RAF1 (CRAF, c-Raf, Raf-1) — curation notes

UniProt P04049, human. HGNC:9829. RAF-family (TKL) Ser/Thr kinase; paralogs BRAF (P15056), ARAF (P10398).

Identity and architecture

• CR1 = RAS-binding domain (RBD) + cysteine-rich domain (CRD, C1-like); CR2 = Ser/Thr-rich with the inhibitory
  pSer259 14-3-3 site; CR3 = kinase domain followed by the pSer621 14-3-3 site
  [file:human/RAF1/RAF1-deep-research-falcon.md "CR2, a serine/threonine-rich regulatory segment containing the inhibitory"].
• Catalysis: EC 2.7.11.1; UniProt catalytic activity evidence is PMID:17603483
  [file:human/RAF1/RAF1-uniprot.txt "Reaction=L-seryl-[protein] + ATP = O-phospho-L-seryl-[protein] + ADP +"].

Core function: MAP3K of the RAS-RAF-MEK-ERK cascade

• UniProt: "Serine/threonine-protein kinase that acts as a regulatory link between the membrane-associated Ras GTPases
  and the MAPK/ERK cascade" [file:human/RAF1/RAF1-uniprot.txt]. RAF1 phosphorylates MAP2K1/MAP2K2, which then
  phosphorylate ERK1/2.
• MEK1/2 are the only broadly accepted physiological substrates; BRAF has higher intrinsic MEK-kinase activity than CRAF
  [file:human/RAF1/RAF1-deep-research-falcon.md "MEK1/MAP2K1 and MEK2/MAP2K2 are the only broadly accepted physiological RAF1 substrates."].
• Reactome: "Although all three RAF kinases can phosphorylate MAP2K1 and MAP2K2, BRAF appears to be the primary
  activator in vivo" [Reactome:R-HSA-5672978].
• Direct kinase assays: carbachol-activated c-Raf phosphorylates MEK in vitro PMID:8063729; IFN-gamma activates Raf-1
  via JAK1 PMID:9446616; IL-8 activates Raf-1 in neutrophils PMID:8576262.

Activation cycle (regulatory interactions that map to informative binding MFs)

• 14-3-3 (GO:0071889): 14-3-3 dimer binds pSer259 and pSer621; pS621 is the high-affinity site
  PMID:14688280;
  S621/14-3-3 binding is required for activity PMID:19595761;
  Noonan mutations near S259 impair 14-3-3 binding PMID:20679480. Many classic papers (PMID:7935795, 7939632, 8085158,
  7882972, 8601312, 7760835).
• RAS-GTP (GO:0031267 small GTPase binding): RBD binds GTP-Ras PMID:8332187;
  RBD/Rap1A crystal structure PMID:7791872; RBD-CRD/KRAS structure PMID:33608534.
  RAF1 does NOT bind Di-Ras PMID:12194967.
• Dimerization with BRAF (GO:0046982) and homodimers (GO:0042802/0042803): RAF inhibitors prime wild-type RAF
  dimers PMID:20130576; kinase-dead BRAF acts through CRAF PMID:20141835; UniProt "Heterodimerizes with BRAF".
• MEK (GO:0031434): substrate complex [PMID:24746704 title "Disruption of CRAF-mediated MEK activation ..."];
  RKIP displaces MEK PMID:17097642.
• HSP90/CDC37 chaperone client (GO:0051879, GO:0051087)
  [file:human/RAF1/RAF1-deep-research-falcon.md "RAF1 also depends strongly on the HSP90–CDC37 chaperone system"].
• MRAS-SHOC2-PP1C holophosphatase dephosphorylates pS259 [PMID:25137548 "which tethers RAS, RAF-1 and the catalytic subunit of protein phosphatase 1c (PP1c)"; PMID:36175670].
• PP2A B-alpha/delta holoenzymes associate with Raf1 and dephosphorylate S259 PMID:16239230.
• Negative regulators: RKIP/PEBP1 PMID:17097642, PAQR3/RKTG Golgi trapping PMID:17724343,
  HERC2 ubiquitylation PMID:36241744.

Kinase-independent / non-canonical roles (non-core but real)

• Suppression of apoptosis by binding/inhibiting ASK1 and MST2
  PMID:21779496;
  MST2/Hippo switch PMID:24929361.
• Mitochondrial pool: Bcl-2 targets Raf-1 to mitochondria, BAD phosphorylation, protection from apoptosis PMID:8929532;
  RAF phosphorylates BAD S75/S99/S118 PMID:19667065.
  Note: GOA row GO:0006915 "acts_upstream_of_or_within_positive_effect" apoptotic process (TAS PMID:8929532) has the sign
  wrong — the paper shows protection from apoptosis.
• Rok-alpha (ROCK2) inhibition, motility, wound healing, differentiation [PMID:21779496, PMID:15943972].
• Adenylyl cyclase phosphorylation/sensitization (AC II, V, VI) PMID:15385642.
• Nuclear pS621 RAF1 with NFATc3 at CXCR5 promoter in RA-induced differentiation PMID:24330068.
• Rb binding/phosphorylation PMID:21139044.

Disease

• Germline gain-of-function: Noonan syndrome 5 / LEOPARD, hypertrophic cardiomyopathy [PMID:17603483, PMID:20052757].

Curation decisions (summary)

• Core: MAP3K activity (GO:0004709), protein Ser/Thr kinase activity, MAPK cascade, cytosol, plasma membrane.
• 225 bare GO:0005515 rows mapped by WITH/FROM partner type, matching the BRAF review:
  14-3-3 -> GO:0071889; RAS/RAP -> GO:0031267; MEK1/2 -> GO:0031434; BRAF -> GO:0046982; HSP90 -> GO:0051879;
  CDC37/BAG2/HSPA5/CCT3 -> GO:0051087; STUB1/HERC2 -> GO:0031625; KSR1/JSAP1/PEBP4/SHOC2 -> GO:0097110;
  PAK1/PAK2/AKT1/STK3/EGFR -> GO:0019901; PP2A/PP1/CDC25A -> GO:0019902; NFATC3 -> GO:0140297.
  Partners with no informative MF (PEBP1, PIN1, PAQR3, RB1, RCAN1, FAM83 family, metabolic/mitochondrial Y2H/AP-MS
  hits, viral proteins, APP, CASP8, SMAD4, MLF2, Lox, MFHAS1) -> REMOVE (not asserting interaction is false).
• NOT small GTPase binding (PMID:12194967, Di-Ras): the Di-Ras negative result is correct, but negating the general
  term contradicts the defining RAS-effector function -> REMOVE.
• Several IntAct IPI RAS rows cite papers where the cached text does not mention RAF1 (RBD pulldowns used as a Ras-GTP
  reporter are likely, e.g. PMID:19222999, 19063885, 19696784, 20133694, 21457714, 17229891, 19164755). Mapped to
  GO:0031267 with low confidence, deferring to the curator.
• PMID:9765203 cache has no abstract; PubMed abstract describes constitutive MEK signaling-induced senescence and does
  not mention RAF1 -> UNDECIDED.
• PMID:19593445 (known batch miscitation) does not appear in RAF1 GOA.