Bra (SpBra) is the single sea urchin orthologue of the vertebrate Brachyury (T/TBXT) transcription factor, a 503-residue protein with an N-terminal T-box DNA-binding domain. It is a node of the endoderm gene regulatory network of the embryo. Zygotic transcription begins in the vegetal plate, where it is confined to the presumptive endoderm of the veg2 lineage and depends on nuclear beta-catenin/Tcf together with Otx and Hox11/13b inputs; by 24 h the expression domain shifts into the veg1 (future hindgut) endoderm and, at gastrulation, forms a ring of cells around the blastopore through which invaginating endoderm cells transit. A second expression domain arises in the oral ectoderm and becomes restricted to the stomodeum, the site of mouth invagination. After the embryonic phase the gene is silenced and is re-expressed in the larval vestibule of the adult rudiment and in the hydrocoel mesoderm. Bra binds genomic cis-regulatory elements and acts mainly as a terminal regulator of the endoderm network: loss of function does not abolish endoderm or mesoderm specification but blocks the morphogenetic movements of gastrulation, and the genes that depend on Bra in the endoderm are effector genes, including cytoskeletal modulators implicated in gut morphogenesis, gut enzymes and the Hedgehog ligand, rather than further regulatory genes. Bra also feeds back positively on the otx-beta transcript of the otx/blimp1b/gatae endoderm kernel, and genome-wide binding studies place it in an ancient Brachyury-FoxA-Wnt feedback loop that predates the cnidarian-bilaterian split.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0000978 RNA polymerase II cis-regulatory region sequence-specific DNA binding | IEA GO_REF:0000002 | ACCEPT | Summary: InterPro2GO mapping from the T-box domain signature. Supported for this gene product by genome-wide Brachyury ChIP-seq in S. purpuratus, which recovered direct target loci including the FoxA/Wnt feedback-loop genes, and by the morpholino/misexpression screen that defined endodermal Brachyury-responsive genes. Reason: Core molecular function; direct genomic binding of S. purpuratus Brachyury has been demonstrated by chromatin immunoprecipitation sequencing. Supporting Evidence: PMID:36396969 we present a genome-wide target-gene screen using chromatin immunoprecipitation sequencing in the sea anemone Nematostella vectensis, an early branching non-bilaterian, and the sea urchin Strongylocentrotus purpuratus PMID:12027442 The latter genes are candidates for direct transcriptional targets of Brachyury. |
| GO:0003677 DNA binding | IEA GO_REF:0000104 | KEEP AS NON CORE | Summary: Generic DNA binding from the UniRule T-box rule. Correct but uninformative; the sequence-specific cis-regulatory binding row already present (GO:0000978) captures the actual activity. Reason: Redundant parent of GO:0000978, which is accepted as the core binding activity. Supporting Evidence: PMID:36396969 Our analysis reveals an ancestral gene regulatory feedback loop connecting Brachyury, FoxA and canonical Wnt signalling involved in axial patterning |
| GO:0003700 DNA-binding transcription factor activity | IEA GO_REF:0000002 | ACCEPT | Summary: InterPro2GO mapping from the T-box and Brachyury signatures. Consistent with the experimental record: Brachyury knockdown or dominant-repressor interference in sea urchin embryos removes expression of a defined set of endodermal target genes, premature misexpression induces them, and the protein binds genomic targets in ChIP-seq. Reason: Core molecular function of the gene product; the RNA polymerase II-specific child term is proposed as NEW. Supporting Evidence: PMID:12027442 the brachyury gene transduces information about the state of endodermal specification to genes that modulate morphogenesis and genes that perform terminal functions in the gut PMID:19895806 perturbation of brachyury expression strongly depressed expression levels of otxβΞ² transcripts |
| GO:0005634 nucleus | IEA GO_REF:0000120 | ACCEPT | Summary: Subcellular-location and InterPro mapping. Expected location of a T-box transcription factor whose chromatin binding has been demonstrated by ChIP-seq in S. purpuratus; an anti-LvBrac serum detects nuclear protein in Lytechinus, but the cached abstract does not state subcellular localization explicitly. Reason: Functionally required location of a sequence-specific transcription factor with demonstrated genomic binding. Supporting Evidence: PMID:36396969 we present a genome-wide target-gene screen using chromatin immunoprecipitation sequencing in the sea anemone Nematostella vectensis, an early branching non-bilaterian, and the sea urchin Strongylocentrotus purpuratus |
| GO:0006355 regulation of DNA-templated transcription | IEA GO_REF:0000002 | KEEP AS NON CORE | Summary: Generic InterPro2GO process term. True, but the positive-regulation row (modified here to the RNA polymerase II-specific term) carries the informative statement. Reason: Generic parent of the positive regulation of transcription annotation; kept for completeness only. Supporting Evidence: PMID:12027442 Here, we identify genes that are expressed downstream of brachyury during gastrulation of the sea urchin embryo. |
| GO:0045893 positive regulation of DNA-templated transcription | IEA GO_REF:0000002 | MODIFY | Summary: InterPro2GO mapping from the T-box DNA-binding domain. The sign is supported in S. purpuratus: brachyury morpholino strongly depresses otx-beta transcripts and removes a set of endodermal effector genes, while premature brachyury misexpression induces target genes; in Lytechinus a Brachyury-Engrailed repressor fusion phenocopies loss of function, as expected for a native activator. Brachyury acts on RNA polymerase II-transcribed genes, so the RNA polymerase II-specific term is the appropriate one. Reason: The essence is right, but the RNA polymerase II-specific child is the correct term for a eukaryotic sequence-specific activator. Proposed replacements: positive regulation of transcription by RNA polymerase II Supporting Evidence: PMID:19895806 perturbation of brachyury expression strongly depressed expression levels of otxβΞ² transcripts PMID:19895806 Otx is activated by its own gene product as well as the products of its target genes blimp1b and brachyury. PMID:12027442 Some of the endodermal genes that respond to Brachyury are cytoskeletal modulators that may play a role in gut morphogenesis. PMID:19393640 At gastrulation the Hh ligand is expressed by the endoderm downstream of the Brachyury and FoxA transcription factors in the endomesoderm gene regulatory network. |
| GO:0000981 DNA-binding transcription factor activity, RNA polymerase II-specific | IDA PMID:36396969 An ancestral Wnt-Brachyury feedback loop in axial patterning... | NEW | Summary: Brachyury is a sequence-specific transcription factor acting on RNA polymerase II-transcribed developmental genes. Genome-wide ChIP-seq in S. purpuratus defined its direct target loci, including the components of a conserved Brachyury-FoxA-Wnt feedback loop, and morpholino knockdown plus premature misexpression in S. purpuratus defined a set of endodermal genes whose transcription depends on Brachyury. The activator child (GO:0001228) is not proposed because no published S. purpuratus reporter or site-mutagenesis assay of a Brachyury target module was found, although all loss- and gain-of-function data point to activation. Reason: Direct chromatin binding (IDA) and target-gene dependence (IMP) in S. purpuratus justify the RNA polymerase II-specific transcription factor term as the core molecular function; mouse T (P20293) carries the same term by IDA. Supporting Evidence: PMID:36396969 we present a genome-wide target-gene screen using chromatin immunoprecipitation sequencing in the sea anemone Nematostella vectensis, an early branching non-bilaterian, and the sea urchin Strongylocentrotus purpuratus PMID:36396969 Our analysis reveals an ancestral gene regulatory feedback loop connecting Brachyury, FoxA and canonical Wnt signalling involved in axial patterning PMID:12027442 the brachyury gene transduces information about the state of endodermal specification to genes that modulate morphogenesis and genes that perform terminal functions in the gut |
| GO:0007369 gastrulation | IMP PMID:12027442 brachyury Target genes in the early sea urchin embryo isolat... | NEW | Summary: Brachyury is expressed in the ring of endoderm cells that invaginate through the blastopore and is required for the movements of gastrulation. In S. purpuratus, interfering with Brachyury function by morpholino blocks gastrulation, and the endodermal genes that depend on Brachyury are cytoskeletal modulators and other effectors of gut morphogenesis rather than regulatory genes; in Lytechinus a dominant-repressor Brachyury-Engrailed fusion blocks gastrulation movements while leaving endoderm and mesoderm specification markers intact. Brachyury therefore performs the regulatory step that turns the endoderm regulatory state into the morphogenetic effector programme of archenteron invagination, which is participation in gastrulation rather than a mere upstream requirement. Reason: Loss-of-function evidence in S. purpuratus (morpholino) corroborated in Lytechinus (dominant negative), with the gene product acting directly on morphogenesis effector genes in the invaginating endoderm. Comparator check: mouse T (P20293) carries GO:0007509 mesoderm migration involved in gastrulation by IMP, so gastrulation-branch process terms are the established convention for Brachyury orthologues. The generic term is chosen because the sea urchin literature describes the gastrulation block as a whole rather than a specific cellular movement. Supporting Evidence: PMID:12027442 Some of the endodermal genes that respond to Brachyury are cytoskeletal modulators that may play a role in gut morphogenesis. This finding is consistent with the block in gastrulation induced by interfering with Brachyury function in sea urchins PMID:11784024 Microinjection of mRNA encoding this LvBrac-EN construct resulted in a block in gastrulation movements but not expression of endoderm and mesoderm marker genes. PMID:11784024 It was then determined that LvBrac is necessary for the morphogenetic movements occurring in both expression regions. PMID:35038441 In sea urchin embryos, Brachyury is expressed in the invaginating endoderm, and in the oral ectoderm of the invaginating mouth opening. |
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Download this section (compressed HTML)Q: Is Brachyury a direct transcriptional activator of its endodermal effector targets (cytoskeletal modulators, hh) and of the otx-beta module, which would justify GO:0001228 and confirm the otx-brachyury positive feedback as a direct linkage in the endoderm kernel?
Q: Does Brachyury perform the same morphogenetic function in the oral ectoderm of S. purpuratus as it does in Lytechinus, that is, is stomodeal invagination and mouth formation dependent on Brachyury, and which effector genes does it control there?
Q: The hindgut (veg1) phase of brachyury expression is driven by Hox11/13b; does Brachyury contribute to hindgut/anus morphogenesis at the pluteus stage, and is the later re-expression in the adult rudiment vestibule and hydrocoel mesoderm functional?
Experiment: Site-directed mutagenesis of T-box (Brachyury) binding sites in the cis-regulatory modules of S. purpuratus endodermal effector genes identified by ChIP-seq and by the macroarray screen, assayed as reporter constructs in Brachyury morpholino and misexpression backgrounds, to establish direct activation.
Experiment: Oral-ectoderm-targeted Brachyury perturbation in S. purpuratus (for example a photoactivatable or late-injected dominant-negative construct) combined with live imaging of stomodeal invagination, to test the mouth-formation function described in Lytechinus.
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