Geminin Is Required for Zygotic Gene Expression at the Xenopus Mid-Blastula Transition

Sarah L Kerns,Tina M Thorne,Thomas J Mcgarry,Kathryn M Schultz,Kelly A Barry,Michael Klymkowsky

doi:10.1371/journal.pone.0038009

Abstract

In many organisms early development is under control of the maternal genome and zygotic gene expression is delayed until the mid-blastula transition (MBT). As zygotic transcription initiates, cell cycle checkpoints become activated and the tempo of cell division slows. The mechanisms that activate zygotic transcription at the MBT are incompletely understood, but they are of interest because they may resemble mechanisms that cause stem cells to stop dividing and terminally differentiate. The unstable regulatory protein Geminin is thought to coordinate cell division with cell differentiation. Geminin is a bi-functional protein. It prevents a second round of DNA replication during S and G2 phase by binding and inhibiting the essential replication factor Cdt1. Geminin also binds and inhibits a number of transcription factors and chromatin remodeling proteins and is thought to keep dividing cells in an undifferentiated state. We previously found that the cells of Geminin-deficient Xenopus embryos arrest in G2 phase just after the MBT then disintegrate at the onset of gastrulation. Here we report that they also fail to express most zygotic genes. The gene expression defect is cell-autonomous and is reproduced by over-expressing Cdt1 or by incubating the embryos in hydroxyurea. Geminin deficient and hydroxyurea-treated blastomeres accumulate DNA damage in the form of double stranded breaks. Bypassing the Chk1 pathway overcomes the cell cycle arrest caused by Geminin depletion but does not restore zygotic gene expression. In fact, bypassing the Chk1 pathway by itself induces double stranded breaks and abolishes zygotic transcription. We did not find evidence that Geminin has a replication-independent effect on transcription. We conclude that Geminin is required to maintain genome integrity during the rapid cleavage divisions, and that DNA damage disrupts zygotic gene transcription at the MBT, probably through activation of DNA damage checkpoint pathways.

Highlights

In many metazoans embryonic development begins with a series of extremely rapid cleavage divisions that quickly produce a blastula containing thousands of cells
Zygotic transcription is deferred until the mid-blastula stage, at a point called the mid-blastula transition (MBT) in Xenopus or the maternal-zygotic transition (MZT) in Drosophila [1]
We previously showed that Xenopus embryos lacking Geminin arrest in G2 phase just after the 13th cell division, one cell cycle after the MBT is reached [28]

Summary

Introduction

In many metazoans embryonic development begins with a series of extremely rapid cleavage divisions that quickly produce a blastula containing thousands of cells During this period development is under the control of maternal RNAs stored in the egg. Zygotic transcription is deferred until the mid-blastula stage, at a point called the mid-blastula transition (MBT) in Xenopus or the maternal-zygotic transition (MZT) in Drosophila [1]. Concomitant with activation of the zygotic genome, the cell cycle slows as gap phases are introduced between S and M phases This pattern of development is thought to be an adaptation that rapidly provides enough cells to form a feeding larva, an important consideration for organisms with eggs that develop outside the mother’s body. The mechanisms that switch on zygotic transcription at the MBT are incompletely understood

Methods

Results

Conclusion