RRC ID 28087
Author Sakaue-Sawano A, Hoshida T, Yo M, Takahashi R, Ohtawa K, Arai T, Takahashi E, Noda S, Miyoshi H, Miyawaki A.
Title Visualizing developmentally programmed endoreplication in mammals using ubiquitin oscillators.
Journal Development
Abstract The majority of mammalian somatic cells maintain a diploid genome. However, some mammalian cell types undergo multiple rounds of genome replication (endoreplication) as part of normal development and differentiation. For example, trophoblast giant cells (TGCs) in the placenta become polyploid through endoreduplication (bypassed mitosis), and megakaryocytes (MKCs) in the bone marrow become polyploid through endomitosis (abortive mitosis). During the normal mitotic cell cycle, geminin and Cdt1 are involved in 'licensing' of replication origins, which ensures that replication occurs only once in a cell cycle. Their protein accumulation is directly regulated by two E3 ubiquitin ligase activities, APC(Cdh1) and SCF(Skp2), which oscillate reciprocally during the cell cycle. Although proteolysis-mediated, oscillatory accumulation of proteins has been documented in endoreplicating Drosophila cells, it is not known whether the ubiquitin oscillators that control normal cell cycle transitions also function during mammalian endoreplication. In this study, we used transgenic mice expressing Fucci fluorescent cell-cycle probes that report the activity of APC(Cdh1) and SCF(Skp2). By performing long-term, high temporal-resolution Fucci imaging, we were able to visualize reciprocal activation of APC(Cdh1) and SCF(Skp2) in differentiating TGCs and MKCs grown in our custom-designed culture wells. We found that TGCs and MKCs both skip cytokinesis, but in different ways, and that the reciprocal activation of the ubiquitin oscillators in MKCs varies with the polyploidy level. We also obtained three-dimensional reconstructions of highly polyploid TGCs in whole, fixed mouse placentas. Thus, the Fucci technique is able to reveal the spatiotemporal regulation of the endoreplicative cell cycle during differentiation.
Volume 140(22)
Pages 4624-32
Published 2013-11-1
DOI 10.1242/dev.099226
PII dev.099226
PMID 24154524
MeSH Animals Cell Survival Cells, Cultured Endoreduplication* Female Mammals / embryology* Megakaryocytes / cytology Megakaryocytes / metabolism Mice Mice, Transgenic Mitosis Molecular Imaging Placenta / cytology Placenta / metabolism Pregnancy Reproducibility of Results Trophoblasts / cytology Trophoblasts / metabolism Ubiquitin / metabolism*
IF 5.611
Times Cited 21
Mice RBRC02706