STE5 (YDR103W, UniProt P32917) curation notes

Identity confirmed from STE5-uniprot.txt: AC P32917; D6VS89, GN Name=STE5; Synonyms=NUL3; OrderedLocusNames=YDR103W, 917 aa, S. cerevisiae S288c. Domains (InterPro): RING_Ste5 (IPR063871), PH_STE5 (IPR063666), Ste5_Fus-binding (IPR021651), Ste5_Fus3-bd_dom (IPR021106), Ste5_vWA-ext (IPR062768), Ste5_C_sf (IPR038382). PDB 2KGN/2L4U (PM helix, aa 44-67), 3FZE/4F2H (VWA "Ste5-ms" domain, aa 583-787). No GO-CAM model in gocams/index.tsv contains Ste5. Context module: modules/scer_mating_fus3_cascade.yaml.

Core function: pheromone-pathway MAPK scaffold

• Two-hybrid: Ste5 binds all three cascade kinases PMID:7851759
• Tethering / copurification [PMID:8062390 "Ste5 copurifies with Ste11, Fus3, and a hypophosphorylated form of Ste7, and all four proteins cosediment in a glycerol gradient as if in a large complex."; "Ste5 also increases the amount of Ste11 complexed to Ste7 and Fus3 and is required for Ste11 to function."]
• RING-H2 domain binds Ste4 (Gbeta); required for oligomerization and function PMID:9311911
• Gbeta binding first shown PMID:7667635.

Membrane recruitment

• Pheromone/Gbetagamma recruits Ste5 to the cell surface at projection tips; artificial PM targeting activates pathway PMID:9732267; localization PMID:9732267.
• Nuclear pool: PMID:9732267; nuclear localization probably not required for signaling PMID:9732267.
• PH domain (aa 388-518) binds PI(4,5)P2 and is essential for PM recruitment [PMID:16847350 "In vitro the Ste5 PH domain binds preferentially to PtdIns(4,5)P 2 ."; "the PH domain is essential for stable membrane recruitment of Ste5"]. PM recruitment lets Ste5-bound Ste11 be activated by Ste20.
• Cln/CDK phosphorylation flanking the PM motif blocks membrane binding, restricting signaling to G1 PMID:17289571.
• Cytoplasm/nucleus + projection tips [PMID:11781566 "Ste5p and Fus3p were found in the nucleus and the cytoplasm"; "Ste5p, Ste7p and Fus3p also localized to tips of mating projections in pheromone-treated cells."].

Active (allosteric) roles beyond tethering

• VWA/"Ste5-ms" domain binds Ste7 tightly (KD 75 nM) and raises kcat of Ste7->Fus3 ~5000-fold, not Ste7->Kss1 [PMID:19303851 "This domain specifically increases the kcat for the Ste7➔ Fus3 reaction by ~5000-fold, while it has no effect on the kcat or KM of the Ste7➔ Kss1 reaction."; "Thus, the Ste5-ms domain is essentially serving as a substrate specific co-catalyst for Ste7➔Fus3 phosphorylation"]. Analogy drawn to cyclin as Cdk co-catalyst. This is the basis for proposing GO:0043539 protein serine/threonine kinase activator activity (NEW, IDA).
• Fus3-binding domain (FBD, aa 288-316) allosterically activates Fus3 autophosphorylation; the resulting partially active Fus3 phosphorylates Ste5 and dampens output [PMID:16424299 "A fragment of Ste5 allosterically activated autophosphorylation of the mitogen-activated protein kinase Fus3."; "autoactivated Fus3 appears to have a negative regulatory role, promoting Ste5 phosphorylation and a decrease in pathway transcriptional output."]. FBD not required for signaling PMID:19303851.
• Evolution: VWA activator is ancient (Fus3/Kss1 split); FBD recent; Ste5-FBD also binds heterologous MAPKs in vitro PMID:23953117. These xeno IPI rows are evolutionary probes, not physiological partners.

Kss1 / invasive growth

• In S288c, Ste4 and Ste5 are needed for Kss1 activation and invasive growth; not in Sigma1278b [PMID:15192700 "Thus, Ste4 and Ste5 are required for Kss1 to be activated by multiple stimuli that induce IG."; "The requirement for Ste4 and Ste5 in activation of Kss1 during IG in S288c contrasted Σ1278b, which does not require Ste4 or Ste5 for IG or SVG"]. Treat as non-core, strain-dependent.

Fus1 SH3 interaction

• Ste5 contains an R(S/T)(S/T)SL motif bound by the non-canonical Fus1 SH3 domain in vitro PMID:18280496. Physiological role unknown.

Curation decisions summary

• GO:0005078 MAP kinase scaffold activity: core MF (ACCEPT, 4 rows).
• 20 bare protein-binding rows: MODIFY to MAPKKK binding (Ste11), MAPKK binding (Ste7), MAPK binding (Fus3), G-protein beta-subunit binding (Ste4), SH3 domain binding (Fus1, 18280496 only); REMOVE xeno MAPK rows (23953117) and the Fus1 two-hybrid row from 15020407 (abstract-only, no domain mapping visible).
• NEW: GO:0043539 protein serine/threonine kinase activator activity (Ste5-ms co-catalyst for Ste7->Fus3; FBD activation of Fus3 autophosphorylation).
• Nucleus, invasive growth, negative regulation of MAPK cascade: KEEP_AS_NON_CORE.