Figure 7
Fbxo41 regulates sonic hedgehog signaling. (a) Scheme depicting sonic hedgehog cascade, which in mammals concentrates in the cilium3. In the basal state, the Ptch1 receptor is in the cilium, and inhibits Smo targeting to cilium. Upon binding of sonic hedgehog (Shh) to Ptch1, Smo can target to the cilium and induces the conversion of the Gli proteins from a repressed (GliRep) to an activated (GliActv) form. Once activated, Gli proteins target to nucleus and promote transcription of Gli1 and Ptch143. (b–e) RPE1 cells were infected with the indicated constructs for three days and then fixed for quantification of percentage of ciliated cells (b) and cilia length (c), or treated with Shh-conditioned media to study sonic hedgehog cascade activation (d,e). (b) Cells expressing Fbxo41 had significantly less cilia than the cells in other conditions. Data from 2 experimental weeks. (c) Cilia length determined in the same cells as in (b), showed significantly smaller cilia in cells expressing Fbxo41WT but not in cells expressing Fbxo41W577A or Fbxo41 ΔCΔF. (d) Fbxo41 overexpression prevents the increase in Gli1 expression in cells stimulated with Shh-N condition media. Data from 4 or 5 experimental weeks. (e) The ratio of Gli1 mRNA levels between cells stimulated with Shh-N CM and control condition media is significantly lower in cells overexpressing Fbxo41, compared to cells expressing GFP, other Fbxo41 mutants and uninfected cells (same cells as in (d)). (f) Gli1 mRNA level increase in response to SAG treatment is not statistically significant in any condition. (g) Neurons expressing GFP but not those expressing Fbxo41, show increased Ptch1 mRNA levels in response to SAG treatment. Data are represented as mean ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001 ****p < 0.0001 Kruskal-Wallis test followed by a Dunn’s multiple comparison post-hoc test
