BIK1 (BOTRYTIS-INDUCED KINASE 1, AT2G39660, UniProt O48814) — research notes

Research journal for the GO annotation review of Arabidopsis thaliana BIK1. Provenance recorded inline as [PMID:xxxx "verbatim quote"].

Identity and architecture

• BIK1 is a receptor-like cytoplasmic kinase (RLCK, subfamily VII) — a genuine, catalytically active Ser/Thr (dual-specificity) protein kinase. UniProt: "RecName: Full=Serine/threonine-protein kinase BIK1"; EC=2.7.11.1; 395 AA; Protein kinase domain 67-356; ATP-binding Lys-105; active-site (proton acceptor) Asp-202.
• Lipid-anchored at the plasma membrane: N-myristoyl Gly-2 and S-palmitoyl Cys-4. Gly-2 mutations mislocalize the protein [PMID:26021844 myristoylation at Gly-2; G->S "Drastic reduction of plasma membrane localization and strong increase of cytoplasmic localization."].
• Dual-specificity kinase: PMID:24104392. Catalytic activity (EC 2.7.11.1) experimentally established [PMID:24104392, PMID:32846426].

Core function: immune signaling hub downstream of cell-surface PRRs

• Central RLCK that relays PTI signaling from PRR–BAK1 complexes: PMID:20018686, and PMID:20018686.
• Trans-phosphorylation order: PMID:20018686.
• BIK1 integrates signaling from multiple immune receptors and is targeted by a Pseudomonas effector (title PMID:20413097). Phosphorylated at Ser-236 by FLS2-complex.
• UniProt FUNCTION: "Plays a central role in immune responses ... Involved in pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI) signaling, including calcium signaling, and defense responses downstream of FLS2."

Core function: phosphorylation of RBOHD and ROS regulation

• BIK1 directly phosphorylates the NADPH oxidase RBOHD to drive the ROS burst: PMID:24629339, and PMID:24629339.
• Specific RBOHD sites: PMID:29649442.

Core function: phosphorylation of Ca2+ channel OSCA1.3 / stomatal immunity

• PMID:32846426, and PMID:32846426.
• Phosphorylation depends on kinase activity: PMID:32846426.
• BIK1/PBL1 required for MAMP/DAMP-induced calcium elevations: PMID:25522736, and PMID:25522736.

Defense response / hormone regulation

• Original identification: required for resistance to necrotrophs, negative regulator of basal resistance to Pst, modulates SA: PMID:16339855, and PMID:16339855. The IEP "response to fungus" annotation derives from: PMID:16339855.
• JA/SA regulation via nuclear WRKY phosphorylation: PMID:29649442, and PMID:29649442.

Subcellular localization assessment

• Plasma membrane is the principal, well-supported location (lipid anchor; multiple focused studies): UniProt SUBCELLULAR LOCATION "Cell membrane ... Lipid-anchor". Supported by PMID:16339855, PMID:26021844, PMID:29649442, PMID:32404997, and the OSCA paper EXP annotation PMID:32846426.
• Nucleus is supported by a focused study with a mechanism (activation-dependent relocation): PMID:29649442; UniProt Note "Linked to the plasma membrane when inactivated, but moves to the nucleus upon pathogen-mediated activation by phosphorylation."
• Endosome / endosome membrane / endomembrane supported by ligand-induced internalization study [PMID:32404997 monoubiquitination and internalization "into endocytic compartments"]. UniProt "Endosome membrane".
• Nucleolus (GO:0005730), cytoplasm (GO:0005737, HDA), Golgi (GO:0005794, HDA), mitochondrion (GO:0005739, ISM) are from generic high-throughput / predictive sources, NOT focused BIK1 studies:
• Nucleolus + cytoplasm + nucleus HDA all come from a high-throughput GFP-ORF transient-expression localization screen: PMID:15610358. This GFP-ORF fusion overexpression assay bypasses native N-myristoylation/palmitoylation membrane targeting and is unreliable for an endogenously membrane-anchored kinase. Nucleolar localization has never been substantiated by any focused BIK1 study.
• Golgi HDA is from a global membrane-protein correlation-profiling proteomics survey: PMID:28887381 — not a localization claim specific to BIK1 biology.
• Mitochondrion is ISM (sequence-model prediction, GO_REF:0000122 "AtSubP analysis") with no experimental support and is inconsistent with the myristoyl/palmitoyl PM anchor.

Developmental / growth-defense tradeoff (non-core)

• bik1 has pleiotropic growth phenotypes: PMID:16339855. UniProt DISRUPTION PHENOTYPE: "Altered growth traits, early flowering, weak stems, small siliques and reduced fertility."
• Opposing role with ERECTA in development; BIK1 phosphorylates ER: PMID:31803215. (Basis for one of the protein-binding IPI annotations.)
• Negative regulator of brassinosteroid signaling via BRI1 interaction [PMID:23818580 title]; ethylene signaling via EIN3 destabilization PMID:26021844.

Protein-binding (IPI) annotations

BIK1 has many GO:0005515 "protein binding" IPI annotations citing interaction studies. The specific, informative relationships captured by these papers are: FLS2/BAK1 (PMID:20018686, PMID:20404519, PMID:20413097), PEPR1 (PMID:23431184), BRI1 (PMID:23818580), RBOHD (PMID:24629339), CPK28 (PMID:25525792), PP2C38 (PMID:27494702), EFR + WRKY TFs (PMID:29649442), ERECTA (PMID:31803215), the AvrAC/XopAC complex (DOI:10.1038/s41586-020-2210-3 references). Bare "protein binding" is uninformative and should not be treated as a core molecular function; the relevant kinase-substrate / receptor-association relationships are captured in the MF kinase term and the BP signaling terms. Per project guidelines, bare protein binding is demoted (MARK_AS_OVER_ANNOTATED / KEEP_AS_NON_CORE), not removed (these are real IPI experimental annotations).

Summary of curation decisions

• Kinase activity terms (GO:0004672, GO:0004674, GO:0106310, GO:0016301) and ATP binding (GO:0005524): ACCEPT (active kinase, experimentally proven; specific Ser/Thr kinase activity is core).
• protein phosphorylation (GO:0006468), protein autophosphorylation (GO:0046777): ACCEPT/KEEP — autophosphorylation is a real PTM but mechanistic detail; protein phosphorylation (of RBOHD/OSCA1.3/FLS2/BAK1/WRKY/ER) is core.
• PTI/immune signaling BP terms (GO:0002221, GO:0002237, GO:0042742, GO:0050832, GO:1900424): ACCEPT (core immune function).
• Stomatal movement (GO:0010119): ACCEPT/KEEP (real, OSCA1.3-RBOHD pathway).
• Hormone regulation (GO:0080141 JA, GO:0080142 SA), response to fungus (GO:0009620): KEEP_AS_NON_CORE (downstream/regulatory, supported but not the primary biochemical function).
• PM (GO:0005886), endosome/endomembrane (GO:0010008, GO:0005768, GO:0012505), nucleus (GO:0005634): ACCEPT/KEEP (supported by focused studies).
• Nucleolus (GO:0005730): MARK_AS_OVER_ANNOTATED — only from a generic GFP-ORF overexpression screen, never substantiated; biologically implausible for a membrane-anchored kinase.
• Cytoplasm (GO:0005737), Golgi (GO:0005794): KEEP_AS_NON_CORE / MARK_AS_OVER_ANNOTATED — generic high-throughput, weakly supported.
• Mitochondrion (GO:0005739, ISM): REMOVE — pure sequence-model prediction (AtSubP), contradicted by PM lipid-anchor and all experimental localization data.
• protein binding (GO:0005515, IPI x many): KEEP_AS_NON_CORE / MARK_AS_OVER_ANNOTATED — real interactions but uninformative as bare MF; specifics captured elsewhere.