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Arf tumor suppressor and miR-205 regulate cell adhesion and formation of extraembryonic endoderm from pluripotent stem cells.

ABSTRACT: Induction of the Arf tumor suppressor (encoded by the alternate reading frame of the Cdkn2a locus) following oncogene activation engages a p53-dependent transcriptional program that limits the expansion of incipient cancer cells. Although the p19(Arf) protein is not detected in most tissues of fetal or young adult mice, it is physiologically expressed in the fetal yolk sac, a tissue derived from the extraembryonic endoderm (ExEn). Expression of the mouse p19(Arf) protein marks late stages of ExEn differentiation in cultured embryoid bodies (EBs) derived from either embryonic stem cells or induced pluripotent stem cells. Arf inactivation delays differentiation of the ExEn lineage within EBs, but not the formation of other germ cell lineages from pluripotent progenitors. Arf is required for the timely induction of ExEn cells in response to Ras/Erk signaling and, in turn, acts through p53 to ensure the development, but not maintenance, of the ExEn lineage. Remarkably, a significant temporal delay in ExEn differentiation detected during the maturation of Arf-null EBs is rescued by enforced expression of mouse microRNA-205 (miR-205), a microRNA up-regulated by p19(Arf) and p53 that controls ExEn cell migration and adhesion. The noncanonical and canonical roles of Arf in ExEn development and tumor suppression, respectively, may be conceptually linked through mechanisms that govern cell attachment and migration.

SUBMITTER: Li C

PROVIDER: S-EPMC3607049 | biostudies-literature | 2013 Mar

REPOSITORIES: biostudies-literature

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Arf tumor suppressor and miR-205 regulate cell adhesion and formation of extraembryonic endoderm from pluripotent stem cells.

Li Chunliang C Finkelstein David D Sherr Charles J CJ

Proceedings of the National Academy of Sciences of the United States of America 20130304 12

Induction of the Arf tumor suppressor (encoded by the alternate reading frame of the Cdkn2a locus) following oncogene activation engages a p53-dependent transcriptional program that limits the expansion of incipient cancer cells. Although the p19(Arf) protein is not detected in most tissues of fetal or young adult mice, it is physiologically expressed in the fetal yolk sac, a tissue derived from the extraembryonic endoderm (ExEn). Expression of the mouse p19(Arf) protein marks late stages of ExE ...[more]

PMID: 23487795

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Project description:Induction of the Arf tumor suppressor in response to hyperproliferative stress following oncogene activation activates a p53-dependent transcriptional program that limits the expansion of incipient cancer cells. Although Arf is not expressed in most tissues of fetal or young adult mice, it is physiologically expressed in the fetal yolk sac, a tissue derived from the extraembryonic endoderm. We demonstrate that expression of the mouse p19Arf protein marks late stages of extraembryonic endoderm differentiation in cultured embryoid bodies derived from either embryonic stem cells or induced pluripotent stem cells, and that Arf inactivation specifically delays the differentiation of the extraembryonic endoderm lineage, but not the formation of other germ cell lineages from pluripotent progenitors. Arf is required for the timely induction of extraembryonic endodermal cells in response to Ras/Erk signaling and, in turn, acts through p53 to ensure extraembryonic endoderm lineage development, but not maintenance. Remarkably, a significant temporal delay in extraembryonic endoderm differentiation detected during the maturation of Arf-null embryoid bodies is rescued by enforced expression of miR-205, a micro-RNA up-regulated by p19Arf and p53. Introduction of miR-205 into Arf-null embryonic stem cells rescues defective ExEn formation and elicits a program of gene expression that controls the migration and adhesion of embryonic endodermal cells. This occurs, at least in part, through atypical regulation of genes that control the epithelial-to-mesenchymal transition in cancer cells. Our findings suggest that noncanonical and canonical roles of Arf in extraembryonic endoderm development and tumor suppression, respectively, may be conceptually linked through mechanisms that govern cell-to-cell attachment and migration. 4 samples total two each at two time points in ESC development At each time point one sample was treted with miR-205 and the other with a GFP control vector

Project description:BackgroundInitial specification of cardiomyocytes in the mouse results from interactions between the extraembryonic anterior visceral endoderm (AVE) and the nascent mesoderm. However the mechanism by which AVE activates cardiogenesis is not well understood, and the identity of specific cardiogenic factors in the endoderm remains elusive. Most mammalian studies of the cardiogenic potential of the endoderm have relied on the use of cell lines that are similar to the heart-inducing AVE. These include the embryonal-carcinoma-derived cell lines, END2 and PYS2. The recent development of protocols to isolate eXtraembryonic ENdoderm (XEN) stem cells, representing the extraembryonic endoderm lineage, from blastocyst stage mouse embryos offers new tools for the genetic dissection of cardiogenesis.Methodology/principal findingsHere, we demonstrate that XEN cell-conditioned media (CM) enhances cardiogenesis during Embryoid Body (EB) differentiation of mouse embryonic stem (ES) cells in a manner comparable to PYS2-CM and END2-CM. Addition of CM from each of these three cell lines enhanced the percentage of EBs that formed beating areas, but ultimately, only XEN-CM and PYS2-CM increased the total number of cardiomyocytes that formed. Furthermore, our observations revealed that both contact-independent and contact-dependent factors are required to mediate the full cardiogenic potential of the endoderm. Finally, we used gene array comparison to identify factors in these cell lines that could mediate their cardiogenic potential.Conclusions/significanceThese studies represent the first step in the use of XEN cells as a molecular genetic tool to study cardiomyocyte differentiation. Not only are XEN cells functionally similar to the heart-inducing AVE, but also can be used for the genetic dissection of the cardiogenic potential of AVE, since they can be isolated from both wild type and mutant blastocysts. These studies further demonstrate the importance of both contact-dependent and contact-independent factors in cardiogenesis and identify potential heart-inducing proteins in the endoderm.

Dataset Information

Arf tumor suppressor and miR-205 regulate cell adhesion and formation of extraembryonic endoderm from pluripotent stem cells.

Publications

Arf tumor suppressor and miR-205 regulate cell adhesion and formation of extraembryonic endoderm from pluripotent stem cells.

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