion disrupts the formation of nucleoli and RNA polymerase II foci, suggestive of global defects in chromosome architecture. We propose that Rad21/cohesin redistribution to active areas of the genome is key to the establishment of chromosome organization and the embryonic developmental program.
Author Summary During the first few hours of existence, early zygotic cellular events are regulated by maternally inherited molecules. From a defined timepoint, the zygotic genome gradually becomes active and is transcribed. How the zygotic genome is first held inactive before becoming rapidly activated is poorly understood. Both gene repression and activation mechanisms are involved, but one aspect that has not yet been investigated is how 3-dimensional chromosome structure influences genome activation. In this study, we used zebrafish embryos to model zygotic genome activation.
The multi-subunit protein complex, cohesin, and the DNA-binding protein CCCTC-binding factor (CTCF) both have well known and overlapping roles in 3-dimensional genome organization. We depleted cohesin subunit Rad21, or CTCF, to determine their effects on zygotic genome activation. Moderate Rad21 depletion delayed transition to zygotic gene expression, without disrupting the cell cycle. By contrast, moderate CTCF depletion had very little effect; however, strong depletion of CTCF was lethal. We surveyed genome-wide binding of Rad21 before and after the zygotic genome is activated, and determined what other chromatin factors and transcription factors coincide with Rad21 binding. Before genome activation, Rad21 was located at satellite DNA and a few noncoding genes, one of which (miR-430) is responsible for degrading maternal transcripts. Following genome activation, there was a mass relocation of Rad21 to genes, particularly active genes and those that are targets of transcriptional activators when the zygotic genome is switched on. Depletion of Rad21 also affected global chromosome structure.
Our study shows that cohesin binding redistributes to active RNA Polymerase II genes at the onset of zygotic gene transcription. Furthermore, we suggest that cohesin contributes to dynamic changes in chromosome architecture that occur upon zygotic genome activation.
