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1.
Development ; 151(11)2024 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-38752427

RESUMO

Bone morphogenic protein (BMP) signaling plays an essential and highly conserved role in embryo axial patterning in animal species. However, in mammalian embryos, which develop inside the mother, early development includes a preimplantation stage, which does not occur in externally developing embryos. During preimplantation, the epiblast is segregated from extra-embryonic lineages that enable implantation and development in utero. Yet, the requirement for BMP signaling is imprecisely defined in mouse early embryos. Here, we show that, in contrast to previous reports, BMP signaling (SMAD1/5/9 phosphorylation) is not detectable until implantation when it is detected in the primitive endoderm - an extra-embryonic lineage. Moreover, preimplantation development appears to be normal following deletion of maternal and zygotic Smad4, an essential effector of canonical BMP signaling. In fact, mice lacking maternal Smad4 are viable. Finally, we uncover a new requirement for zygotic Smad4 in epiblast scaling and cavitation immediately after implantation, via a mechanism involving FGFR/ERK attenuation. Altogether, our results demonstrate no role for BMP4/SMAD4 in the first lineage decisions during mouse development. Rather, multi-pathway signaling among embryonic and extra-embryonic cell types drives epiblast morphogenesis postimplantation.


Assuntos
Implantação do Embrião , Camadas Germinativas , Morfogênese , Transdução de Sinais , Proteína Smad4 , Animais , Proteína Smad4/metabolismo , Proteína Smad4/genética , Camadas Germinativas/metabolismo , Implantação do Embrião/genética , Camundongos , Morfogênese/genética , Feminino , Proteína Morfogenética Óssea 4/metabolismo , Proteína Morfogenética Óssea 4/genética , Regulação da Expressão Gênica no Desenvolvimento , Desenvolvimento Embrionário/genética , Camundongos Knockout , Embrião de Mamíferos/metabolismo , Endoderma/metabolismo , Endoderma/embriologia , Blastocisto/metabolismo , Blastocisto/citologia
2.
bioRxiv ; 2024 Jan 24.
Artigo em Inglês | MEDLINE | ID: mdl-38328075

RESUMO

Bone Morphogenic Protein (BMP) signaling plays an essential and highly conserved role in axial patterning in embryos of many externally developing animal species. However, in mammalian embryos, which develop inside the mother, early development includes an additional stage known as preimplantation. During preimplantation, the epiblast lineage is segregated from the extraembryonic lineages that enable implantation and development in utero. Yet, the requirement for BMP signaling in mouse preimplantation is imprecisely defined. We show that, in contrast to prior reports, BMP signaling (as reported by SMAD1/5/9 phosphorylation) is not detectable until implantation, when it is detected in the primitive endoderm - an extraembryonic lineage. Moreover, preimplantation development appears normal following deletion of maternal and zygotic Smad4, an essential effector of BMP signaling. In fact, mice lacking maternal Smad4 are viable. Finally, we uncover a new requirement for zygotic Smad4 in epiblast scaling and cavitation immediately after implantation, via a mechanism involving FGFR/ERK attenuation. Altogether, our results demonstrate no role for BMP4/SMAD4 in the first lineage decisions during mouse development. Rather, multi-pathway signaling among embryonic and extraembryonic cell types drives epiblast morphogenesis post-implantation.

3.
bioRxiv ; 2024 Jan 22.
Artigo em Inglês | MEDLINE | ID: mdl-38328220

RESUMO

During development, progenitors of embryonic stem (ES) and extraembryonic endoderm stem (XEN) cells are concomitantly specified within the inner cell mass (ICM) of the mouse blastocyst. Similarly, XEN cells are induced (iXEN cells) alongside induced pluripotent stem (iPS) cells following overexpression of Oct4, Sox2, Klf4 and Myc (OSKM) during somatic cell reprogramming. It is unclear how or why this cocktail produces both stem cell types, but OCT4 has been associated with non-pluripotent outcomes. In this report, we show that, during OSKM reprogramming, many individual Oct4-GFP-expressing cells are fated to become iXEN cells. Interestingly, SKM alone was also sufficient to induce iXEN cell formation, likely via activation of endogenous Oct4. These observations indicate that iXEN cell formation is not strictly an artifact of Oct4 overexpression. Moreover, our results suggest that a pathway to XEN may be an integral feature of establishing pluripotency during reprogramming, as in early embryo development.

4.
Science ; 382(6676): eadi5516, 2023 12 15.
Artigo em Inglês | MEDLINE | ID: mdl-38096290

RESUMO

Pioneer transcription factors (TFs), such as OCT4 and SOX2, play crucial roles in pluripotency regulation. However, the master TF-governed pluripotency regulatory circuitry was largely inferred from cultured cells. In this work, we investigated SOX2 binding from embryonic day 3.5 (E3.5) to E7.5 in the mouse. In E3.5 inner cell mass (ICM), SOX2 regulates the ICM-trophectoderm program but is dispensable for opening global enhancers. Instead, SOX2 occupies preaccessible enhancers in part opened by early-stage expressing TFs TFAP2C and NR5A2. SOX2 then widely redistributes when cells adopt naive and formative pluripotency by opening enhancers or poising them for rapid future activation. Hence, multifaceted pioneer TF-enhancer interaction underpins pluripotency progression in embryos, including a distinctive state in E3.5 ICM that bridges totipotency and pluripotency.


Assuntos
Blastocisto , Linhagem da Célula , Cromatina , Elementos Facilitadores Genéticos , Regulação da Expressão Gênica no Desenvolvimento , Fatores de Transcrição SOXB1 , Animais , Camundongos , Blastocisto/citologia , Blastocisto/metabolismo , Células Cultivadas , Cromatina/metabolismo , Fator 3 de Transcrição de Octâmero/genética , Fator 3 de Transcrição de Octâmero/metabolismo , Fatores de Transcrição SOXB1/genética , Fatores de Transcrição SOXB1/metabolismo , Diferenciação Celular/genética , Linhagem da Célula/genética
5.
Cell Reprogram ; 24(6): 353-362, 2022 12.
Artigo em Inglês | MEDLINE | ID: mdl-36342671

RESUMO

Somatic cell reprogramming was first developed to create induced pluripotent stem (iPS) cells. Since that time, the highly dynamic and heterogeneous nature of the reprogramming process has come to be appreciated. Remarkably, a distinct type of stem cell, called induced extraembryonic endoderm (iXEN) stem cell, is also formed during reprogramming of mouse somatic cells by ectopic expression of the transcription factors, OCT4, SOX2, KLF4, and MYC (OSKM). The mechanisms leading somatic cells to adopt differing stem cell fates are challenging to resolve given that formation of either stem cell type is slow, stochastic, and rare. For these reasons, fluorescent gene expression reporters have provided an invaluable tool for revealing the path from the somatic state to pluripotency. However, no such reporters have been established for comparable studies of iXEN cell formation. In this study, we examined the expression of multiple fluorescent reporters, including Nanog, Oct4, and the endodermal genes, Gata4 and Gata6-alone and in combination, during reprogramming. We show that only simultaneous evaluation of Nanog and Gata4 reliably distinguishes iPS and iXEN cell colonies during reprogramming.


Assuntos
Reprogramação Celular , Células-Tronco Pluripotentes Induzidas , Camundongos , Animais , Fatores de Transcrição SOXB1/genética , Fator 3 de Transcrição de Octâmero/genética , Fator 3 de Transcrição de Octâmero/metabolismo , Células-Tronco Pluripotentes Induzidas/metabolismo , Endoderma/metabolismo , Diferenciação Celular/genética
6.
Development ; 149(14)2022 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-35900100

RESUMO

Adults contracting Zika virus (ZIKV) typically exhibit mild symptoms, yet ZIKV infection of pregnant individuals can cause miscarriage or birth defects in their offspring. Many studies have focused on maternal-to-fetal ZIKV transmission via blood and placenta. Notably, however, ZIKV is also transmitted sexually, raising the possibility that ZIKV could infect the embryo shortly after fertilization, long before the placenta is established. Here, we evaluate the consequences of ZIKV infection in mouse embryos during the first few days of embryogenesis. We show that divergent strains of ZIKV can infect the fetal lineage and can cause developmental arrest, raising concern for the developmental consequences of sexual ZIKV transmission. This article has an associated 'The people behind the papers' interview.


Assuntos
Infecção por Zika virus , Zika virus , Animais , Suscetibilidade a Doenças , Feminino , Fertilização , Feto , Humanos , Transmissão Vertical de Doenças Infecciosas , Camundongos , Gravidez
7.
Cell Stem Cell ; 29(5): 661-662, 2022 05 05.
Artigo em Inglês | MEDLINE | ID: mdl-35523135

RESUMO

In this issue of Cell Stem Cell, a new study by Rostovskaya et al. (2022) uses human stem cells to understand the formation of the amnion, a vital extraembryonic tissue that encircles the fetus during gestation. They provide insight into human amnion formation and reveal differences in developmental competency of naive and primed human pluripotent stem cells.


Assuntos
Âmnio , Células-Tronco Pluripotentes , Diferenciação Celular , Humanos
8.
Development ; 148(13)2021 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-34036333

RESUMO

Fluorescent proteins and epitope tags can reveal protein localization in cells and animals, yet the large size of many tags hinders efficient genome targeting. Accordingly, many studies have relied on characterizing overexpressed proteins, which might not recapitulate endogenous protein activities. Here, we present two strategies for higher throughput production of endogenous protein reporters in mice, focusing on the blastocyst model of development. Our first strategy makes use of a split fluorescent protein, mNeonGreen2 (mNG2). Knock-in of a small portion of the mNG2 gene, in frame with gene coding regions of interest, was highly efficient in embryos, potentially obviating the need to establish mouse lines. When complemented by the larger portion of the mNG2 gene, fluorescence was reconstituted and endogenous protein localization faithfully reported in living embryos. Our second strategy achieves in-frame knock-in of a relatively small protein tag, which provides high efficiency and higher sensitivity protein reporting. Together, these two approaches provide complementary advantages and enable broad downstream applications.


Assuntos
Blastocisto/fisiologia , Genoma , Animais , Sistemas CRISPR-Cas , Linhagem Celular , Imunofluorescência , Técnicas de Introdução de Genes , Proteínas de Fluorescência Verde/genética , Camundongos , Fatores de Transcrição
9.
Nature ; 587(7834): 370-371, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-33067590
10.
Reproduction ; 160(1): V5-V7, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32484160

RESUMO

The mouse preimplantation embryo is a paradigm for discovery of the molecular principles governing formation of specific cell types during development. In this Point of View Article, we show that conditions commonly used for ex vivo culture of preimplantation development are themselves antagonistic to a pathway that is critical for blastocyst lineage commitment.


Assuntos
Blastocisto/fisiologia , Técnicas de Cultura de Células/métodos , Linhagem da Célula , Embrião de Mamíferos/citologia , Desenvolvimento Embrionário , Animais , Embrião de Mamíferos/fisiologia , Camundongos , Transdução de Sinais
11.
Development ; 146(17)2019 09 06.
Artigo em Inglês | MEDLINE | ID: mdl-31444221

RESUMO

In mice, pluripotent cells are thought to derive from cells buried inside the embryo around the 16-cell stage. Sox2 is the only pluripotency gene known to be expressed specifically within inside cells at this stage. To understand how pluripotency is established, we therefore investigated the mechanisms regulating the initial activation of Sox2 expression. Surprisingly, Sox2 expression initiated normally in the absence of both Nanog and Oct4 (Pou5f1), highlighting differences between embryo and stem cell models of pluripotency. However, we observed precocious ectopic expression of Sox2 prior to the 16-cell stage in the absence of Yap1, Wwtr1 and Tead4 Interestingly, the repression of premature Sox2 expression was sensitive to LATS kinase activity, even though LATS proteins normally do not limit activity of TEAD4, YAP1 and WWTR1 during these early stages. Finally, we present evidence for direct transcriptional repression of Sox2 by YAP1, WWTR1 and TEAD4. Taken together, our observations reveal that, while embryos are initially competent to express Sox2 as early as the four-cell stage, transcriptional repression prevents the premature expression of Sox2, thereby restricting the pluripotency program to the stage when inside cells are first created.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ligação a DNA/metabolismo , Proteínas Musculares/metabolismo , Fatores de Transcrição SOXB1/genética , Fatores de Transcrição SOXB1/metabolismo , Transativadores/metabolismo , Fatores de Transcrição/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Blastocisto/citologia , Proteínas de Ciclo Celular/genética , Proteínas de Ligação a DNA/genética , Expressão Ectópica do Gene , Embrião de Mamíferos/metabolismo , Células-Tronco Embrionárias/metabolismo , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Masculino , Camundongos/embriologia , Camundongos Transgênicos , Proteínas Musculares/genética , Células-Tronco Pluripotentes/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Fatores de Transcrição de Domínio TEA , Transativadores/genética , Fatores de Transcrição/genética , Proteínas Supressoras de Tumor/metabolismo , Proteínas de Sinalização YAP
12.
Methods Mol Biol ; 1893: 335-352, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30565145

RESUMO

The HIPPO signaling pathway plays an early and essential role in mammalian embryogenesis. The earliest known roles for HIPPO signaling during mouse development include segregating fetal and extraembryonic lineages and establishing the pluripotent progenitors of embryonic stem (ES) cells. In the mouse early embryo, HIPPO signaling responds to multiple cell biological inputs, including cell polarization, cytoskeleton, and cell environment, to influence gene expression and the first cell fate decisions in development. Methods to monitor and manipulate HIPPO signaling in the mouse early embryo are fundamental to discovering mechanisms regulating pluripotency in vivo, but properties of the early embryo, such as small cell number and spherical architecture, pose unique challenges for signaling pathway analysis. Here, we share approaches for visualizing HIPPO signaling in mouse early embryos. In addition, these methods can be applied to visualize HIPPO signaling in other spherical or cystic structures comprised of relatively few cells, such as organoids, or for the examination of other signaling pathways in these contexts.


Assuntos
Desenvolvimento Embrionário , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais , Animais , Técnicas de Cultura de Células , Transferência Embrionária , Embrião de Mamíferos , Desenvolvimento Embrionário/genética , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Genótipo , Via de Sinalização Hippo , Imuno-Histoquímica , Masculino , Camundongos , Microscopia Confocal , Proteínas Serina-Treonina Quinases/genética
13.
Elife ; 72018 12 11.
Artigo em Inglês | MEDLINE | ID: mdl-30526858

RESUMO

During mammalian development, the challenge for the embryo is to override intrinsic cellular plasticity to drive cells to distinct fates. Here, we unveil novel roles for the HIPPO signaling pathway in controlling cell positioning and expression of Sox2, the first marker of pluripotency in the mouse early embryo. We show that maternal and zygotic YAP1 and WWTR1 repress Sox2 while promoting expression of the trophectoderm gene Cdx2 in parallel. Yet, Sox2 is more sensitive than Cdx2 to Yap1/Wwtr1 dosage, leading cells to a state of conflicted cell fate when YAP1/WWTR1 activity is moderate. Remarkably, HIPPO signaling activity resolves conflicted cell fate by repositioning cells to the interior of the embryo, independent of its role in regulating Sox2 expression. Rather, HIPPO antagonizes apical localization of Par complex components PARD6B and aPKC. Thus, negative feedback between HIPPO and Par complex components ensure robust lineage segregation.


Assuntos
Linhagem da Célula/genética , Retroalimentação Fisiológica , Células-Tronco Embrionárias Murinas/metabolismo , Células-Tronco Pluripotentes/metabolismo , Proteínas Serina-Treonina Quinases/genética , Fatores de Transcrição SOXB1/genética , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Fator de Transcrição CDX2/genética , Fator de Transcrição CDX2/metabolismo , Proteínas de Ciclo Celular , Diferenciação Celular , Movimento Celular , Embrião de Mamíferos , Dosagem de Genes , Regulação da Expressão Gênica no Desenvolvimento , Via de Sinalização Hippo , Camundongos , Células-Tronco Embrionárias Murinas/citologia , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , Células-Tronco Pluripotentes/citologia , Proteína Quinase C/genética , Proteína Quinase C/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Fatores de Transcrição SOXB1/metabolismo , Transdução de Sinais , Transativadores , Proteínas de Sinalização YAP
14.
Mol Reprod Dev ; 85(7): 635-648, 2018 07.
Artigo em Inglês | MEDLINE | ID: mdl-29900695

RESUMO

Structural maintenance of chromosome flexible domain containing 1 (Smchd1) is a chromatin regulatory gene for which mutations are associated with facioscapulohumeral muscular dystrophy and arhinia. The contribution of oocyte- and zygote-expressed SMCHD1 to early development was examined in mice ( Mus musculus) using a small interfering RNA knockdown approach. Smchd1 knockdown compromised long-term embryo viability, with reduced embryo nuclear volumes at the morula stage, reduced blastocyst cell number, formation and hatching, and reduced viability to term. RNA sequencing analysis of Smchd1 knockdown morulae revealed aberrant increases in expression of a small number of trophectoderm (TE)-related genes and reduced expression of cell proliferation genes, including S-phase kinase-associated protein 2 ( Skp2). Smchd1 expression was elevated in embryos deficient for Caudal-type homeobox transcription factor 2 ( Cdx2, a key regulator of TE specification), indicating that Smchd1 is normally repressed by CDX2. These results indicate that Smchd1 plays an important role in the preimplantation embryo, regulating early gene expression and contributing to long-term embryo viability. These results extend the known functions of SMCHD1 to the preimplantation period and highlight important function for maternally expressed Smchd1 messenger RNA and protein.


Assuntos
Blastocisto/citologia , Proteínas Cromossômicas não Histona/genética , Desenvolvimento Embrionário/genética , Regulação da Expressão Gênica no Desenvolvimento/genética , Mórula/citologia , Animais , Fator de Transcrição CDX2/genética , Proliferação de Células , Sobrevivência Celular/genética , Proteínas Cromossômicas não Histona/metabolismo , Embrião de Mamíferos/citologia , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos DBA , Gravidez , Interferência de RNA , RNA Interferente Pequeno/genética , Proteínas Quinases Associadas a Fase S/biossíntese
15.
Cell ; 173(3): 544-545, 2018 04 19.
Artigo em Inglês | MEDLINE | ID: mdl-29677506

RESUMO

A new study by Zenker et al. uses time-lapse imaging to discover how dynamic actin movements contribute to epithelialization of living mouse embryos. Together with work from other labs, this study presents exciting new ways to think about the emergence of cell fates during mammalian development.


Assuntos
Actinas , Embrião de Mamíferos , Animais , Atenção , Blastocisto , Desenvolvimento Embrionário , Camundongos
16.
Curr Top Dev Biol ; 128: 181-202, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29477163

RESUMO

Cells of the early embryo are totipotent because they will differentiate to produce the fetus and its surrounding extraembryonic tissues. By contrast, embryonic stem (ES) cells are considered to be merely pluripotent because they lack the ability to efficiently produce extraembryonic cell types. The relatively limited developmental potential of ES cells can be explained by the observation that ES cells are derived from the embryo after its cells have already begun to specialize and lose totipotency. Meanwhile, at the time that pluripotent ES cell progenitors are specified, so are the multipotent progenitors of two extraembryonic stem cell types: trophoblast stem (TS) cells and extraembryonic endoderm stem (XEN) cells. Notably, all three embryo-derived stem cell types are capable of either self-renewing or differentiating in a lineage-appropriate manner. These three types of embryo-derived stem cell serve as paradigms for defining the genes and pathways that define and maintain unique stem cell identities. Remarkably, some of the mechanisms that maintain the specific developmental potential of each stem cell line do so by preventing conversion to another stem cell fate. This chapter highlights noteworthy studies that have identified the genes and pathways that normally limit the interconversion of stem cell identities.


Assuntos
Técnicas de Cultura de Células/métodos , Células-Tronco Embrionárias/citologia , Animais , Linhagem da Célula , Reprogramação Celular , Modelos Biológicos , Trofoblastos/citologia
17.
Adv Anat Embryol Cell Biol ; 229: 69-78, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29177765

RESUMO

The extraembryonic endoderm is one of the first cell types specified during mammalian development. This extraembryonic lineage is known to play multiple important roles throughout mammalian development, including guiding axial patterning and inducing formation of the first blood cells during embryogenesis. Moreover, recent studies have uncovered striking conservation between mouse and human embryos during the stages when extraembryonic endoderm cells are first specified, in terms of both gene expression and morphology. Therefore, mouse embryos serve as an excellent model for understanding the pathways that maintain extraembryonic endoderm cell fate. In addition, self-renewing multipotent stem cell lines, called XEN cells, have been derived from the extraembryonic endoderm of mouse embryos. Mouse XEN cell lines provide an additional tool for understanding the basic mechanisms that contribute to maintaining lineage potential, a resource for identifying how extraembryonic ectoderm specifies fetal cell types, and serve as a paradigm for efforts to establish human equivalents. Given the potential conservation of essential extraembryonic endoderm roles, human XEN cells would provide a considerable advance. However, XEN cell lines have not yet been successfully derived from human embryos. Given the potential utility of human XEN cell lines, this chapter focuses on reviewing the mechanisms known to govern the stem cell properties of mouse XEN, in hopes of facilitating new ways to establish human XEN cell lines.


Assuntos
Diferenciação Celular , Endoderma , Células-Tronco Multipotentes , Animais , Linhagem Celular , Embrião de Mamíferos , Endoderma/crescimento & desenvolvimento , Humanos , Camundongos , Células-Tronco Multipotentes/fisiologia
18.
Sci Rep ; 7(1): 17156, 2017 12 07.
Artigo em Inglês | MEDLINE | ID: mdl-29214996

RESUMO

The first cellular differentiation event in mouse development leads to the formation of the blastocyst consisting of the inner cell mass (ICM) and trophectoderm (TE). The transcription factor CDX2 is required for proper TE specification, where it promotes expression of TE genes, and represses expression of Pou5f1 (OCT4). However its downstream network in the developing embryo is not fully characterized. Here, we performed high-throughput single embryo qPCR analysis in Cdx2 null embryos to identify CDX2-regulated targets in vivo. To identify genes likely to be regulated by CDX2 directly, we performed CDX2 ChIP-Seq on trophoblast stem (TS) cells. In addition, we examined the dynamics of gene expression changes using inducible CDX2 embryonic stem (ES) cells, so that we could predict which CDX2-bound genes are activated or repressed by CDX2 binding. By integrating these data with observations of chromatin modifications, we identify putative novel regulatory elements that repress gene expression in a lineage-specific manner. Interestingly, we found CDX2 binding sites within regulatory elements of key pluripotent genes such as Pou5f1 and Nanog, pointing to the existence of a novel mechanism by which CDX2 maintains repression of OCT4 in trophoblast. Our study proposes a general mechanism in regulating lineage segregation during mammalian development.


Assuntos
Fator de Transcrição CDX2/metabolismo , Linhagem da Célula/genética , Embrião de Mamíferos/citologia , Células-Tronco Embrionárias/citologia , Regulação da Expressão Gênica no Desenvolvimento , Transcrição Gênica , Trofoblastos/citologia , Animais , Fator de Transcrição CDX2/genética , Diferenciação Celular , Células Cultivadas , Embrião de Mamíferos/fisiologia , Células-Tronco Embrionárias/fisiologia , Camundongos , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Análise de Célula Única , Transcriptoma , Trofoblastos/fisiologia
19.
J Biol Chem ; 292(10): 4138-4151, 2017 03 10.
Artigo em Inglês | MEDLINE | ID: mdl-28126904

RESUMO

Transforming growth factor ß (TGF-ß) pathways are key determinants of cell fate in animals. Their basic mechanism of action is simple. However, to produce cell-specific responses, TGF-ß pathways are heavily regulated by secondary factors, such as membrane-associated EGF-CFC family proteins. Cellular activities of EGF-CFC proteins have been described, but their molecular functions, including how the mammalian homologs Cripto-1 and Cryptic recognize and regulate TGF-ß family ligands, are less clear. Here we use purified human Cripto-1 and mouse Cryptic produced in mammalian cells to show that these two EGF-CFC homologs have distinct, highly specific ligand binding activities. Cripto-1 interacts with BMP-4 in addition to its known partner Nodal, whereas Cryptic interacts only with Activin B. These interactions depend on the integrity of the protein, as truncated or deglycosylated Cripto-1 lacked BMP-4 binding activity. Significantly, Cripto-1 and Cryptic blocked binding of their cognate ligands to type I and type II TGF-ß receptors, indicating that Cripto-1 and Cryptic contact ligands at their receptor interaction surfaces and, thus, that they could inhibit their ligands. Indeed, soluble Cripto-1 and Cryptic inhibited ligand signaling in various cell-based assays, including SMAD-mediated luciferase reporter gene expression, and differentiation of a multipotent stem cell line. But in agreement with previous work, the membrane bound form of Cripto-1 potentiated signaling, revealing a critical role of membrane association for its established cellular activity. Thus, our studies provide new insights into the mechanism of ligand recognition by this enigmatic family of membrane-anchored TGF-ß family signaling regulators and link membrane association with their signal potentiating activities.


Assuntos
Membrana Celular/metabolismo , Proteínas Ligadas por GPI/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Proteínas de Neoplasias/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Sequência de Aminoácidos , Diferenciação Celular , Células Hep G2 , Humanos , Ligantes , Ligação Proteica , Receptor do Fator de Crescimento Transformador beta Tipo II , Homologia de Sequência de Aminoácidos , Transdução de Sinais
20.
Trends Genet ; 32(8): 457-458, 2016 08.
Artigo em Inglês | MEDLINE | ID: mdl-27256906

RESUMO

Understanding how and when cells become different during embryogenesis is a goal that is at the forefront of investigations in mammalian development. Two recent studies from the laboratories of Nicholas Plachta and Magdalena Zernicka-Goetz present evidence that cellular heterogeneities detected in four-cell mouse embryos bias the process of cell fate acquisition thereafter.


Assuntos
Padronização Corporal/genética , Diferenciação Celular/genética , Desenvolvimento Embrionário/genética , Animais , Linhagem da Célula/genética , Embrião de Mamíferos , Mamíferos , Camundongos
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