CoBra (Capsaspora owczarzaki Brachyury) - curation notes

Automated deep research was unavailable for this gene (no provider keys), so no
-deep-research-*.md file exists. These notes are compiled from the cached full
texts of PMID:24043797 and PMID:27114036, the UniProt record and our own locus
assignment analysis.

Identity and the locus assignment caveat

• UniProt A0A0D2VUC6, ORF CAOG_005512, automatic name "TBX19 protein", 1130 aa,
  one T-box domain at 447-625 (PROSITE PS50252), InterPro IPR002070 (TF_Brachyury)
  [file:CAPO3/CoBra/CoBra-uniprot.txt].
• PMID:24043797 names CoBra but gives no locus ID. The assignment of CoBra to
  A0A0D2VUC6 is our own inference
  [file:projects/ORIGINS_OF_MULTICELLULARITY/capsaspora-tbox/RESULTS.md "A0A0D2VUC6 (CAOG_005512) is CoBra."]:
• best T-subfamily match (T-box domain scores TBX19 534.5, TBXT 504.5, well above
  the other two Capsaspora T-box proteins);
• an Arg at the position aligned to Xenopus Bra K149 (context LKLTNRPNTKG), which
  is the residue the paper reports for CoBra
  PMID:24043797.
• Caveat: similarity plus one marker residue, not a phylogeny. The gene models in
  UniProt may differ from those used in the paper.
• PANTHER places the protein in PTHR11267:SF181 "OPTOMOTOR-BLIND PROTEIN" (a
  Tbx2-class subfamily name). This disagrees with the Brachyury identity from the
  paper's phylogeny and our domain scoring. None of the GOA rows come from the
  subfamily; the TreeGrafter rows come from PTN000137774, the deep T-box family node
  (the same node that gives human TBXT its IBA cell fate specification row, with
  fly, mouse, worm and zebrafish donors).
• Side note: the paper calls its second tested gene "CoTbx3" but places it in a
  new Tbx7 class PMID:24043797; RESULTS.md matches A0A0D2WSA5 to
  the Tbx2/3 class. Not relevant to CoBra's rows.

Evidence on the protein

Phylogeny (PMID:24043797)

• Brachyury is the oldest T-box class; filasterean Bra genes cluster at the base of
  the class and keep most DNA-binding and dimerization residues
  PMID:24043797.
• T-box genes are absent in choanoflagellates
  PMID:24043797.

DNA binding in vitro (PMID:24043797, universal PBM)

• PMID:24043797
• Motif conserved across T-box classes; specificity between classes attributed to
  cofactors PMID:24043797.

Heterologous activity in Xenopus (PMID:24043797) - NOT Capsaspora evidence

• Partial rescue of XBra_En dominant-negative embryos (MyoD in situ, qRT-PCR), with
  the authors' own caveat PMID:24043797.
• Overexpression activates all mesendodermal genes tested, unlike metazoan Bra
  PMID:24043797.
• XBra/CoBra chimeras map the difference to the N- and C-terminal regions
  PMID:24043797.
• "pan-Tbox" behaviour PMID:24043797.
• Capsaspora lacks Smad1, the cofactor implicated in restricting metazoan Bra.

In vivo chromatin occupancy in Capsaspora (PMID:27114036)

• ~900 Bra motif instances in ATAC-defined regulatory sites, mostly first intron and
  5' UTR, associated with filopodial amoeba and aggregative stages and with H3K4me3
  and H3K27ac PMID:27114036.
• Anti-CoBra antibody ChIP-qPCR at 20 sites vs 10 random motif sites
  PMID:27114036.
• Inferred target network: cell polarity, phagocytosis, metabolism, TFs, GPCR
  signalling; 63 orthologs shared with mouse Bra targets, enriched in actin
  cytoskeleton and amoeboid motility PMID:27114036.
• Caveat: the antibody was raised against "CoBra" as defined by that group; which
  UniProt entry it corresponds to is again our inference. Target genes are inferred
  (motif + ATAC), only 20 sites validated by ChIP-qPCR; no knockout of CoBra exists.

Decisions summary

• MF rows (DNA-binding TF activity, Pol II cis-regulatory DNA binding): ACCEPT.
• Chromatin, nucleus: ACCEPT (ChIP shows chromatin occupancy in Capsaspora).
• Transcription regulation process rows: ACCEPT; positive regulation supported by
  Xenopus activation and by association of Bra sites with active marks.
• Cell fate specification (TreeGrafter from deep T-box node): REMOVE. Embryonic
  fate specification by animal T-box factors; no evidence that CoBra designates a
  cell fate in Capsaspora (which has temporal life stages, not developmental
  lineages), and the Xenopus rescue is heterologous and nonspecific. Propagation
  review: PROPAGATION_BAD / LINEAGE_OR_TAXON_MISMATCH (not a formal taxon-constraint
  violation: GO:0001708 is only_in_taxon cellular organisms, QuickGO 2026-10-01).
• NEW: GO:0043565 sequence-specific DNA binding, IDA, PMID:24043797 (in vitro PBM).
  The Capsaspora ChIP-qPCR (PMID:27114036) would support IDA GO:0000978, but that term
  is already in GOA (IEA), so it is recorded in the review of that row.