Ets1 (Sp-Ets1/2) is the single sea urchin orthologue of the vertebrate ETS1/ETS2 subfamily of ETS-domain transcription factors, with an N-terminal PNT (pointed/SAM) domain and a C-terminal winged-helix ETS DNA-binding domain. Its transcript is maternally supplied and ubiquitous through cleavage; zygotic transcription is confined to the large micromere lineage, the founder cells of the skeletogenic primary mesenchyme cells (PMCs), because ets1 is one of the direct targets of the Pmar1/HesC double-negative gate, and it later extends to the non-skeletogenic mesoderm. The protein is activated in the micromere lineage by Raf/MEK/ERK phosphorylation of a conserved threonine (T107) that lies in a P-P-T-P MAP-kinase consensus motif. Ets1 is a transcriptional activator that helps lock in the skeletogenic regulatory state (feeding into alx1, tbr and foxN2/3) and is the most widely used positive driver of the skeletogenic differentiation gene batteries, acting directly through ETS sites in cis-regulatory modules of effector genes such as cyclophilin-1 and the spicule matrix gene SM50. Loss of Ets1 function gives skeleton-less embryos whose PMCs fail to ingress, whereas ectopic ets1 mRNA converts non-PMC cells into migratory mesenchyme, so Ets1 is required both for specification of the skeletogenic mesenchyme and for its epithelial-mesenchymal transition. Across echinoderms Ets1/2 is a conserved larval mesoderm regulator whose skeletogenic target set was co-opted in the sea urchin lineage.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0000981 DNA-binding transcription factor activity, RNA polymerase II-specific | IBA GO_REF:0000033 | ACCEPT | Summary: PAINT inference from the ETS family node, seeded by the experimentally characterized vertebrate and fly ETS factors. Fully consistent with the sea urchin record: Ets1 is a sequence-specific ETS-domain factor whose direct positive inputs into skeletogenic differentiation genes (Sp-cyp1) and regulatory genes (tbr) have been shown by cis-regulatory analysis in S. purpuratus. Reason: Core molecular function of the gene product, supported by direct cis-regulatory evidence; the activator child term is proposed as NEW below. Supporting Evidence: PMID:16574094 Ets1 and Dri target sites within the 218 bp fragment are required for normal expression. This indicates that the predicted inputs are direct. PMID:19679118 a dedicated distal module utilizing positive Ets1/2 inputs contributes to definitive expression in the skeletogenic mesenchyme |
| GO:0003677 DNA binding | IEA GO_REF:0000104 | MODIFY | Summary: Generic ETS-domain DNA binding from UniRule. True but uninformative; the demonstrated binding is to ETS sites in RNA polymerase II cis-regulatory modules (the Sp-cyp1 module requires its Ets1 target sites, the SM50 promoter has a functional positive ets site, and Ets1 motifs are enriched in PMC CRMs). Reason: Replace with the cis-regulatory sequence-specific term that describes what the protein actually binds. Note that no EMSA/ChIP with sea urchin Ets1 protein is cached; the inference rests on site-mutagenesis in reporter constructs. Proposed replacements: RNA polymerase II cis-regulatory region sequence-specific DNA binding Supporting Evidence: PMID:16574094 Ets1 and Dri target sites within the 218 bp fragment are required for normal expression PMID:10096062 In the upstream region of the SM50 gene there exists an ets binding site that functions as a positive cis-regulatory element |
| GO:0003700 DNA-binding transcription factor activity | IEA GO_REF:0000002 | ACCEPT | Summary: InterPro2GO mapping from the ETS domain and ETS family signatures. Correct; it is the parent of the RNA polymerase II-specific IBA term above, which is the more informative statement. Reason: Correct general molecular function, subsumed by GO:0000981 and by the activator term proposed as NEW. Supporting Evidence: PMID:16574094 This indicates that the predicted inputs are direct. |
| GO:0005634 nucleus | IBA GO_REF:0000033 | ACCEPT | Summary: PAINT inference of nuclear activity across the ETS family. No immunolocalization of sea urchin Ets1 is cached, but the protein is a transcription factor acting through cis-regulatory target sites, so nuclear activity is not in doubt. Reason: Expected and functionally required location of a transcription factor with demonstrated direct cis-regulatory inputs. Supporting Evidence: PMID:16574094 Ets1 and Dri target sites within the 218 bp fragment are required for normal expression |
| GO:0005634 nucleus | IEA GO_REF:0000120 | ACCEPT | Summary: Rule-based nuclear localization from the ETS domain; consistent with the IBA row and with the transcriptional function. Reason: Correct location; redundant with the IBA row. |
| GO:0006351 DNA-templated transcription | IEA GO_REF:0000104 | MODIFY | Summary: UniRule mapping to the transcription process itself. Ets1 is a sequence-specific transcription factor that regulates transcription of its target genes; it is not part of the RNA polymerase machinery that carries out transcription. GO practice annotates transcription factors to regulation-of-transcription terms. Reason: Replace the over-general process with the demonstrated positive regulation of RNA polymerase II transcription (direct activating inputs into cyp1 and tbr cis-regulatory modules; loss of ets1 depresses target gene expression). Proposed replacements: positive regulation of transcription by RNA polymerase II Supporting Evidence: PMID:16574094 elimination of either Ets1 or Dri inputs severely depresses the activity of expression constructs containing this DNA fragment |
| GO:0006355 regulation of DNA-templated transcription | IEA GO_REF:0000120 | KEEP AS NON CORE | Summary: Generic InterPro/UniRule process term. Correct but redundant with the RNA polymerase II-specific terms present and with the positive-regulation term proposed as NEW. Reason: True by parentage of the demonstrated function; the informative terms are the RNA polymerase II-specific child and the lineage-specification process. |
| GO:0006357 regulation of transcription by RNA polymerase II | IBA GO_REF:0000033 | ACCEPT | Summary: PAINT inference of RNA polymerase II transcriptional regulation. In the sea urchin the demonstrated sign is positive (Ets1 is the most common A-driver of the skeletogenic differentiation batteries and an activator of tbr and alx1), so the sign-neutral parent is accurate but the positive child is proposed as NEW. Reason: Supported by direct cis-regulatory and perturbation evidence in S. purpuratus. Supporting Evidence: PMID:18413610 the ets1 gene ( 9 ) is used in these gene batteries only as the A driver, and it is the most common of these PMID:19679118 both pmar mRNA and hesC MASO injection cause the ectopic expression of ets1/2, an activator of tbr |
| GO:0006357 regulation of transcription by RNA polymerase II | IEA GO_REF:0000002 | ACCEPT | Summary: InterPro2GO duplicate of the IBA row above; same assessment. Reason: Correct, redundant with the IBA annotation. Supporting Evidence: PMID:18413610 the ets1 gene ( 9 ) is used in these gene batteries only as the A driver, and it is the most common of these |
| GO:0030154 cell differentiation | IBA GO_REF:0000033 | MODIFY | Summary: PAINT places a generic cell-differentiation role at the ETS family node, drawing on the many lineage-differentiation roles of vertebrate ETS factors. For the sea urchin gene the role is specific and well characterized: Ets1 is an essential regulator of the skeletogenic mesenchyme (PMC) lineage, whose loss gives skeleton-minus embryos and whose ectopic expression converts non-PMC cells to mesenchyme. Reason: The IBA is not wrong, but the informative term for this gene is the specification of the mesodermal (skeletogenic mesenchyme) cell fate, proposed as the replacement and as a NEW row. Propagation Review Root cause: TERM SCOPING PROBLEM Failure modes: GRANULARITY MISMATCH Sources checked: PANTHER:PTN000218930 SUPPORTS TRANSFER ETS-family node seeded by vertebrate, fly and worm ETS factors with lineage-differentiation roles; the transfer of a differentiation role to Sp-Ets1 is sound, but the family-level term is far less specific than the skeletogenic mesenchyme specification role established experimentally in S. purpuratus. Proposed replacements: mesodermal cell fate specification Supporting Evidence: PMID:16997294 Functional analysis of Sp-Ets1/2 has previously demonstrated an essential role of this gene in the specification of the skeletogenic mesenchyme lineage PMID:18413610 interference with their expression produces a skeleton-minus phenotype, as illustrated in Fig. 3 B–G for tbr , hex , tgif , ets1 , and alx1 |
| GO:0043565 sequence-specific DNA binding | IEA GO_REF:0000002 | ACCEPT | Summary: InterPro2GO mapping from the ETS and pointed domains. Consistent with the requirement for Ets1 target sites in the Sp-cyp1 cis-regulatory module and the enrichment of Ets1 motifs in PMC cis-regulatory modules; the cis-regulatory-region child is proposed as the replacement for the generic DNA binding row. Reason: Sequence-specific binding is the basis of the demonstrated direct cis-regulatory inputs. Supporting Evidence: PMID:29558892 Binding sites for Ets1 and Alx1, two PMC-enriched transcription factors that have direct or indirect inputs into half of the known PMC effector genes in the PMC gene regulatory network [21], were found to be significantly enriched |
| GO:0001228 DNA-binding transcription activator activity, RNA polymerase II-specific | IDA PMID:16574094 cis-Regulatory control of cyclophilin, a member of the ETS-D... | NEW | Summary: Ets1 is a direct transcriptional activator: in S. purpuratus, Ets1 target sites in the 218-bp Sp-cyp1 cis-regulatory module are required for reporter expression and removing the Ets1 input depresses it; the distal tbr module carries positive Ets1/2 inputs; and in Hemicentrotus the SM50 promoter has a positive ets site. Reason: Direct reporter/cis-regulatory evidence for activator function in S. purpuratus; the sign is shown, so the activator child of GO:0000981 is warranted. Supporting Evidence: PMID:16574094 Ets1 and Dri target sites within the 218 bp fragment are required for normal expression. This indicates that the predicted inputs are direct. PMID:19679118 a dedicated distal module utilizing positive Ets1/2 inputs contributes to definitive expression in the skeletogenic mesenchyme PMID:10096062 In the upstream region of the SM50 gene there exists an ets binding site that functions as a positive cis-regulatory element |
| GO:0045944 positive regulation of transcription by RNA polymerase II | IMP PMID:18413610 Global regulatory logic for specification of an embryonic ce... | NEW | Summary: Ets1 positively regulates transcription of the skeletogenic regulatory genes alx1 and tbr (cross-regulatory input, maintenance of alx1) and is the most common positive driver of the skeletogenic differentiation gene batteries; ets1 knockdown abolishes these outputs and ets1/2 mRNA drives tbr ectopically. Reason: Perturbation (MASO, mRNA, dominant-negative) plus cis-regulatory evidence that Ets1 activates Pol II target genes in the micromere lineage. Supporting Evidence: PMID:18413610 This gene is also a cross-regulatory target of the ets1 gene, among the initial double negative gate genes, and ets1 expression may lock itself on by autoactivation PMID:20181745 this gene is regulated by MAPK signaling and by the transcription factor Ets1; however, these inputs influence only the maintenance of alx1 expression and not its activation PMID:19679118 both pmar mRNA and hesC MASO injection cause the ectopic expression of ets1/2, an activator of tbr |
| GO:0007501 mesodermal cell fate specification | IMP PMID:18413610 Global regulatory logic for specification of an embryonic ce... | NEW | Summary: Ets1 is one of the four essential regulators of the skeletogenic micromere lineage activated by the double-negative gate. It performs the regulatory work of specification directly: it feeds forward into alx1 and tbr and drives the differentiation gene batteries. Loss of function gives skeleton-minus embryos in S. purpuratus; gain of function in Hemicentrotus and Paracentrotus converts non-PMC cells into migratory mesenchyme, and a dominant-negative blocks SM50 and spicule formation. Reason: The lineage specified is skeletogenic mesoderm (large micromere/PMC); Ets1 is a direct transcriptional driver of the specifying regulatory state, not merely required upstream. Comparators Alx1 and Tbr in the same network carry or warrant the same term. Supporting Evidence: PMID:18413610 They are four essential regulators of downstream micromere lineage function: alx1 ( 29 ), ets1 ( 9 , 33 ), tbr ( 34 , 35 ), and tel ( 9 ) PMID:16997294 Functional analysis of Sp-Ets1/2 has previously demonstrated an essential role of this gene in the specification of the skeletogenic mesenchyme lineage PMID:10096062 The overexpression of HpEts by mRNA injection into fertilized eggs alters the cell fate of non-PMC to migratory PMC PMID:14973284 Overexpression of ets1 by injection of synthetic mRNA in the egg caused a dramatic increase in the number of cells becoming mesenchymal at the blastula stage |
| GO:0010718 positive regulation of epithelial to mesenchymal transition | IMP PMID:18413610 Global regulatory logic for specification of an embryonic ce... | NEW | Summary: Ets1 target genes are required for ingression (EMT) of the skeletogenic cells: ets1 morpholino blocks PMC ingression in S. purpuratus, ERK-phosphorylated Ets1 is required for the epithelial-to-mesenchymal transition of PMCs in Paracentrotus, and ectopic ets1 mRNA increases the number of cells becoming mesenchymal. Reason: Ets1 is a transcriptional driver of the ingression program (a regulatory role, so the regulation term rather than EMT itself); the same term is proposed for the co-regulator Alx1 in this network. Supporting Evidence: PMID:18413610 both it and alx1 target genes are required for ingression of the skeletogenic cells (ref. 36 and Fig. 3 F and G ) PMID:14973284 the MAP kinase pathway, working through phosphorylation of Ets1, is required for full specification of the PMCs and their subsequent transition from epithelial to mesenchymal state |
| GO:0070169 positive regulation of biomineral tissue development | IMP PMID:18413610 Global regulatory logic for specification of an embryonic ce... | NEW | Summary: Ets1 directly activates skeletogenic differentiation genes (Sp-cyp1 through required Ets1 sites; SM50 through a positive ets site) and is the most common A-driver of the biomineralization gene batteries; ets1 knockdown gives skeleton-minus embryos and dominant-negative Ets blocks SM50 expression and spicule formation. Reason: Direct cis-regulatory input into biomineralization effector genes plus loss-of-function skeleton phenotype; parallels the term proposed for Alx1. Supporting Evidence: PMID:18413610 interference with their expression produces a skeleton-minus phenotype, as illustrated in Fig. 3 B–G for tbr , hex , tgif , ets1 , and alx1 PMID:16574094 Ets1 and Dri target sites within the 218 bp fragment are required for normal expression PMID:10096062 The overexpression of dominant negative delta HpEts which lacks the N terminal domain, in contrast, specifically represses SM50 expression and development of the spicule |
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Download this section (compressed HTML)Q: Is the later Ets1 input into non-skeletogenic mesoderm (where ets1 expression expands after ingression) a distinct regulatory function that would warrant an additional mesoderm-development annotation, or is it covered by mesodermal cell fate specification?
Q: Does ERK phosphorylation of T107 regulate Ets1 DNA binding, its interaction with a coactivator, or its stability, and is the same mechanism operative in S. purpuratus as shown in P. lividus?
Q: Is ets1 auto-activation (proposed from network analysis) a direct cis-regulatory input, which would justify an autoregulation annotation?
Experiment: ChIP-seq (or CUT&RUN) of tagged Sp-Ets1 in PMCs at mesenchyme blastula stage, combined with EMSA on the Sp-cyp1 and tbr distal module ETS sites, to establish direct binding genome-wide and distinguish direct from indirect Ets1 inputs in the PMC GRN.
Experiment: Phospho-site mutant (T107A/T107D) rescue of ets1 morphants in S. purpuratus, scored for alx1/tbr expression and PMC ingression, to confirm that ERK phosphorylation is the switch for Ets1 activity in this species.
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