BUB1B (BubR1, UniProt O60566) - curation notes

Session 2026-09-25 - full review of GOA annotations

Identity and architecture

• Human BUB1B encodes BubR1 (hBUBR1, MAD3L, SSK1), the Mad3-related paralog of BUB1. Taylor et al. noted
  at the outset that it is "perhaps not an additional member of the Bub1 family, but more likely a Mad3-related
  protein" and that its kinase domain "lacks several of the residues that are usually highly conserved among
  most protein kinases" PMID:9660858.
• Domain map (UniProt + literature): N-terminal TPR domain (KNL1/Blinkin binding, KEN1 box at residue 26,
  ABBA motifs, D-box), GLEBS/Bub3-binding motif, KEN2, KARD (LxxIxE PP2A-B56 motif; S670 CDK1, S676 PLK1
  sites), C-terminal kinase-like domain (766-1050).
  PMID:22331848

Core function 1 - MCC subunit / APC/C-CDC20 inhibitor (GO:1990948, GO:0033597, GO:0007094)

• hBUBR1 is essential for the mitotic checkpoint: antibody microinjection abrogates nocodazole arrest
  PMID:10477750.
• The MCC was purified from HeLa cells as hBUBR1/hBUB3/CDC20/MAD2 in near-equal stoichiometry and inhibits
  APC/C ubiquitin ligase activity; the inhibitor co-fractionates with hBUBR1 at every step
  PMID:11535616.
  A preformed MCC exists in interphase
  PMID:11535616.
• BubR1 inhibits APC/C-Cdc20 independently of Mad2 and of its kinase activity
  PMID:11702782.
• Mechanism (cryo-EM, APC/C-MCC): BubR1 degron-like motifs block Cdc20 degron-recognition sites and the
  BubR1 TPR obstructs UbcH10
  PMID:27509861;
  MCC forms when C-Mad2-Cdc20 binds the BubR1-Bub3 dimer
  PMID:27509861.
• KEN1 (K26EN) is essential for the core MCC; D-box/KEN2 let the MCC inhibit a second, APC/C-bound CDC20
  PMID:25383541.
• Acetylation at K250 by PCAF switches BubR1 from APC/C inhibitor (pseudosubstrate) to APC/C-Cdc20 substrate
  PMID:19407811.
• Checkpoint silencing: p31comet drives ATP-dependent MCC disassembly, dissociating Cdc20 from BubR1
  PMID:21300909.

Core function 2 - kinetochore recruitment and KARD-dependent PP2A-B56 recruitment

• Kinetochore localization requires Bub3 binding through the GLEBS segment
  PMID:9660858;
  BubR1 assembles onto kinetochores in prophase, after CENP-F and before CENP-E
  PMID:9763420.
• Recruitment is Bub1- and Bub3-dependent and needed both for SAC arrest and stable K-MT attachment
  PMID:20220147.
  TPR-KI2 (Knl1) contacts are dispensable for recruitment
  PMID:22331848.
• BubR1 sits in the outer kinetochore (Spindly colocalizes with BubR1 adjacent to CREST)
  PMID:19468067.
• PP2A-B56 recruitment via the KARD (not in the local publication cache; from deep research):
  [file:human/BUB1B/BUB1B-deep-research-falcon.md "BUBR1 therefore acts as a targeting platform that positions PP2A-B56 where kinetochore phosphosignalling must be reversed."]
  [file:human/BUB1B/BUB1B-deep-research-falcon.md "Mutation of BUBR1 residues required for B56 binding disrupts chromosome congression; Aurora B inhibition can partially reverse this phenotype, supporting the proposed kinase–phosphatase balance."]
  Primary papers to cache for a future pass: Suijkerbuijk et al. 2012 Dev Cell; Kruse et al. 2013 J Cell Sci;
  Xu et al. 2013 Biol Open; Wang et al. 2016 Protein Cell; Braga et al. 2020 Cell Rep.
  GOA carries no annotation for this function; captured in core_functions with GO:0140483 (kinetochore adaptor
  activity) and flagged in suggested_questions rather than asserted as NEW.

The kinase question (GO:0004672 / GO:0004674 / GO:0106310 / EC 2.7.11.1)

• Historical evidence: autophosphorylation in GST pull-downs, with the caveat of co-purifying kinases
  PMID:9660858;
  immunoprecipitate kinase activity stimulated by nocodazole
  PMID:10477750;
  CENP-E stimulates the activity
  PMID:12925705.
• Pseudokinase consensus (deep research; Suijkerbuijk 2012, Braga 2020):
  [file:human/BUB1B/BUB1B-deep-research-falcon.md "Phylogenomic, structural and direct biochemical analyses found no convincing intrinsic phosphotransfer activity."]
  [file:human/BUB1B/BUB1B-deep-research-falcon.md "Deleting or mutating the region reduced kinetochore PP2A-B56, delayed checkpoint silencing and caused chromosome-alignment defects."]
• Contrary report: Huang et al. 2019 claim human BubR1 is an active kinase phosphorylating CENP-E S2639
  PMID:31201382,
  while conceding the active-kinase signature is not conserved
  PMID:31201382
  and acknowledging the controversy
  PMID:31201382.
• Decision: all seven kinase rows (IBA, IDA x2, NAS, TAS, EC-IEA, Rhea-IEA) graded MARK_AS_OVER_ANNOTATED, consistently.
  Not REMOVE because a direct experimental claim exists and has not been formally refuted; not ACCEPT because
  the activity is disputed, dispensable for the core function, and the field consensus is pseudokinase.
  PMID:31201382 flagged DISPUTED in reference_review. The IBA is annotated with a propagation_review
  (PROPAGATION_BAD / PSEUDO_OR_SUBACTIVITY_LOSS + WRONG_ORTHOLOG_OR_PARALOG): node PTN000361607 predates the
  Bub1/BubR1 split and kinase activity is genuine on the BUB1 branch.

Localization rows

• Cytoplasm (interphase) is the dominant pool
  PMID:9763420;
  spindle/midzone in late anaphase
  PMID:9763420.
• Nucleus (IBA, UniProt keyword): KEEP_AS_NON_CORE - phylogenetic (closed-mitosis yeasts) and inferred for human.
• Centrosome (UniProt keyword from PubMed:19503101, not cached): KEEP_AS_NON_CORE.
• Perinuclear (PMID:20531406, B-cell interactome co-localization with MCM3): KEEP_AS_NON_CORE
  PMID:20531406.
• Cytosol: HPA IDA and 24 Reactome TAS rows all ACCEPT (same location claim); the 11 rows derived from
  cohesin/separase/astral-MT-capture/EML4-NUDC reactions are noted as peripheral pathway-membership rows.
• Anaphase-promoting complex (TAS, PMID:10477750): MODIFY -> GO:0033597. BubR1 binds APC/C-CDC20 as an
  inhibitor within the MCC; it is not an APC/C subunit
  PMID:10477750.

Protein binding (GO:0005515) - 55 IPI rows

Policy: MODIFY where the cited paper supports an informative MF or complex; REMOVE (uninformative, interaction
not disputed) for high-throughput datasets, PTM-enzyme partners (BubR1 as substrate) and recruitment partners
without an MF; UNDECIDED where the cached abstract gives no handle.

• CDC20 (19 rows): mechanistic papers (PMID:11030144, 15525512, 19407811, 20212161, 21300909, 21407176,
  22000412, 24581499, 25383541) -> MODIFY to GO:1990948; HT/pharmacology (PMID:20360068, 25241761, 25502805,
  25852190, 31515488, 32707033, 33961781, 35271311, 37926298, 40205054) -> REMOVE.
• MAD2L1 (8) and BUB3 (11): MCC-context papers -> MODIFY to GO:0033597 mitotic checkpoint complex; HT -> REMOVE.
• BUB1 (4, all HT) -> REMOVE. KNL1 (4) -> REMOVE (recruitment interaction; CC rows carry it).
• CENPE (PMID:9763420), PLK1 (PMID:16760428) -> REMOVE (no settled MF for BubR1).
• KAT2B, UBC, CREBBP, SIRT2 -> REMOVE (BubR1 is the substrate). YWHAE (14-3-3 interactome) -> REMOVE.
• RIPK3 (PMID:29883609, abstract-only, no BubR1 mention) -> UNDECIDED.
• PMID:15525512 and PMID:16760428 abstracts foreground Bub1/Plk1-Bub1; no mis-attribution asserted (curator had
  the full text); reference_review notes this.

Meiotic centromeric cohesion (GO:0051754, IBA)

• Sources: fly BubR1 and pombe bub1 at PTN000361607. KEEP_AS_NON_CORE with propagation_review
  (NO_FAILURE_NON_CORE / CONTEXT_OR_TISSUE_MISMATCH): the shugoshin-recruiting kinase function is BUB1's in
  human; BubR1-PP2A-B56 does participate in cohesion protection, and mouse oocyte studies (Touati et al. 2015,
  not cached) support a BubR1 requirement in meiosis. Raised in suggested_questions.

Disease

• MVA1: biallelic BUB1B variants; R727C/L844F destabilise the protein and abolish its interactions
  PMID:25502805.
• Ageing: BubR1 levels decline with age via K668 acetylation balance (CBP vs SIRT2), mouse
  PMID:24825348.
  Not annotated as a GO process for BUB1B; treated as regulation of BubR1 abundance, not a BubR1 function.

Repository context

• modules/metaphase_anaphase_transition_and_mitotic_exit.yaml models BubR1/Mad3 as the "pseudokinase/KEN-box
  subunit that blocks Cdc20 substrate-binding sites" in the MCC - consistent with this review.
• gocams/67369e7600002505 (pombe) types the mad3-containing MCC activity as GO:0140678 molecular function
  inhibitor activity, occurring in the kinetochore, part of GO:0007094; the human review uses the more specific
  GO:1990948 ubiquitin ligase inhibitor activity, consistent with the MAD2L1 review.

Validation

• just validate human BUB1B: valid, 3 warnings (core-function terms GO:0140483, GO:0007080, GO:0051315 not
  present in existing_annotations - deliberate, see above). All supporting_text snippets checked verbatim.