Ste5 is the scaffold protein of the Saccharomyces cerevisiae mating-pheromone MAP kinase pathway. It is a non-catalytic 917-residue protein with an N-terminal basic amphipathic plasma-membrane (PM) helix, a RING-H2 zinc-finger domain that binds the G-protein beta subunit Ste4, a Fus3-binding domain (residues 288-316), a cryptic pleckstrin-homology (PH) domain that binds PtdIns(4,5)P2 and the MAPKKK Ste11, and a C-terminal von Willebrand type-A (VWA; "Ste5-ms") domain that binds the MAPKK Ste7. Ste5 binds Ste11, Ste7 and Fus3 and, upon pheromone receptor activation, is recruited by the released Gbeta-gamma (Ste4-Ste18) dimer, together with direct membrane and PI(4,5)P2 binding, to the cytoplasmic face of the plasma membrane at the mating projection tip. Membrane recruitment delivers Ste5-bound Ste11 to the membrane kinase Ste20 and is sufficient to activate the pathway. Ste5 is also an allosteric activator: its VWA domain acts as a co-catalyst that raises the rate of Ste7 phosphorylation of Fus3 about 5000-fold without affecting Ste7 phosphorylation of Kss1, making Fus3 activation specific to the mating pathway, while the Fus3-binding domain stimulates Fus3 autophosphorylation, leading to feedback phosphorylation of Ste5 that dampens pathway output. Ste5 shuttles between nucleus and cytoplasm; G1 Cln/CDK phosphorylation of sites flanking the PM helix blocks its membrane binding, restricting pheromone signaling to G1. In some strain backgrounds Ste5 also supports Kss1 activation during invasive growth.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0001403 invasive growth in response to glucose limitation | IMP PMID:15192700 Differential input by Ste5 scaffold and Msg5 phosphatase rou... | KEEP AS NON CORE | Summary: In S288c (FLO8-corrected), ste5 deletion abolishes agar invasion and Ste5 is needed for Kss1 activation by several invasive-growth stimuli including glucose depletion. Reason: The IMP data support a role of Ste5 in invasive growth in the S288c background, via generating active Ste11 that feeds the Kss1 cascade. This is strain dependent (not required in Sigma1278b) and secondary to the mating-pathway scaffold role, so kept as non-core. Supporting Evidence: PMID:15192700 showed that ste4 Ξ and ste5 Ξ strains were as deficient in invading the agar as flo11 Ξ and ste20 Ξ strains PMID:15192700 Thus, Ste4 and Ste5 are required for Kss1 to be activated by multiple stimuli that induce IG. PMID:15192700 The requirement for Ste4 and Ste5 in activation of Kss1 during IG in S288c contrasted Ξ£1278b, which does not require Ste4 or Ste5 for IG or SVG |
| GO:0001934 positive regulation of protein phosphorylation | IDA PMID:19303851 The Ste5 scaffold directs mating signaling by catalytically ... | ACCEPT | Summary: Reconstitution shows the Ste5 VWA (Ste5-ms) domain increases the kcat of Ste7-catalysed Fus3 phosphorylation about 5000-fold, i.e. Ste5 positively regulates Fus3 phosphorylation. Reason: Directly demonstrated by in vitro kinetics (IDA). This regulatory process is a core aspect of Ste5 function: it is what makes Fus3 activation mating-pathway specific. The corresponding MF is captured by the proposed NEW protein serine/threonine kinase activator activity annotation. Supporting Evidence: PMID:19303851 We identify and structurally characterize a domain in Ste5 that catalytically unlocks Fus3 for phosphorylation by Ste7. PMID:19303851 This domain specifically increases the kcat for the Ste7β Fus3 reaction by ~5000-fold, while it has no effect on the kcat or KM of the Ste7β Kss1 reaction. PMID:19303851 Thus, the Ste5-ms domain is essentially serving as a substrate specific co-catalyst for Ste7βFus3 phosphorylation |
| GO:0005078 MAP kinase scaffold activity | IDA PMID:8062390 Ste5 tethers multiple protein kinases in the MAP kinase casc... | ACCEPT | Summary: Ste5 is the prototypic MAPK scaffold of the pheromone pathway, binding Ste11, Ste7 and Fus3 on distinct domains and the Gbeta subunit Ste4 (IDA, PMID:8062390). Reason: Core molecular function. Ste5 binds all three kinases of the mating MAPK cascade through separable domains, copurifies with them in a large complex, and increases association of Ste11 with Ste7 and Fus3; it also binds the upstream Gbeta Ste4, exactly as described in the term definition (binds multiple kinases of the cascade and upstream signaling proteins). Supporting Evidence: PMID:8062390 By two-hybrid analysis, Ste11, Ste7, and Fus3 associate with different domains of Ste5, while Kss1, another MAPK, associates with the same domain as Fus3, thus implying that Ste5 simultaneously binds a MEKK, MEK, and MAPK. 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. PMID:8062390 Ste5 also increases the amount of Ste11 complexed to Ste7 and Fus3 and is required for Ste11 to function. PMID:7851759 Ste5p, a protein of unknown biochemical function, interacted with protein kinases that operate at each step of the MAP kinase cascade, specifically with Ste11p (an MEKK), Ste7p (an MEK), and Fus3p (a MAP kinase). PMID:7851759 This finding suggests that one role of Ste5p is to serve as a scaffold to facilitate interactions among members of the kinase cascade. |
| GO:0005078 MAP kinase scaffold activity | IPI PMID:7851759 Protein-protein interactions in the yeast pheromone response... | ACCEPT | Summary: Ste5 is the prototypic MAPK scaffold of the pheromone pathway, binding Ste11, Ste7 and Fus3 on distinct domains and the Gbeta subunit Ste4 (IPI, PMID:7851759). Reason: Core molecular function. Ste5 binds all three kinases of the mating MAPK cascade through separable domains, copurifies with them in a large complex, and increases association of Ste11 with Ste7 and Fus3; it also binds the upstream Gbeta Ste4, exactly as described in the term definition (binds multiple kinases of the cascade and upstream signaling proteins). Supporting Evidence: PMID:7851759 Ste5p, a protein of unknown biochemical function, interacted with protein kinases that operate at each step of the MAP kinase cascade, specifically with Ste11p (an MEKK), Ste7p (an MEK), and Fus3p (a MAP kinase). PMID:7851759 This finding suggests that one role of Ste5p is to serve as a scaffold to facilitate interactions among members of the kinase cascade. PMID:8062390 By two-hybrid analysis, Ste11, Ste7, and Fus3 associate with different domains of Ste5, while Kss1, another MAPK, associates with the same domain as Fus3, thus implying that Ste5 simultaneously binds a MEKK, MEK, and MAPK. |
| GO:0005078 MAP kinase scaffold activity | IPI PMID:7851759 Protein-protein interactions in the yeast pheromone response... | ACCEPT | Summary: Ste5 is the prototypic MAPK scaffold of the pheromone pathway, binding Ste11, Ste7 and Fus3 on distinct domains and the Gbeta subunit Ste4 (IPI, PMID:7851759). Reason: Core molecular function. Ste5 binds all three kinases of the mating MAPK cascade through separable domains, copurifies with them in a large complex, and increases association of Ste11 with Ste7 and Fus3; it also binds the upstream Gbeta Ste4, exactly as described in the term definition (binds multiple kinases of the cascade and upstream signaling proteins). Supporting Evidence: PMID:7851759 Ste5p, a protein of unknown biochemical function, interacted with protein kinases that operate at each step of the MAP kinase cascade, specifically with Ste11p (an MEKK), Ste7p (an MEK), and Fus3p (a MAP kinase). PMID:7851759 This finding suggests that one role of Ste5p is to serve as a scaffold to facilitate interactions among members of the kinase cascade. PMID:8062390 By two-hybrid analysis, Ste11, Ste7, and Fus3 associate with different domains of Ste5, while Kss1, another MAPK, associates with the same domain as Fus3, thus implying that Ste5 simultaneously binds a MEKK, MEK, and MAPK. |
| GO:0005078 MAP kinase scaffold activity | IPI PMID:7851759 Protein-protein interactions in the yeast pheromone response... | ACCEPT | Summary: Ste5 is the prototypic MAPK scaffold of the pheromone pathway, binding Ste11, Ste7 and Fus3 on distinct domains and the Gbeta subunit Ste4 (IPI, PMID:7851759). Reason: Core molecular function. Ste5 binds all three kinases of the mating MAPK cascade through separable domains, copurifies with them in a large complex, and increases association of Ste11 with Ste7 and Fus3; it also binds the upstream Gbeta Ste4, exactly as described in the term definition (binds multiple kinases of the cascade and upstream signaling proteins). Supporting Evidence: PMID:7851759 Ste5p, a protein of unknown biochemical function, interacted with protein kinases that operate at each step of the MAP kinase cascade, specifically with Ste11p (an MEKK), Ste7p (an MEK), and Fus3p (a MAP kinase). PMID:7851759 This finding suggests that one role of Ste5p is to serve as a scaffold to facilitate interactions among members of the kinase cascade. PMID:8062390 By two-hybrid analysis, Ste11, Ste7, and Fus3 associate with different domains of Ste5, while Kss1, another MAPK, associates with the same domain as Fus3, thus implying that Ste5 simultaneously binds a MEKK, MEK, and MAPK. |
| GO:0005515 protein binding | IPI PMID:15020407 Fus1p interacts with components of the Hog1p mitogen-activat... | REMOVE | Summary: Two-hybrid matrix detection of Ste5 interacting with the cell-fusion protein Fus1 (Nelson et al. 2004). Reason: Bare protein binding is uninformative. The cached record is abstract-only and does not show which Fus1 region mediated the Ste5 interaction, so a more specific MF (e.g. SH3 domain binding, supported separately by PMID:18280496) cannot be assigned from this reference; the physiological role of a Ste5-Fus1 interaction is also unknown. Removal of the generic term does not mean the reported interaction is false. Supporting Evidence: PMID:15020407 A two-hybrid matrix containing fusion proteins and pheromone response pathway signaling molecules reveals that Fus1p may participate in a complex network of interactions. |
| GO:0005515 protein binding | IPI PMID:15020407 Fus1p interacts with components of the Hog1p mitogen-activat... | MODIFY | Summary: Two-hybrid matrix of pheromone-pathway signaling molecules (Nelson et al. 2004) detecting Ste5 binding to the MAPK Fus3. Reason: Bare protein binding is uninformative. The Ste5-Fus3 interaction is well established (two-hybrid and copurification; PMID:7851759, PMID:8062390) and is mediated by the Ste5 Fus3-binding domain, so mitogen-activated protein kinase binding is the informative MF. Removal of the generic term does not mean the reported interaction is false. Proposed replacements: mitogen-activated protein kinase binding Supporting Evidence: PMID:15020407 A two-hybrid matrix containing fusion proteins and pheromone response pathway signaling molecules reveals that Fus1p may participate in a complex network of interactions. PMID:7851759 Ste5p, a protein of unknown biochemical function, interacted with protein kinases that operate at each step of the MAP kinase cascade, specifically with Ste11p (an MEKK), Ste7p (an MEK), and Fus3p (a MAP kinase). |
| GO:0005515 protein binding | IPI PMID:17289571 A mechanism for cell-cycle regulation of MAP kinase signalin... | MODIFY | Summary: IPI evidence for Ste5 binding Ste4 (G-protein beta subunit) (PMID:17289571). Reason: Ste5 binds the G-protein beta subunit Ste4 via its N-terminal RING-H2 domain, coupling pheromone receptor activation to membrane recruitment of the scaffold. The informative MF is G-protein beta-subunit binding. Bare protein binding is uninformative; removal of the generic term does not mean the reported interaction is false. The cached text is partial; the Ste5-Gbetagamma interaction is well established (PMID:7667635). Proposed replacements: G-protein beta-subunit binding Supporting Evidence: PMID:17289571 Instead, Ste5 also binds directly to membranes, and the cooperative effect of these two weak interactions (Ste5-GΞ²Ξ³ and Ste5-membrane) controls membrane recruitment PMID:7667635 A haploid-specific interaction between the amino terminus of Ste5p and the G protein beta subunit Ste4p was also detected in a two-hybrid assay, and the product of a signaling-defective allele of STE4 was defective in this interaction. |
| GO:0005515 protein binding | IPI PMID:18280496 Recognition of non-canonical peptides by the yeast Fus1p SH3... | MODIFY | Summary: IPI evidence for Ste5 binding Fus1 (PMID:18280496). Reason: A Ste5-derived R(S/T)(S/T)SL peptide binds the non-canonical SH3 domain of Fus1 with low-micromolar affinity in vitro. The informative MF is SH3 domain binding. Bare protein binding is uninformative; removal of the generic term does not mean the reported interaction is false. Note that in vivo relevance of the Ste5-Fus1 interaction is not established, so this is not a core function. Proposed replacements: SH3 domain binding Supporting Evidence: PMID:18280496 Here, we have demonstrated that this domain binds to R(S/T)(S/T)SL-containing peptides derived from two putative in vivo binding partners from yeast proteins, Bnr1p and Ste5p, with K(d) values in the low micromolar range. |
| GO:0005515 protein binding | IPI PMID:19303851 The Ste5 scaffold directs mating signaling by catalytically ... | MODIFY | Summary: IPI evidence for Ste5 binding Ste7 (MAPKK) (PMID:19303851). Reason: Ste5 binds the MAPKK Ste7, most tightly through its C-terminal VWA (Ste5-ms) domain (KD ~75 nM), which then acts as a co-catalyst for Ste7-mediated Fus3 phosphorylation. The informative MF is mitogen-activated protein kinase kinase binding. Bare protein binding is uninformative; removal of the generic term does not mean the reported interaction is false. Proposed replacements: mitogen-activated protein kinase kinase binding Supporting Evidence: PMID:19303851 The Ste5-ms domain binds tightly to Ste7, but only very weakly to Fus3. |
| GO:0005515 protein binding | IPI PMID:19303851 The Ste5 scaffold directs mating signaling by catalytically ... | MODIFY | Summary: IPI evidence for Ste5 binding Fus3 (MAPK) (PMID:19303851). Reason: Ste5 binds the MAPK Fus3 through the Fus3-binding domain (residues 288-316), which allosterically promotes Fus3 autophosphorylation and feedback phosphorylation of Ste5. The informative MF is mitogen-activated protein kinase binding. Bare protein binding is uninformative; removal of the generic term does not mean the reported interaction is false. Proposed replacements: mitogen-activated protein kinase binding Supporting Evidence: PMID:19303851 Second, we show here that the previously identified Fus3 binding site in Ste5 (residues 288β316 in Ste5) is not required to promote the Ste7βFus3 reaction. PMID:19303851 We identify and structurally characterize a domain in Ste5 that catalytically unlocks Fus3 for phosphorylation by Ste7. |
| GO:0005515 protein binding | IPI PMID:23953117 Exploitation of latent allostery enables the evolution of ne... | REMOVE | Summary: In vitro binding of the S. cerevisiae Ste5 Fus3-binding domain (FBD) to a heterologous MAPK, Aspergillus nidulans MpkB, used to probe the evolution of Ste5 allosteric activation of MAPKs. Reason: Bare protein binding is uninformative, and this partner is a MAPK from another fungal species tested in vitro as an evolutionary probe (latent allostery), not a physiological partner of S. cerevisiae Ste5. The informative MF (MAPK binding) is captured by the S. cerevisiae Fus3 rows. Removal does not mean the reported in vitro interaction is false. Supporting Evidence: PMID:23953117 Several kinases were not allosterically affected by the S. cer. Ste5-FBD (D. han. Cek2, C. alb. Cek1, A. nid. MpkB, N. cra. MpkB) even though these kinases readily bound to S. cer. Ste5-FBD PMID:23953117 Surprisingly, both Ste5 activator motifs could regulate MAP kinases that diverged from Fus3 prior to the emergence of Ste5, suggesting that Ste5 activators arose by exploiting latent regulatory features already present in the MAPK ancestor. |
| GO:0005515 protein binding | IPI PMID:23953117 Exploitation of latent allostery enables the evolution of ne... | MODIFY | Summary: IPI evidence for Ste5 binding Fus3 (MAPK) (PMID:23953117). Reason: Ste5 binds the MAPK Fus3 through the Fus3-binding domain (residues 288-316), which allosterically promotes Fus3 autophosphorylation and feedback phosphorylation of Ste5. The informative MF is mitogen-activated protein kinase binding. Bare protein binding is uninformative; removal of the generic term does not mean the reported interaction is false. Proposed replacements: mitogen-activated protein kinase binding Supporting Evidence: PMID:23953117 The second allosteric interaction involves a linear motif in Ste5 called the βFus3 binding domainβ (Ste5-FBD), which binds Fus3 and allosterically activates autophosphorylation of the MAP kinase on its activation loop tyrosine |
| GO:0005515 protein binding | IPI PMID:23953117 Exploitation of latent allostery enables the evolution of ne... | REMOVE | Summary: In vitro binding of the S. cerevisiae Ste5 Fus3-binding domain (FBD) to a heterologous MAPK, Candida albicans Cek1, used to probe the evolution of Ste5 allosteric activation of MAPKs. Reason: Bare protein binding is uninformative, and this partner is a MAPK from another fungal species tested in vitro as an evolutionary probe (latent allostery), not a physiological partner of S. cerevisiae Ste5. The informative MF (MAPK binding) is captured by the S. cerevisiae Fus3 rows. Removal does not mean the reported in vitro interaction is false. Supporting Evidence: PMID:23953117 Several kinases were not allosterically affected by the S. cer. Ste5-FBD (D. han. Cek2, C. alb. Cek1, A. nid. MpkB, N. cra. MpkB) even though these kinases readily bound to S. cer. Ste5-FBD PMID:23953117 Surprisingly, both Ste5 activator motifs could regulate MAP kinases that diverged from Fus3 prior to the emergence of Ste5, suggesting that Ste5 activators arose by exploiting latent regulatory features already present in the MAPK ancestor. |
| GO:0005515 protein binding | IPI PMID:23953117 Exploitation of latent allostery enables the evolution of ne... | REMOVE | Summary: In vitro binding of the S. cerevisiae Ste5 Fus3-binding domain (FBD) to a heterologous MAPK, Kluyveromyces lactis Fus3/Kss1-family MAPK (KLLA0_E10539g), used to probe the evolution of Ste5 allosteric activation of MAPKs. Reason: Bare protein binding is uninformative, and this partner is a MAPK from another fungal species tested in vitro as an evolutionary probe (latent allostery), not a physiological partner of S. cerevisiae Ste5. The informative MF (MAPK binding) is captured by the S. cerevisiae Fus3 rows. Removal does not mean the reported in vitro interaction is false. Supporting Evidence: PMID:23953117 Several kinases were not allosterically affected by the S. cer. Ste5-FBD (D. han. Cek2, C. alb. Cek1, A. nid. MpkB, N. cra. MpkB) even though these kinases readily bound to S. cer. Ste5-FBD PMID:23953117 Surprisingly, both Ste5 activator motifs could regulate MAP kinases that diverged from Fus3 prior to the emergence of Ste5, suggesting that Ste5 activators arose by exploiting latent regulatory features already present in the MAPK ancestor. |
| GO:0005515 protein binding | IPI PMID:23953117 Exploitation of latent allostery enables the evolution of ne... | REMOVE | Summary: In vitro binding of the S. cerevisiae Ste5 Fus3-binding domain (FBD) to a heterologous MAPK, Candida glabrata Fus3, used to probe the evolution of Ste5 allosteric activation of MAPKs. Reason: Bare protein binding is uninformative, and this partner is a MAPK from another fungal species tested in vitro as an evolutionary probe (latent allostery), not a physiological partner of S. cerevisiae Ste5. The informative MF (MAPK binding) is captured by the S. cerevisiae Fus3 rows. Removal does not mean the reported in vitro interaction is false. Supporting Evidence: PMID:23953117 Several kinases were not allosterically affected by the S. cer. Ste5-FBD (D. han. Cek2, C. alb. Cek1, A. nid. MpkB, N. cra. MpkB) even though these kinases readily bound to S. cer. Ste5-FBD PMID:23953117 Surprisingly, both Ste5 activator motifs could regulate MAP kinases that diverged from Fus3 prior to the emergence of Ste5, suggesting that Ste5 activators arose by exploiting latent regulatory features already present in the MAPK ancestor. |
| GO:0005515 protein binding | IPI PMID:7851759 Protein-protein interactions in the yeast pheromone response... | MODIFY | Summary: IPI evidence for Ste5 binding Ste7 (MAPKK) (PMID:7851759). Reason: Ste5 binds the MAPKK Ste7, most tightly through its C-terminal VWA (Ste5-ms) domain (KD ~75 nM), which then acts as a co-catalyst for Ste7-mediated Fus3 phosphorylation. The informative MF is mitogen-activated protein kinase kinase binding. Bare protein binding is uninformative; removal of the generic term does not mean the reported interaction is false. Proposed replacements: mitogen-activated protein kinase kinase binding Supporting Evidence: PMID:7851759 Ste5p, a protein of unknown biochemical function, interacted with protein kinases that operate at each step of the MAP kinase cascade, specifically with Ste11p (an MEKK), Ste7p (an MEK), and Fus3p (a MAP kinase). |
| GO:0005515 protein binding | IPI PMID:7851759 Protein-protein interactions in the yeast pheromone response... | MODIFY | Summary: IPI evidence for Ste5 binding Fus3 (MAPK) (PMID:7851759). Reason: Ste5 binds the MAPK Fus3 through the Fus3-binding domain (residues 288-316), which allosterically promotes Fus3 autophosphorylation and feedback phosphorylation of Ste5. The informative MF is mitogen-activated protein kinase binding. Bare protein binding is uninformative; removal of the generic term does not mean the reported interaction is false. Proposed replacements: mitogen-activated protein kinase binding Supporting Evidence: PMID:7851759 Ste5p, a protein of unknown biochemical function, interacted with protein kinases that operate at each step of the MAP kinase cascade, specifically with Ste11p (an MEKK), Ste7p (an MEK), and Fus3p (a MAP kinase). |
| GO:0005515 protein binding | IPI PMID:7851759 Protein-protein interactions in the yeast pheromone response... | MODIFY | Summary: IPI evidence for Ste5 binding Ste11 (MAPKKK) (PMID:7851759). Reason: Ste5 binds the MAPKKK Ste11 (in part via its PH domain region) and is required for Ste11 function in the mating pathway. The informative MF is mitogen-activated protein kinase kinase kinase binding. Bare protein binding is uninformative; removal of the generic term does not mean the reported interaction is false. Proposed replacements: mitogen-activated protein kinase kinase kinase binding Supporting Evidence: PMID:7851759 Ste5p, a protein of unknown biochemical function, interacted with protein kinases that operate at each step of the MAP kinase cascade, specifically with Ste11p (an MEKK), Ste7p (an MEK), and Fus3p (a MAP kinase). |
| GO:0005515 protein binding | IPI PMID:8062390 Ste5 tethers multiple protein kinases in the MAP kinase casc... | MODIFY | Summary: IPI evidence for Ste5 binding Ste7 (MAPKK) (PMID:8062390). Reason: Ste5 binds the MAPKK Ste7, most tightly through its C-terminal VWA (Ste5-ms) domain (KD ~75 nM), which then acts as a co-catalyst for Ste7-mediated Fus3 phosphorylation. The informative MF is mitogen-activated protein kinase kinase binding. Bare protein binding is uninformative; removal of the generic term does not mean the reported interaction is false. Proposed replacements: mitogen-activated protein kinase kinase binding Supporting Evidence: PMID:8062390 By two-hybrid analysis, Ste11, Ste7, and Fus3 associate with different domains of Ste5, while Kss1, another MAPK, associates with the same domain as Fus3, thus implying that Ste5 simultaneously binds a MEKK, MEK, and MAPK. 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. |
| GO:0005515 protein binding | IPI PMID:8062390 Ste5 tethers multiple protein kinases in the MAP kinase casc... | MODIFY | Summary: IPI evidence for Ste5 binding Fus3 (MAPK) (PMID:8062390). Reason: Ste5 binds the MAPK Fus3 through the Fus3-binding domain (residues 288-316), which allosterically promotes Fus3 autophosphorylation and feedback phosphorylation of Ste5. The informative MF is mitogen-activated protein kinase binding. Bare protein binding is uninformative; removal of the generic term does not mean the reported interaction is false. Proposed replacements: mitogen-activated protein kinase binding Supporting Evidence: PMID:8062390 By two-hybrid analysis, Ste11, Ste7, and Fus3 associate with different domains of Ste5, while Kss1, another MAPK, associates with the same domain as Fus3, thus implying that Ste5 simultaneously binds a MEKK, MEK, and MAPK. 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. |
| GO:0005515 protein binding | IPI PMID:8062390 Ste5 tethers multiple protein kinases in the MAP kinase casc... | MODIFY | Summary: IPI evidence for Ste5 binding Ste11 (MAPKKK) (PMID:8062390). Reason: Ste5 binds the MAPKKK Ste11 (in part via its PH domain region) and is required for Ste11 function in the mating pathway. The informative MF is mitogen-activated protein kinase kinase kinase binding. Bare protein binding is uninformative; removal of the generic term does not mean the reported interaction is false. Proposed replacements: mitogen-activated protein kinase kinase kinase binding Supporting Evidence: PMID:8062390 By two-hybrid analysis, Ste11, Ste7, and Fus3 associate with different domains of Ste5, while Kss1, another MAPK, associates with the same domain as Fus3, thus implying that Ste5 simultaneously binds a MEKK, MEK, and MAPK. 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. |
| GO:0005515 protein binding | IPI PMID:9311911 Ste5 RING-H2 domain: role in Ste4-promoted oligomerization f... | MODIFY | Summary: IPI evidence for Ste5 binding Ste7 (MAPKK) (PMID:9311911). Reason: Ste5 binds the MAPKK Ste7, most tightly through its C-terminal VWA (Ste5-ms) domain (KD ~75 nM), which then acts as a co-catalyst for Ste7-mediated Fus3 phosphorylation. The informative MF is mitogen-activated protein kinase kinase binding. Bare protein binding is uninformative; removal of the generic term does not mean the reported interaction is false. Proposed replacements: mitogen-activated protein kinase kinase binding Supporting Evidence: PMID:9311911 In vitro Ste5(C177A C180A) bound each component of the MAPK cascade, but not Ste4. |
| GO:0005515 protein binding | IPI PMID:9311911 Ste5 RING-H2 domain: role in Ste4-promoted oligomerization f... | MODIFY | Summary: IPI evidence for Ste5 binding Fus3 (MAPK) (PMID:9311911). Reason: Ste5 binds the MAPK Fus3 through the Fus3-binding domain (residues 288-316), which allosterically promotes Fus3 autophosphorylation and feedback phosphorylation of Ste5. The informative MF is mitogen-activated protein kinase binding. Bare protein binding is uninformative; removal of the generic term does not mean the reported interaction is false. Proposed replacements: mitogen-activated protein kinase binding Supporting Evidence: PMID:9311911 In vitro Ste5(C177A C180A) bound each component of the MAPK cascade, but not Ste4. |
| GO:0005515 protein binding | IPI PMID:9311911 Ste5 RING-H2 domain: role in Ste4-promoted oligomerization f... | MODIFY | Summary: IPI evidence for Ste5 binding Ste4 (G-protein beta subunit) (PMID:9311911). Reason: Ste5 binds the G-protein beta subunit Ste4 via its N-terminal RING-H2 domain, coupling pheromone receptor activation to membrane recruitment of the scaffold. The informative MF is G-protein beta-subunit binding. Bare protein binding is uninformative; removal of the generic term does not mean the reported interaction is false. Proposed replacements: G-protein beta-subunit binding Supporting Evidence: PMID:9311911 In vitro Ste5(C177A C180A) bound each component of the MAPK cascade, but not Ste4. PMID:9311911 Thus, the RING-H2 domain mediates Ste4-Ste5 interaction, which is a prerequisite for Ste5-Ste5 self-association and signaling. |
| GO:0005515 protein binding | IPI PMID:9311911 Ste5 RING-H2 domain: role in Ste4-promoted oligomerization f... | MODIFY | Summary: IPI evidence for Ste5 binding Ste11 (MAPKKK) (PMID:9311911). Reason: Ste5 binds the MAPKKK Ste11 (in part via its PH domain region) and is required for Ste11 function in the mating pathway. The informative MF is mitogen-activated protein kinase kinase kinase binding. Bare protein binding is uninformative; removal of the generic term does not mean the reported interaction is false. Proposed replacements: mitogen-activated protein kinase kinase kinase binding Supporting Evidence: PMID:9311911 In vitro Ste5(C177A C180A) bound each component of the MAPK cascade, but not Ste4. |
| GO:0005515 protein binding | IPI PMID:9428767 Interaction of a G-protein beta-subunit with a conserved seq... | MODIFY | Summary: IPI evidence for Ste5 binding Ste4 (G-protein beta subunit) (PMID:9428767). Reason: Ste5 binds the G-protein beta subunit Ste4 via its N-terminal RING-H2 domain, coupling pheromone receptor activation to membrane recruitment of the scaffold. The informative MF is G-protein beta-subunit binding. Bare protein binding is uninformative; removal of the generic term does not mean the reported interaction is false. The cached abstract of PMID:9428767 focuses on Gbeta binding to Ste20; the curator (full text) recorded a Ste5-Ste4 interaction, consistent with the well-established RING-H2-mediated binding (PMID:7667635, PMID:9311911), so the interaction is accepted and re-typed. Proposed replacements: G-protein beta-subunit binding Supporting Evidence: PMID:9428767 In the yeast Saccharomyces cerevisiae, Ste20 is involved in transmitting the mating-pheromone signal from the betagamma-subunits (encoded by the STE4 and STE18 genes, respectively) of a heterotrimeric G protein to a downstream MAP kinase cascade. PMID:7667635 A haploid-specific interaction between the amino terminus of Ste5p and the G protein beta subunit Ste4p was also detected in a two-hybrid assay, and the product of a signaling-defective allele of STE4 was defective in this interaction. PMID:9311911 Thus, the RING-H2 domain mediates Ste4-Ste5 interaction, which is a prerequisite for Ste5-Ste5 self-association and signaling. |
| GO:0005546 phosphatidylinositol-4,5-bisphosphate binding | IDA PMID:16847350 Function of the MAPK scaffold protein, Ste5, requires a cryp... | ACCEPT | Summary: The isolated Ste5 PH domain (residues 388-518) binds PtdIns(4,5)P2 preferentially in overlay, liposome flotation and SPR assays; R407S/K411S abolishes binding. Reason: Direct in vitro evidence with a specificity-defining mutant that also abolishes membrane recruitment and function in vivo. PI(4,5)P2 binding is part of the mechanism that anchors the scaffold at the plasma membrane. Supporting Evidence: PMID:16847350 In vitro the Ste5 PH domain binds preferentially to PtdIns(4,5)P 2 . PMID:16847350 Indeed, in multiple trials, the R407S K411S mutations decreased PtdIns(4,5)P 2 binding of the Ste5 PH domain to the background level PMID:16847350 Thus, the PH domain is essential for stable membrane recruitment of Ste5, and this association is critical for initiation of downstream signaling because it allows Ste5-bound Ste11 (MAPKKK) to be activated by membrane-bound Ste20 (MAPKKKK). |
| GO:0005634 nucleus | IDA PMID:9732267 Membrane recruitment of the kinase cascade scaffold protein ... | KEEP AS NON CORE | Summary: GFP-Ste5 is enriched in nuclei of untreated cells; Ste5 shuttles between nucleus and cytoplasm. Reason: Nuclear localization is directly observed, but membrane-targeted Ste5 that is depleted from the nucleus still signals, so the nucleus is not where Ste5 performs its scaffold function. Kept as a genuine but non-core location. Supporting Evidence: PMID:9732267 In untreated cells, GFPβSte5 was present diffusely throughout the cytoplasm, and was enriched in the nuclei PMID:9732267 suggesting that nuclear localization of Ste5 may not be required for signaling PMID:11781566 Here we show that Ste11p and Ste7p were predominantly cytoplasmic proteins, while Ste5p and Fus3p were found in the nucleus and the cytoplasm. |
| GO:0005737 cytoplasm | IDA PMID:11781566 MAP kinase dynamics in response to pheromones in budding yea... | ACCEPT | Summary: Ste5 localizes to the cytoplasm and nucleus in vegetative cells (van Drogen et al. 2001). Reason: Directly observed; the cytoplasmic pool is the reservoir that is recruited to the plasma membrane by Gbetagamma on pheromone stimulation, and cytoplasmic Ste5-kinase complexes have been quantified by FCCS. Supporting Evidence: PMID:11781566 Here we show that Ste11p and Ste7p were predominantly cytoplasmic proteins, while Ste5p and Fus3p were found in the nucleus and the cytoplasm. PMID:17952059 We quantified the abundance of complexes in the cytoplasm among the MAPKs Ste11, Ste7, Fus3 and the scaffold protein Ste5 in yeast pheromone signalling using fluorescence cross-correlation spectroscopy (FCCS). |
| GO:0005737 cytoplasm | IEA GO_REF:0000044 | ACCEPT | Summary: UniProt subcellular location mapping (Cytoplasm). Reason: Consistent with experimental localization (PMID:11781566, PMID:9732267). Broad but correct. Supporting Evidence: file:yeast/STE5/STE5-uniprot.txt SUBCELLULAR LOCATION: Cytoplasm. PMID:11781566 Here we show that Ste11p and Ste7p were predominantly cytoplasmic proteins, while Ste5p and Fus3p were found in the nucleus and the cytoplasm. |
| GO:0005886 plasma membrane | IMP PMID:16847350 Function of the MAPK scaffold protein, Ste5, requires a cryp... | ACCEPT | Summary: The PH-domain mutant Ste5(R407S K411S) fails to be recruited to the plasma membrane in response to pheromone, and artificial membrane tethering restores signaling. Reason: The plasma membrane (cytoplasmic face, mating projection tip) is where Ste5 performs its core function: Gbetagamma- and PI(4,5)P2-dependent recruitment brings Ste5-bound Ste11 to Ste20. Supported by multiple independent studies. Supporting Evidence: PMID:16847350 Ste5(R407S K411S) is expressed normally, retains Gbetagamma and Ste11 binding, and oligomerizes, yet is not recruited to the membrane in response to pheromone. PMID:16847350 Thus, the PH domain is essential for stable membrane recruitment of Ste5, and this association is critical for initiation of downstream signaling because it allows Ste5-bound Ste11 (MAPKKK) to be activated by membrane-bound Ste20 (MAPKKKK). PMID:9732267 Furthermore, this event has functional significance, as artificial targeting of Ste5 to the plasma membrane, but not intracellular membranes, activates the pathway in the absence of pheromone or Gbetagamma. PMID:11781566 Ste5p remains stably bound at the plasma membrane, unlike activated Fus3p, which dissociates from Ste5p and translocates to the nucleus. |
| GO:0019900 kinase binding | IEA GO_REF:0000002 | MODIFY | Summary: InterPro2GO mapping from IPR021651 (Ste5_Fus-binding) to kinase binding. Reason: Correct but very general. The IPR021651 region corresponds to the Fus3-binding region of Ste5, and experimental work shows it binds the MAPK Fus3, so mitogen-activated protein kinase binding is the more informative term. (Ste5 also binds the MAPKK Ste7 and MAPKKK Ste11, captured by the replacement terms on the protein-binding rows.) Proposed replacements: mitogen-activated protein kinase binding Supporting Evidence: PMID:23953117 The second allosteric interaction involves a linear motif in Ste5 called the βFus3 binding domainβ (Ste5-FBD), which binds Fus3 and allosterically activates autophosphorylation of the MAP kinase on its activation loop tyrosine PMID:19303851 Second, we show here that the previously identified Fus3 binding site in Ste5 (residues 288β316 in Ste5) is not required to promote the Ste7βFus3 reaction. |
| GO:0031681 G-protein beta-subunit binding | IPI PMID:7667635 Association of the yeast pheromone response G protein beta g... | ACCEPT | Summary: Two-hybrid and co-immunoprecipitation evidence that the Ste5 N-terminus binds the Gbeta subunit Ste4 (Whiteway et al. 1995). Reason: Core binding activity: the RING-H2 domain-mediated Ste4 interaction couples pheromone receptor activation to membrane recruitment of the scaffold and is required for signaling. It is part of the MAP kinase scaffold function (binding upstream signaling proteins). Supporting Evidence: PMID:7667635 A haploid-specific interaction between the amino terminus of Ste5p and the G protein beta subunit Ste4p was also detected in a two-hybrid assay, and the product of a signaling-defective allele of STE4 was defective in this interaction. PMID:7667635 In cells with a constitutively activated pheromone response pathway, epitope-tagged Ste4p was coimmunoprecipitated with Ste5p. PMID:9311911 Thus, the RING-H2 domain mediates Ste4-Ste5 interaction, which is a prerequisite for Ste5-Ste5 self-association and signaling. |
| GO:0043332 mating projection tip | IDA PMID:11781566 MAP kinase dynamics in response to pheromones in budding yea... | ACCEPT | Summary: Ste5, Ste7 and Fus3 localize to tips of mating projections in pheromone-treated cells. Reason: Directly observed and reproduced (PMID:9732267); the shmoo tip is the site of Gbetagamma-dependent Ste5 recruitment and cascade activation. Supporting Evidence: PMID:11781566 Ste5p, Ste7p and Fus3p also localized to tips of mating projections in pheromone-treated cells. PMID:9732267 Pheromone caused some of the GFPβSte5 to accumulate at the cell surface, at the tips of pheromone-induced projections |
| GO:0043332 mating projection tip | IDA PMID:17952059 Spatial regulation of Fus3 MAP kinase activity through a rea... | ACCEPT | Summary: FCCS/FLIM study of Fus3 activity gradients emanating from the mating projection tip, where Fus3-activating kinases on the Ste5 scaffold are partitioned. Reason: The cached abstract describes partitioning of Fus3-activating kinases to the projection tip; the curator (full text) recorded Ste5 localization there, which agrees with independent studies (PMID:11781566, PMID:9732267). Deferring to the curator. Supporting Evidence: PMID:17952059 Spatial partitioning of Fus3 activating kinases to this site and deactivating phosphatases in the cytoplasm maintain this Fus3(PP)-activity distribution. PMID:11781566 Ste5p, Ste7p and Fus3p also localized to tips of mating projections in pheromone-treated cells. |
| GO:0043409 negative regulation of MAPK cascade | IMP PMID:16424299 The Ste5 scaffold allosterically modulates signaling output ... | KEEP AS NON CORE | Summary: The Ste5 Fus3-binding domain allosterically promotes Fus3 autophosphorylation; this partially active Fus3 phosphorylates Ste5, reducing transcriptional output (negative feedback). Mutation of the FBD increases output. Reason: The IMP evidence supports a Ste5-mediated negative-feedback role that tunes mating pathway output. This is genuine but a modulatory, secondary role relative to the positive scaffold function, so kept as non-core. Supporting Evidence: PMID:16424299 A fragment of Ste5 allosterically activated autophosphorylation of the mitogen-activated protein kinase Fus3. PMID:16424299 Unexpectedly, at a systems level, autoactivated Fus3 appears to have a negative regulatory role, promoting Ste5 phosphorylation and a decrease in pathway transcriptional output. PMID:23953117 The second allosteric interaction involves a linear motif in Ste5 called the βFus3 binding domainβ (Ste5-FBD), which binds Fus3 and allosterically activates autophosphorylation of the MAP kinase on its activation loop tyrosine |
| GO:0070867 mating projection tip membrane | IDA PMID:9732267 Membrane recruitment of the kinase cascade scaffold protein ... | ACCEPT | Summary: Pheromone or activated Gbetagamma recruits GFP-Ste5 to the cell surface at the tips of mating projections. Reason: Directly observed; Ste5 is a peripheral protein on the cytoplasmic face of the projection-tip plasma membrane, held by Gbetagamma, PI(4,5)P2 binding and its PM amphipathic helix. This is the functional site of the scaffold. Supporting Evidence: PMID:9732267 Pheromone caused some of the GFPβSte5 to accumulate at the cell surface, at the tips of pheromone-induced projections PMID:9732267 We also show that pheromone stimulates translocation of the kinase cascade scaffold protein Ste5 to the cell surface. PMID:9732267 Furthermore, this event has functional significance, as artificial targeting of Ste5 to the plasma membrane, but not intracellular membranes, activates the pathway in the absence of pheromone or Gbetagamma. |
| GO:0071507 pheromone response MAPK cascade | IDA PMID:8062390 Ste5 tethers multiple protein kinases in the MAP kinase casc... | ACCEPT | Summary: Ste5 forms a multikinase complex with Ste11, Ste7 and Fus3 required for efficient Fus3 activation in the pheromone response MAPK cascade. Reason: Core process. Ste5 does work in the cascade: it assembles the kinases, delivers Ste11 to membrane Ste20, and catalytically unlocks Fus3 for Ste7 phosphorylation. ste5 mutants block pheromone signaling. Supporting Evidence: 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. PMID:8062390 Ste5 also increases the amount of Ste11 complexed to Ste7 and Fus3 and is required for Ste11 to function. PMID:9732267 Furthermore, this event has functional significance, as artificial targeting of Ste5 to the plasma membrane, but not intracellular membranes, activates the pathway in the absence of pheromone or Gbetagamma. PMID:19303851 This domain specifically increases the kcat for the Ste7β Fus3 reaction by ~5000-fold, while it has no effect on the kcat or KM of the Ste7β Kss1 reaction. file:yeast/STE5/STE5-uniprot.txt Component of the pheromone signal transduction pathway. It mediates pheromone signals acting between STE20 and STE11. file:yeast/STE5/STE5-deep-research-falcon.md Its primary function is to couple pheromone-activated GΞ²Ξ³ at the plasma membrane to the Ste11βSte7βFus3 kinase cascade |
| GO:0043539 protein serine/threonine kinase activator activity | IDA PMID:19303851 The Ste5 scaffold directs mating signaling by catalytically ... | NEW | Summary: Ste5 VWA (Ste5-ms) domain binds the MAPKK Ste7 and increases the kcat of Ste7-catalysed Fus3 phosphorylation ~5000-fold (a cyclin-like co-catalyst); separately the Ste5 FBD allosterically activates Fus3 autophosphorylation. Reason: This is a molecular activity of Ste5 itself, beyond passive tethering: purified Ste5-ms binds Ste7 (KD ~75 nM) and specifically raises the catalytic rate of Ste7 toward Fus3 (not Kss1), and the FBD fragment allosterically activates Fus3 autophosphorylation. Both fit the definition (binds to and increases the activity of a protein serine/threonine kinase). The existing GO:0001934 positive regulation of protein phosphorylation (IDA, same paper) captures the process consequence but not the MF. Participation test satisfied: Ste5 itself performs the activating step. Supporting Evidence: PMID:19303851 This domain specifically increases the kcat for the Ste7β Fus3 reaction by ~5000-fold, while it has no effect on the kcat or KM of the Ste7β Kss1 reaction. PMID:19303851 Thus, the Ste5-ms domain is essentially serving as a substrate specific co-catalyst for Ste7βFus3 phosphorylation PMID:19303851 The Ste5-ms domain binds tightly to Ste7, but only very weakly to Fus3. PMID:16424299 A fragment of Ste5 allosterically activated autophosphorylation of the mitogen-activated protein kinase Fus3. PMID:23953117 The second allosteric interaction involves a linear motif in Ste5 called the βFus3 binding domainβ (Ste5-FBD), which binds Fus3 and allosterically activates autophosphorylation of the MAP kinase on its activation loop tyrosine |
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Download this section (compressed HTML)Q: Should the Ste5 VWA-domain co-catalyst activity (raising Ste7 kcat toward Fus3) be captured as protein serine/threonine kinase activator activity, or does GO need a more specific term for a substrate-specific kinase co-catalyst/"substrate unlocking" function distinct from general kinase activation?
Q: Is the Ste5 R(S/T)(S/T)SL motif bound by the Fus1 SH3 domain used in vivo during cell fusion, or is the Ste5-Fus1 interaction an in vitro/two-hybrid observation without physiological role?
Experiment: Test Ste5 alleles that separately disrupt the VWA catalytic interface (retaining Ste7 binding) and the Fus1-binding R(S/T)(S/T)SL motif for mating efficiency and cell fusion, to separate scaffold, co-catalyst and potential fusion roles in vivo.
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Download this section (compressed HTML)Pheromone-responsive MAPK scaffold; couples activation of the G-protein-coupled pheromone receptor to MAPK activation; intramolecular interaction of PH and VWA domains blocks activation of the assembled signaling cascade (Ste11p, Ste7p and Fus3p) under basal conditions; GΞ²-Ξ³ (Ste4p-Ste18p)-dependent docking at the plasma membrane and PI(4,5)P2 binding by the PH domain relieves autoinhibition, activating the pathway; interacts with GΞ± (Gpa1p) to enhance the efficiency of polarity site alignment
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