Imported:YPM/Ste11/Ste50 interactions

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Ste11/Ste50 Interactions

  • Overexpression of Ste50 and Ste50(T42D) are partially able to compensate for a Ste11ΔSAM mutant (mating assay). Overexpression of Ste50(T42A) is not. Wu et al. 2003 PMID 14555477
    • This suggests that phosphorylation of T42 is important for Ste50's ability to suppress the defect caused by the Ste11ΔSAM mutant.
    • Though overexpression of Ste50 can restore mating in Ste11ΔSAM strain, it does not result in sustained cell cycle arrest (by halo assay), or gene expression (FUS1-LacZ reporter).
    • Overexpression of Ste50 cannot compensate for deletion of Ste11. This is interesting because binding between Ste11ΔSAM and Ste50 is not detectable, suggesting that there is still some undetectable level of binding that is important for signal transmission, or that Ste50 is able to activate Ste11 through some means other than binding it.
  • Similar amounts of Ste11-Myc coprecipitated with WT Ste50, Ste50(T42A) and Ste50(T42D), suggesting that phosphorylation of Ste50 at T42 does not affect its binding to Ste11. Wu et al. 2003 PMID 14555477
  • Overexpression of Ste11 is able to partially compensate for the mating defect suffered from deletion of Ste50 (judged by mating efficiency, halo formation, gene expression, and shmoo formation). Ramezani-Rad et al. 1998 PMID 9738877
  • Ste11 amino-terminal domain (residues 1-424) mutants Ste11(S302A S306A T307A) and Ste11(S302D S306D T307D) both interact efficiently with Ste50 by yeast two-hybrid, suggesting that phosphorylation of Ste11 does not modulate its binding to Ste50. van Drogen et al. 2000 PMID 10837245
  • Ste11's SAM domain (residues 26-129) interacts with Ste50's N-terminal 115 residues (by coimmunoprecipitation). Wu et al. 1999 PMID 10397774
  • Ste11's SAM (Sterile Alpha Motif) domain (contained within residues 1-108) interacts with full length Ste50 as well as Ste50's SAM domain (by two-hybrid). Jansen et al. 2001 PMID 11370856
  • Deletion of Ste11 SAM domain (residues 26-130) has same effect as deletion of Ste50. Since Ste50 interacts with Ste11's SAM domain, this suggests that Ste50's contribution to pheromone response requires its interaction with Ste11. Wu et al. 1999 PMID 10397774
  • Deletion of Ste50's SAM domain has similar effects to deletion of Ste50 itself (assaying mating efficiency, transcriptional response, shmoo formation, and halo formation). Jansen et al. 2001 PMID 11370856
    • Overexperession of Ste11-ΔSAM partially compensates for the deletion of the SAM domain in both mating response and filamentous growth, but not in the Sho1 branch of the HOG pathway.
  • The amount of Ste11-myc that is coprecipitated with Ste50 is unchanged by exposure to pheromone and high salt concentrations. Wu et al. 1999 PMID 10397774
  • Gel filtration experiments suggest that the SAM domain from Ste11 (residues 14-86) and the SAM domain from Ste50 (residues 27-108) form a trimer (two Ste11-SAM, one Ste50-SAM). Grimshaw et al. 2004 PMID 14573615
  • Gel shift assays suggest that Ste11 SAM domain (residues 37-104) and GST-tagged Ste50 SAM domain (residues 27-131) form a herotrimer (two Ste11SAM and one Ste50SAM). Bhattacharjya et al. 2004 PMID 15544813
  • Ste11SAM (residues 15-92) interacts with Ste50SAM (residues 14-118) via yeast two-hybrid in a diploid (using Gal4's activating domain and Gal4's DNA-binding domain). Kwan et al. 2004 PMID 15327964
    • GST-Ste11SAM was able to pull down purified His-Ste50SAM, and vice versa.
  • By surface plasmon resonance, the kinetics of Ste50SAM (residues 14-118) binding to His-tagged immobilized Ste11SAM (residues 15-92) were measured to be kon = 0.413 ± 0.011 μM-1s-1, koff = 0.0295 ± 0.002 s-1 (Kd = 71.4 ± 3.1 nM). The kinetics of Ste11SAM binding to His-tagged immobilized Ste50SAM were measured to be kon = 0.389 ± 0.033 μM-1s-1, koff = 0.0232 ± 0.0001 s-1 (Kd = 59.6 ± 3.4 nM). Kwan et al. 2004 PMID 15327964
  • Mutation of Ste50's RA domain does not disrupt Ste50's binding to Ste11. Truckses et al. 2006 PMID 16428446
  • The SAM domains from Ste11 and Ste50 can be swapped for each other. Wu et al. 2006 PMID 16543225
    • Replacing just the Ste11 SAM domain with the Ste50 SAM domain, or just the Ste50 SAM domain with the Ste11 SAM domain prevents cells from growing on high-osmotic media.
    • Swapping Ste11's SAM domain and Ste50's SAM domain with each other restores growth on high-osmotic media.
  • Ste50's function in the HOG pathway appears to be to target Ste11 to the membrane via its RA domain. Wu et al. 2006 16543225
    • Expression of Ste50's RA domain by itself (in ste11Δ ste50Δ ssk2Δ ssk22Δ cells), or expression of Ste11ΔSAM by itself does not permit growth on high-osmotic media.
    • Expression of Ste50's RA domain fused to Ste11ΔSAM (in ste11Δ ste50Δ ssk2Δ ssk22Δ cells) restores growth on high-osmotic media.
    • Expression of Ste50's SAM domain tagged with a myristoylation signal restores growth on high-osmotic media in ste50Δ ssk2Δ ssk22Δ cells.
    • Addition of a myristoylation signal to Ste11 bypasses the need for Ste50 for growth on high-osmotic media. Ste5 was deleted in these experiments to prevent improper activation of mating response and (presumably) cell-cycle arrest.
  • Cell expressing myristoylated Ste11 showed abnormal morphology consistent with partial activation of the pheromone response pathway, whereas cells expressing myristoylated Ste50 displayed normal morphology. Wu et al. 2006 16543225
    • This suggests that localization of Ste11 to the membrane via its interaction with Ste50 is not sufficient for activation of the mating pathway.

Reaction Definition

Since it does not appear that Ste50/Ste11 binding is modulated at all by pheromone exposure, we will ignore the details of Ste50 binding to Ste11, and assume that the effects of this interaction are taken into account by the rates of reaction involving Ste11.