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1.
Mol Biol Evol ; 37(1): 100-109, 2020 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-31504761

RESUMO

The GLIS family transcription factors, GLIS1 and GLIS3, potentiate generation of induced pluripotent stem cells (iPSCs). In contrast, another GLIS family member, GLIS2, suppresses cell reprograming. To understand how these disparate roles arose, we examined evolutionary origins and genomic organization of GLIS genes. Comprehensive phylogenetic analysis shows that GLIS1 and GLIS3 originated during vertebrate whole genome duplication, whereas GLIS2 is a sister group to the GLIS1/3 and GLI families. This result is consistent with their opposing functions in cell reprograming. Glis1 evolved faster than Glis3, losing many protein-interacting motifs. This suggests that Glis1 acquired new functions under weakened evolutionary constraints. In fact, GLIS1 induces induced pluripotent stem cells more strongly. Transcriptomic data from various animal embryos demonstrate that glis1 is maternally expressed in some tetrapods, whereas vertebrate glis3 and invertebrate glis1/3 genes are rarely expressed in oocytes, suggesting that vertebrate (or tetrapod) Glis1 acquired a new expression domain and function as a maternal factor. Furthermore, comparative genomic analysis reveals that glis1/3 is part of a bilaterian-specific gene cluster, together with rfx3, ndc1, hspb11, and lrrc42. Because known functions of these genes are related to cilia formation and function, the last common ancestor of bilaterians may have acquired this cluster by shuffling gene order to establish more sophisticated epithelial tissues involving cilia. This evolutionary study highlights the significance of GLIS1/3 for cell reprograming, development, and diseases in ciliated organs such as lung, kidney, and pancreas.


Assuntos
Evolução Molecular , Fatores de Transcrição Kruppel-Like/genética , Motivos de Aminoácidos , Animais , Reprogramação Celular , Humanos , Fatores de Transcrição Kruppel-Like/metabolismo , Oócitos/metabolismo , Filogenia , Sintenia
2.
Development ; 145(5)2018 03 12.
Artigo em Inglês | MEDLINE | ID: mdl-29440302

RESUMO

The homeodomain transcription factor Otx2 has essential roles in head and eye formation via the negative and positive regulation of its target genes, but it remains elusive how this dual activity of Otx2 affects cellular functions. In the current study, we first demonstrated that both exogenous and endogenous Otx2 are phosphorylated at multiple sites. Using Xenopus embryos, we identified three possible cyclin-dependent kinase (Cdk) sites and one Akt site, and analyzed the biological activities of phosphomimetic (4E) and nonphosphorylatable (4A) mutants for those sites. In the neuroectoderm, the 4E but not the 4A mutant downregulated the Cdk inhibitor gene p27xic1 (cdknx) and posterior genes, and promoted cell proliferation, possibly forming a positive-feedback loop consisting of Cdk, Otx2 and p27xic1 for cell proliferation, together with anteriorization. Conversely, the 4A mutant functioned as an activator on its own and upregulated the expression of eye marker genes, resulting in enlarged eyes. Consistent with these results, the interaction of Otx2 with the corepressor Tle1 is suggested to be phosphorylation dependent. These data suggest that Otx2 orchestrates cell proliferation, anteroposterior patterning and eye formation via its phosphorylation state.


Assuntos
Padronização Corporal , Proliferação de Células , Fatores de Transcrição Otx/metabolismo , Xenopus laevis/embriologia , Xenopus laevis/metabolismo , Animais , Animais Geneticamente Modificados , Padronização Corporal/genética , Quinases Ciclina-Dependentes/metabolismo , Embrião não Mamífero , Olho/embriologia , Olho/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Proteína Oncogênica v-akt/metabolismo , Fosforilação , Processamento de Proteína Pós-Traducional , Xenopus laevis/genética
3.
Dev Growth Differ ; 62(6): 379-390, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32275068

RESUMO

The notochord is a defining feature of chordates. During notochord formation in vertebrates and tunicates, notochord cells display dynamic morphogenetic movement, called convergent extension, in which cells intercalate and align at the dorsal midline. However, in cephalochordates, the most basal group of chordates, the notochord is formed without convergent extension. It is simply developed from mesodermal cells at the dorsal midline. This suggests that convergent extension movement of notochord cells is a secondarily acquired developmental attribute in the common ancestor of olfactores (vertebrates + tunicates), and that the chordate ancestor innovated the notochord upon a foundation of morphogenetic mechanisms independent of cell movement. Therefore, this review focuses on biological features specific to notochord cells, which have been well studied using clawed frogs, zebrafish, and tunicates. Attributes of notochord cells, such as vacuolation, membrane trafficking, extracellular matrix formation, and apoptosis, can be understood in terms of two properties: turgor pressure of vacuoles and strength of the notochord sheath. To maintain the straight rod-like structure of the notochord, these parameters must be counterbalanced. In the future, the turgor pressure-sheath strength model, proposed in this review, will be examined in light of quantitative molecular data and mathematical simulations, illuminating the evolutionary origin of the notochord.


Assuntos
Modelos Biológicos , Morfogênese , Notocorda/crescimento & desenvolvimento , Notocorda/metabolismo , Animais , Apoptose , Proliferação de Células , Notocorda/citologia
4.
Dev Growth Differ ; 62(5): 279-300, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32479656

RESUMO

Chordates comprise three major groups, cephalochordates (amphioxus), tunicates (urochordates), and vertebrates. Since cephalochordates were the early branching group, comparisons between amphioxus and other chordates help us to speculate about ancestral chordates. Here, I summarize accumulating data from functional studies analyzing amphioxus cis-regulatory modules (CRMs) in model systems of other chordate groups, such as mice, chickens, clawed frogs, fish, and ascidians. Conservatism and variability of CRM functions illustrate how gene regulatory networks have evolved in chordates. Amphioxus CRMs, which correspond to CRMs deeply conserved among animal phyla, govern reporter gene expression in conserved expression domains of the putative target gene in host animals. In addition, some CRMs located in similar genomic regions (intron, upstream, or downstream) also possess conserved activity, even though their sequences are divergent. These conservative CRM functions imply ancestral genomic structures and gene regulatory networks in chordates. However, interestingly, if expression patterns of amphioxus genes do not correspond to those of orthologs of experimental models, some amphioxus CRMs recapitulate expression patterns of amphioxus genes, but not those of endogenous genes, suggesting that these amphioxus CRMs are close to the ancestral states of chordate CRMs, while vertebrates/tunicates innovated new CRMs to reconstruct gene regulatory networks subsequent to the divergence of the cephalochordates. Alternatively, amphioxus CRMs may have secondarily lost ancestral CRM activity and evolved independently. These data help to solve fundamental questions of chordate evolution, such as neural crest cells, placodes, a forebrain/midbrain, and genome duplication. Experimental validation is crucial to verify CRM functions and evolution.


Assuntos
Evolução Biológica , Cordados/genética , Elementos Facilitadores Genéticos/genética , Regulação da Expressão Gênica no Desenvolvimento/genética , Animais , Cordados/metabolismo
5.
Dev Biol ; 426(2): 270-290, 2017 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-28089430

RESUMO

Cell signaling pathways, such as Wnt, Hedgehog (Hh), Notch, and Hippo, are essential for embryogenesis, organogenesis, and tissue homeostasis. In this study, we analyzed 415 genes involved in these pathways in the allotetraploid frog, Xenopus laevis. Most genes are retained in two subgenomes called L and S (193 homeologous gene pairs and 29 singletons). This conservation rate of homeologs is much higher than that of all genes in the X. laevis genome (86.9% vs 60.2%). Among singletons, 24 genes are retained in the L subgenome, a rate similar to the average for all genes (82.8% vs 74.6%). In addition, as general components of signal transduction, we also analyzed 32 heparan sulfate proteoglycan (HSPG)-related genes and eight TLE/Groucho transcriptional corepressors-related genes. In these gene sets, all homeologous pairs have been retained. Transcriptome analysis using RNA-seq data from developmental stages and adult tissues demonstrated that most homeologous pairs of signaling components have variable expression patterns, in contrast to the conservative expression profiles of homeologs for transcription factors. Our results indicate that homeologous gene pairs for cell signaling regulation have tended to become subfunctionalized after allotetraploidization. Diversification of signaling pathways by subfunctionalization of homeologs may enhance environmental adaptability. These results provide insights into the evolution of signaling pathways after polyploidization.


Assuntos
Perfilação da Expressão Gênica , Proteínas Hedgehog/genética , Receptores Notch/genética , Transdução de Sinais/genética , Proteínas Wnt/genética , Proteínas de Xenopus/genética , Xenopus laevis/genética , Animais , Receptores Frizzled/biossíntese , Receptores Frizzled/genética , Expressão Gênica , Genoma , Proteínas Hedgehog/biossíntese , Anotação de Sequência Molecular , Receptores Notch/biossíntese , Frações Subcelulares/metabolismo , Sintenia , Tetraploidia , Transcriptoma , Proteínas Wnt/biossíntese , Via de Sinalização Wnt/genética , Proteínas de Xenopus/biossíntese
6.
Dev Biol ; 426(2): 301-324, 2017 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-27810169

RESUMO

Xenopus laevis has an allotetraploid genome of 3.1Gb, in contrast to the diploid genome of a closely related species, Xenopus tropicalis. Here, we identified 412 genes (189 homeolog pairs, one homeologous gene cluster pair, and 28 singletons) encoding transcription factors (TFs) in the X. laevis genome by comparing them with their orthologs from X. tropicalis. Those genes include the homeobox gene family (Mix/Bix, Lhx, Nkx, Paired, POU, and Vent), Sox, Fox, Pax, Dmrt, Hes, GATA, T-box, and some clock genes. Most homeolog pairs for TFs are retained in two X. laevis subgenomes, named L and S, at higher than average rates (87.1% vs 60.2%). Among the 28 singletons, 82.1% were deleted from chromosomes of the S subgenome, a rate similar to the genome-wide average (82.1% vs 74.6%). Interestingly, nkx2-1, nkx2-8, and pax9, which reside consecutively in a postulated functional gene cluster, were deleted from the S chromosome, suggesting cluster-level gene regulation. Transcriptome correlation analysis demonstrated that TF homeolog pairs tend to have more conservative developmental expression profiles than most other types of genes. In some cases, however, either of the homeologs may show strongly different spatio-temporal expression patterns, suggesting neofunctionalization, subfunctionalization, or nonfunctionalization after allotetraploidization. Analyses of otx1 suggests that homeologs with much lower expression levels have undergone greater amino acid sequence diversification. Our comprehensive study implies that TF homeologs are highly conservative after allotetraploidization, possibly because the DNA sequences that they bind were also duplicated, but in some cases, they differed in expression levels or became singletons due to dosage-sensitive regulation of their target genes.


Assuntos
Perfilação da Expressão Gênica , Fatores de Transcrição/genética , Xenopus laevis/genética , Animais
7.
Biochem Biophys Res Commun ; 451(4): 522-8, 2014 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-25111819

RESUMO

It has been reported that GCS1 (Generative Cell Specific 1) is a transmembrane protein that is exclusively expressed in sperm cells and is essential for gamete fusion in flowering plants. The GCS1 gene is present not only in angiosperms but also in unicellular organisms and animals, implying the occurrence of a common or ancestral mechanism of GCS1-mediated gamete fusion. In order to elucidate the common mechanism, we investigated the role of GCS1 in animal fertilization using a sea anemone (Cnidaria), Nematostella vectensis. Although the existence of the GCS1 gene in N. vectensis has been reported, the expression of GCS1 in sperm and the role of GCS1 in fertilization are not known. In this study, we showed that the GCS1 gene is expressed in the testis and that GCS1 protein exists in sperm by in situ hybridization and proteomic analysis, respectively. Then we made four peptide antibodies against the N-terminal extracellular region of NvGCS1. These antibodies specifically reacted to NvGCS1 among sperm proteins on the basis of Western analysis and potently inhibited fertilization in a concentration-dependent manner. These results indicate that sperm GCS1 plays a pivotal role in fertilization, most probably in sperm-egg fusion, in a starlet sea anemone, suggesting a common gamete-fusion mechanism shared by eukaryotic organisms.


Assuntos
Fertilização/fisiologia , Hormônios de Invertebrado/fisiologia , Proteínas de Membrana/fisiologia , Interações Espermatozoide-Óvulo/efeitos dos fármacos , Sequência de Aminoácidos , Animais , Masculino , Plantas/genética , Anêmonas-do-Mar , Alinhamento de Sequência , Espermatozoides/metabolismo
8.
Front Cell Dev Biol ; 11: 957805, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36998246

RESUMO

Keratan sulfate (KS) is a glycosaminoglycan that is enriched in vertebrate cornea, cartilage, and brain. During embryonic development, highly sulfated KS (HSKS) is first detected in the developing notochord and then in otic vesicles; therefore, HSKS has been used as a molecular marker of the notochord. However, its biosynthetic pathways and functional roles in organogenesis are little known. Here, I surveyed developmental expression patterns of genes related to HSKS biosynthesis in Xenopus embryos. Of these genes, the KS chain-synthesizing glycosyltransferase genes, beta-1,3-N-acetylglucosaminyltransferase (b3gnt7) and beta-1,4-galactosyltransferase (b4galt4), are strongly expressed in the notochord and otic vesicles, but also in other tissues. In addition, their notochord expression is gradually restricted to the posterior end at the tailbud stage. In contrast, carbohydrate sulfotransferase (Chst) genes, chst2, chst3, and chst5.1, are expressed in both notochord and otic vesicles, whereas chst1, chst4/5-like, and chst7 are confined to otic vesicles. Because the substrate for Chst1 and Chst3 is galactose, while that for others is N-acetylglucosamine, combinatorial, tissue-specific expression patterns of Chst genes should be responsible for tissue-specific HSKS enrichment in embryos. As expected, loss of function of chst1 led to loss of HSKS in otic vesicles and reduction of their size. Loss of chst3 and chst5.1 resulted in HSKS loss in the notochord. These results reveal that Chst genes are critical for HSKS biosynthesis during organogenesis. Being hygroscopic, HSKS forms "water bags" in embryos to physically maintain organ structures. In terms of evolution, in ascidian embryos, b4galt and chst-like genes are also expressed in the notochord and regulate notochord morphogenesis. Furthermore, I found that a chst-like gene is also strongly expressed in the notochord of amphioxus embryos. These conserved expression patterns of Chst genes in the notochord of chordate embryos suggest that Chst is an ancestral component of the chordate notochord.

9.
Cell Rep ; 38(7): 110364, 2022 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-35172134

RESUMO

Mesendodermal specification is one of the earliest events in embryogenesis, where cells first acquire distinct identities. Cell differentiation is a highly regulated process that involves the function of numerous transcription factors (TFs) and signaling molecules, which can be described with gene regulatory networks (GRNs). Cell differentiation GRNs are difficult to build because existing mechanistic methods are low throughput, and high-throughput methods tend to be non-mechanistic. Additionally, integrating highly dimensional data composed of more than two data types is challenging. Here, we use linked self-organizing maps to combine chromatin immunoprecipitation sequencing (ChIP-seq)/ATAC-seq with temporal, spatial, and perturbation RNA sequencing (RNA-seq) data from Xenopus tropicalis mesendoderm development to build a high-resolution genome scale mechanistic GRN. We recover both known and previously unsuspected TF-DNA/TF-TF interactions validated through reporter assays. Our analysis provides insights into transcriptional regulation of early cell fate decisions and provides a general approach to building GRNs using highly dimensional multi-omic datasets.


Assuntos
Endoderma/embriologia , Redes Reguladoras de Genes , Genômica , Mesoderma/embriologia , Xenopus/embriologia , Xenopus/genética , Animais , Cromatina/metabolismo , Sequência Consenso/genética , DNA/metabolismo , Gastrulação/genética , Regulação da Expressão Gênica no Desenvolvimento , Ligação Proteica , RNA/metabolismo , Fatores de Transcrição/metabolismo , Transcrição Gênica
10.
Curr Top Dev Biol ; 145: 113-166, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34074528

RESUMO

The field of molecular embryology started around 1990 by identifying new genes and analyzing their functions in early vertebrate embryogenesis. Those genes encode transcription factors, signaling molecules, their regulators, etc. Most of those genes are relatively highly expressed in specific regions or exhibit dramatic phenotypes when ectopically expressed or mutated. This review focuses on one of those genes, Lim1/Lhx1, which encodes a transcription factor. Lim1/Lhx1 is a member of the LIM homeodomain (LIM-HD) protein family, and its intimate partner, Ldb1/NLI, binds to two tandem LIM domains of LIM-HDs. The most ancient LIM-HD protein and its partnership with Ldb1 were innovated in the metazoan ancestor by gene fusion combining LIM domains and a homeodomain and by creating the LIM domain-interacting domain (LID) in ancestral Ldb, respectively. The LIM domain has multiple interacting interphases, and Ldb1 has a dimerization domain (DD), the LID, and other interacting domains that bind to Ssbp2/3/4 and the boundary factor, CTCF. By means of these domains, LIM-HD-Ldb1 functions as a hub protein complex, enabling more intricate and elaborate gene regulation. The common, ancestral role of LIM-HD proteins is neuron cell-type specification. Additionally, Lim1/Lhx1 serves crucial roles in the gastrula organizer and in kidney development. Recent studies using Xenopus embryos have revealed Lim1/Lhx1 functions and regulatory mechanisms during development and regeneration, providing insight into evolutionary developmental biology, functional genomics, gene regulatory networks, and regenerative medicine. In this review, we also discuss recent progress at unraveling participation of Ldb1, Ssbp, and CTCF in enhanceosomes, long-distance enhancer-promoter interactions, and trans-interactions between chromosomes.


Assuntos
Proteínas com Homeodomínio LIM/metabolismo , Animais , Biologia do Desenvolvimento , Genômica , Humanos , Ligação Proteica , Medicina Regenerativa
11.
Cold Spring Harb Protoc ; 2019(1)2019 01 02.
Artigo em Inglês | MEDLINE | ID: mdl-30131366

RESUMO

Introducing exogenous DNA into an embryo can promote misexpression of a gene of interest via transcription regulated by an attached enhancer-promoter. This protocol describes plasmid DNA microinjection into Xenopus embryos for misexpression of genes after zygotic gene expression begins. It also describes a method for coinjecting a reporter plasmid with mRNA or antisense morpholinos to perform luciferase reporter assays, which are useful for quantitative analysis of cis-regulatory sequences responding to endogenous or exogenous stimuli in embryos.


Assuntos
DNA/administração & dosagem , Regulação da Expressão Gênica no Desenvolvimento , Microinjeções/métodos , Xenopus/embriologia , Animais , Plasmídeos/administração & dosagem
12.
Zoological Lett ; 5: 27, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31388442

RESUMO

BACKGROUND: In cephalochordates (amphioxus), the notochord runs along the dorsal to the anterior tip of the body. In contrast, the vertebrate head is formed anterior to the notochord, as a result of head organizer formation in anterior mesoderm during early development. A key gene for the vertebrate head organizer, goosecoid (gsc), is broadly expressed in the dorsal mesoderm of amphioxus gastrula. Amphioxus gsc expression subsequently becomes restricted to the posterior notochord from the early neurula. This has prompted the hypothesis that a change in expression patterns of gsc led to development of the vertebrate head during chordate evolution. However, molecular mechanisms of head organizer evolution involving gsc have never been elucidated. RESULTS: To address this question, we compared cis-regulatory modules of vertebrate organizer genes between amphioxus, Branchiostoma japonicum, and frogs, Xenopus laevis and Xenopus tropicalis. Here we show conservation and diversification of gene regulatory mechanisms through cis-regulatory modules for gsc, lim1/lhx1, and chordin in Branchiostoma and Xenopus. Reporter analysis using Xenopus embryos demonstrates that activation of gsc by Nodal/FoxH1 signal through the 5' upstream region, that of lim1 by Nodal/FoxH1 signal through the first intron, and that of chordin by Lim1 through the second intron, are conserved between amphioxus and Xenopus. However, activation of gsc by Lim1 and Otx through the 5' upstream region in Xenopus are not conserved in amphioxus. Furthermore, the 5' region of amphioxus gsc recapitulated the amphioxus-like posterior mesoderm expression of the reporter gene in transgenic Xenopus embryos. CONCLUSIONS: On the basis of this study, we propose a model, in which the gsc gene acquired the cis-regulatory module bound with Lim1 and Otx at its 5' upstream region to be activated persistently in anterior mesoderm, in the vertebrate lineage. Because Gsc globally represses trunk (notochord) genes in the vertebrate head organizer, this cooption of gsc in vertebrates appears to have resulted in inhibition of trunk genes and acquisition of the head organizer and its derivative prechordal plate.

13.
Biol Open ; 8(3)2019 Mar 04.
Artigo em Inglês | MEDLINE | ID: mdl-30651235

RESUMO

hox genes are found as clusters in the genome in most bilaterians. The order of genes in the cluster is supposed to be correlated with the site of expression along the anterior-posterior body axis and the timing of expression during development, and these correlations are called spatial and temporal collinearity, respectively. Here we studied the expression dynamics of all hox genes of the diploid species Xenopus tropicalis in four Hox clusters (A-D) by analyzing high-temporal-resolution RNA-seq databases and the results showed that temporal collinearity is not supported, which is consistent with our previous data from allotetraploid X enopus laevis Because the temporal collinearity hypothesis implicitly assumes the collinear order of gene activation, not mRNA accumulation, we determined for the first time the timing of when new transcripts of hox genes are produced, by detecting pre-spliced RNA in whole embryos with reverse transcription and quantitative PCR (RT-qPCR) for all hoxa genes as well as several selected hoxb, hox c and hoxd genes. Our analyses showed that, coinciding with the RNA-seq results, hoxa genes started to be transcribed in a non-sequential order, and found that multiple genes start expression almost simultaneously or more posterior genes could be expressed earlier than anterior ones. This tendency was also found in hoxb and hoxc genes. These results suggest that temporal collinearity of hox genes is not held during early development of Xenopus.

14.
Zoological Lett ; 3: 4, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28344820

RESUMO

BACKGROUND: The T-box family transcription-factor gene, Brachyury, has two expression domains with discrete functions during animal embryogenesis. The primary domain, associated with the blastopore, is shared by most metazoans, while the secondary domain, involved in the notochord, is specific to chordates. In most animals, Brachyury is present in a single copy, but in cephalochordates, the most basal of the chordates, the gene is present in two copies, suggesting allotment of the two domains to each of the duplicates. RESULTS: In order to clarify whether Brachyury duplication occurred in the common ancestor of chordates after which one of duplicates was lost in the urochordate and vertebrate lineages, we estimated phylogenetic relationships of Brachyury genes and examined the synteny of a Brachyury-containing genomic region of deuterostomes with decoded genomes. The monophyletic origin of tandemly arranged Brachyury genes of cephalochordates indicates that the tandem duplication occurred in the cephalochordate lineage, but not in the chordate ancestor. CONCLUSIONS: Our results thus suggest that, in the common ancestor of chordates, a single copy of Brachyury acquired two expression domains and that the duplication was not involved in the acquisition of the notochord. However, in relation to regulatory mechanisms, both possibilities-namely a single copy with two domains and two copies with different domains-should be considered in future studies of Brachyury.

15.
Dev Cell ; 40(6): 595-607.e4, 2017 03 27.
Artigo em Inglês | MEDLINE | ID: mdl-28325473

RESUMO

The interplay between transcription factors and chromatin dictates gene regulatory network activity. Germ layer specification is tightly coupled with zygotic gene activation and, in most metazoans, is dependent upon maternal factors. We explore the dynamic genome-wide interactions of Foxh1, a maternal transcription factor that mediates Nodal/TGF-ß signaling, with cis-regulatory modules (CRMs) during mesendodermal specification. Foxh1 marks CRMs during cleavage stages and recruits the co-repressor Tle/Groucho in the early blastula. We highlight a population of CRMs that are continuously occupied by Foxh1 and show that they are marked by H3K4me1, Ep300, and Fox/Sox/Smad motifs, suggesting interplay between these factors in gene regulation. We also propose a molecular "hand-off" between maternal Foxh1 and zygotic Foxa at these CRMs to maintain enhancer activation. Our findings suggest that Foxh1 functions at the top of a hierarchy of interactions by marking developmental genes for activation, beginning with the onset of zygotic gene expression.


Assuntos
Endoderma/metabolismo , Fatores de Transcrição Forkhead/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Mesoderma/metabolismo , Fatores de Transcrição/metabolismo , Proteínas de Xenopus/metabolismo , Xenopus/embriologia , Xenopus/genética , Animais , Blástula/metabolismo , Fase de Clivagem do Zigoto/metabolismo , Proteínas Correpressoras/metabolismo , Embrião não Mamífero/metabolismo , Endoderma/embriologia , Elementos Facilitadores Genéticos/genética , Fatores de Transcrição Forkhead/genética , Genoma , Histonas/metabolismo , Lisina/metabolismo , Mesoderma/embriologia , Metilação , Proteína Nodal/metabolismo , Ligação Proteica/genética , RNA Polimerase II/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Sequências Reguladoras de Ácido Nucleico/genética , Análise de Sequência de RNA , Transdução de Sinais/genética , Transcrição Gênica , Xenopus/metabolismo , Proteínas de Xenopus/genética
16.
Curr Biol ; 26(21): 2885-2892, 2016 11 07.
Artigo em Inglês | MEDLINE | ID: mdl-27693135

RESUMO

Blastoporal expression of the T-box transcription factor gene brachyury is conserved in most metazoans [1, 2]. Its role in mesoderm formation has been intensively studied in vertebrates [3-6]. However, its fundamental function near the blastopore is poorly understood in other phyla. Cnidarians are basal metazoans that are important for understanding evolution of metazoan body plans [7, 8]. Because they lack mesoderm, they have been used to investigate the evolutionary origins of mesoderm [1, 9-11]. Here, we focus on corals, a primitive clade of cnidarians that diverged from sea anemones ∼500 mya [12]. We developed a microinjection method for coral eggs to examine Brachyury functions during embryogenesis of the scleractinian coral, Acropora digitifera. Because Acropora embryos undergo pharynx formation after the blastopore closes completely [13-15], they are useful to understand Brachyury functions in gastrulation movement and pharynx formation. We show that blastoporal expression of brachyury is directly activated by Wnt/ß-catenin signaling in the ectoderm of coral embryos, indicating that the regulatory axis from Wnt/ß-catenin signaling to brachyury is highly conserved among eumetazoans. Loss-of-function analysis demonstrated that Brachyury is required for pharynx formation but not for gastrulation movement. Genome-wide transcriptome analysis demonstrated that genes positively regulated by Brachyury are expressed in the ectoderm of Acropora gastrulae, while negatively regulated genes are in endoderm. Therefore, germ layer demarcation around the blastopore appears to be the evolutionarily conserved role of Brachyury during gastrulation. Compared with Brachyury functions in vertebrate mesoderm-ectoderm and mesoderm-endoderm demarcation [4-6], our results suggest that the vertebrate-type mesoderm may have originated from brachyury-expressing ectoderm adjacent to endoderm.


Assuntos
Antozoários/embriologia , Antozoários/genética , Proteínas Fetais/genética , Proteínas com Domínio T/genética , Animais , Ectoderma/embriologia , Endoderma/embriologia , Proteínas Fetais/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Proteínas com Domínio T/metabolismo
17.
Nat Commun ; 5: 4322, 2014 Jul 09.
Artigo em Inglês | MEDLINE | ID: mdl-25005894

RESUMO

Head specification by the head-selector gene, orthodenticle (otx), is highly conserved among bilaterian lineages. However, the molecular mechanisms by which Otx and other transcription factors (TFs) interact with the genome to direct head formation are largely unknown. Here we employ ChIP-seq and RNA-seq approaches in Xenopus tropicalis gastrulae and find that occupancy of the corepressor, TLE/Groucho, is a better indicator of tissue-specific cis-regulatory modules (CRMs) than the coactivator p300, during early embryonic stages. On the basis of TLE binding and comprehensive CRM profiling, we define two distinct types of Otx2- and TLE-occupied CRMs. Using these devices, Otx2 and other head organizer TFs (for example, Lim1/Lhx1 (activator) or Goosecoid (repressor)) are able to upregulate or downregulate a large battery of target genes in the head organizer. An underlying principle is that Otx marks target genes for head specification to be regulated positively or negatively by partner TFs through specific types of CRMs.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Cabeça/embriologia , Fatores de Transcrição Otx/metabolismo , Proteínas de Xenopus/metabolismo , Xenopus/metabolismo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Sítios de Ligação , Feminino , Gástrula/embriologia , Gástrula/metabolismo , Masculino , Especificidade de Órgãos , Fatores de Transcrição Otx/genética , Regiões Promotoras Genéticas , Ligação Proteica , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Xenopus/embriologia , Xenopus/genética , Proteínas de Xenopus/genética
18.
Curr Biol ; 22(7): 601-7, 2012 Apr 10.
Artigo em Inglês | MEDLINE | ID: mdl-22386310

RESUMO

Teleosts have an asymmetrical caudal fin skeleton formed by the upward bending of the caudal-most portion of the body axis, the ural region. This homocercal type of caudal fin ensures powerful and complex locomotion and is regarded as one of the most important innovations for teleosts during adaptive radiation in an aquatic environment. However, the mechanisms that create asymmetric caudal fin remain largely unknown. The spontaneous medaka (teleost fish) mutant, Double anal fin (Da), exhibits a unique symmetrical caudal skeleton that resembles the diphycercal type seen in Polypterus and Coelacanth. We performed a detailed analysis of the Da mutant to obtain molecular insight into caudal fin morphogenesis. We first demonstrate that a large transposon, inserted into the enhancer region of the zic1 and zic4 genes (zic1/zic4) in Da, is associated with the mesoderm-specific loss of their transcription. We then show that zic1/zic4 are strongly expressed in the dorsal part of the ural mesenchyme and thereby induce asymmetric caudal fin development in wild-type embryos, whereas their expression is lost in Da. Comparative analysis further indicates that the dorsal mesoderm expression of zic1/zic4 is conserved in teleosts, highlighting the crucial role of zic1/zic4 in caudal fin morphogenesis.


Assuntos
Nadadeiras de Animais/embriologia , Proteínas de Peixes/genética , Morfogênese , Oryzias/embriologia , Oryzias/genética , Fatores de Transcrição/genética , Nadadeiras de Animais/metabolismo , Animais , Evolução Biológica , Elementos de DNA Transponíveis , Proteínas de Peixes/metabolismo , Mesoderma/embriologia , Mesoderma/metabolismo , Dados de Sequência Molecular , Mutação , Oryzias/metabolismo , Reação em Cadeia da Polimerase , Cauda/embriologia , Cauda/metabolismo , Fatores de Transcrição/metabolismo , Dedos de Zinco
19.
Development ; 136(12): 2005-14, 2009 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-19439497

RESUMO

Expression of the LIM homeobox gene lhx1 (lim1) is specific to the vertebrate gastrula organizer. Lhx1 functions as a transcriptional regulatory core protein to exert ;organizer' activity in Xenopus embryos. Its ancient paralog, lhx3 (lim3), is expressed around the blastopore in amphioxus and ascidian, but not vertebrate, gastrulae. These two genes are thus implicated in organizer evolution, and we addressed the evolutionary origins of their blastoporal expression and organizer activity. Gene expression analysis of organisms ranging from cnidarians to chordates suggests that blastoporal expression has its evolutionary root in or before the ancestral eumetazoan for lhx1, but possibly in the ancestral chordate for lhx3, and that in the ascidian lineage, blastoporal expression of lhx1 ceased, whereas endodermal expression of lhx3 has persisted. Analysis of organizer activity using Xenopus embryos suggests that a co-factor of LIM homeodomain proteins, Ldb, has a conserved function in eumetazoans to activate Lhx1, but that Lhx1 acquired organizer activity in the bilaterian lineage, Lhx3 acquired organizer activity in the deuterostome lineage and ascidian Lhx3 acquired a specific transactivation domain to confer organizer activity on this molecule. Knockdown analysis using cnidarian embryos suggests that Lhx1 is required for chordin expression in the blastoporal region. These data suggest that Lhx1 has been playing fundamental roles in the blastoporal region since the ancestral eumetazoan arose, that it contributed as an 'original organizer gene' to the evolution of the vertebrate gastrula organizer, and that Lhx3 could be involved in the establishment of organizer gene networks.


Assuntos
Evolução Molecular , Gástrula/fisiologia , Proteínas de Homeodomínio/fisiologia , Organizadores Embrionários/fisiologia , Proteínas de Xenopus/fisiologia , Sequência de Aminoácidos , Animais , Padronização Corporal/fisiologia , Embrião não Mamífero/fisiologia , Gástrula/embriologia , Regulação da Expressão Gênica no Desenvolvimento , Técnicas de Silenciamento de Genes , Glicoproteínas/metabolismo , Proteínas de Homeodomínio/genética , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Proteínas com Homeodomínio LIM , Dados de Sequência Molecular , Anêmonas-do-Mar/embriologia , Anêmonas-do-Mar/genética , Anêmonas-do-Mar/metabolismo , Fatores de Transcrição , Xenopus/embriologia , Proteínas de Xenopus/genética
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