ERK2 MAP kinase regulates SUFU binding by multisite phosphorylation of GLI1

Crosstalk between the Hedgehog and MAPK signaling pathways occurs in several types of cancer and contributes to clinical resistance to Hedgehog pathway inhibitors. Here we show that MAP kinase-mediated phosphorylation weakens the binding of the GLI1 transcription factor to its negative regulator SUFU. ERK2 phosphorylates GLI1 on three evolutionarily conserved target sites (S102, S116, and S130) located near the high-affinity binding site for SUFU; these phosphorylations cooperate to weaken the affinity of GLI1-SUFU binding by over 25-fold. Phosphorylation of any one, or even any two, of the three sites does not result in the level of SUFU release seen when all three sites are phosphorylated. Tumor-derived mutations in R100 and S105, residues bordering S102, also diminish SUFU binding, collectively defining a novel evolutionarily conserved SUFU affinity-modulating region. In cultured mammalian cells, GLI1 variants containing phosphomimetic substitutions of S102, S116, and S130 displayed an increased ability to drive transcription. We conclude that multisite phosphorylation of GLI1 by ERK2 or other MAP kinases weakens GLI1-SUFU binding, thereby facilitating GLI1 activation and contributing to both physiological and pathological crosstalk.

Automated summary

There is crosstalk between the Hedgehog and MAPK signaling pathways in cancer, leading to resistance to Hedgehog pathway inhibitors. The study demonstrates that MAP kinase-mediated phosphorylation weakens the binding of the GLI1 transcription factor to its negative regulator SUFU. The phosphorylation of GLI1 at three specific target sites decreases the affinity of GLI1-SUFU binding, and this effect is not observed when only one or two sites are phosphorylated.