Wnt target gene activation requires ß-catenin separation into biomolecular condensates.

The Wnt/ß-catenin signaling pathway plays numerous essential roles in animal development and tissue/stem cell maintenance. The activation of genes regulated by Wnt/ß-catenin signaling requires the nuclear accumulation of ß-catenin, a transcriptional co-activator. ß-catenin is recruited to many Wnt-regulated enhancers through direct binding to T-cell factor/lymphoid enhancer factor (TCF/LEF) family transcription factors. ß-catenin has previously been reported to form phase-separated biomolecular condensates (BMCs), which was implicated as a component of ß-catenin's mechanism of action. This function required aromatic amino acid residues in the intrinsically disordered regions (IDRs) at the N- and C-termini of the protein. In this report, we further explore a role for ß-catenin BMCs in Wnt target gene regulation. We find that ß-catenin BMCs are miscible with LEF1 BMCs in vitro and in cultured cells. We characterized a panel of ß-catenin mutants with different combinations of aromatic residue mutations in human cell culture and Drosophila melanogaster. Our data support a model in which aromatic residues across both IDRs contribute to BMC formation and signaling activity. Although different Wnt targets have different sensitivities to loss of ß-catenin's aromatic residues, the activation of every target examined was compromised by aromatic substitution. These mutants are not defective in nuclear import or co-immunoprecipitation with several ß-catenin binding partners. In addition, residues in the N-terminal IDR with no previously known role in signaling are clearly required for the activation of various Wnt readouts. Consistent with this, deletion of the N-terminal IDR results in a loss of signaling activity, which can be rescued by the addition of heterologous IDRs enriched in aromatic residues. Overall, our work supports a model in which the ability of ß-catenin to form biomolecular condensates in the nucleus is tightly linked to its function as a transcriptional co-regulator.

PI3K Promotes Basal Cell Carcinoma Growth Through Kinase-Induced p21 Degradation.

Basal cell carcinoma (BCC) is a locally invasive epithelial cancer that is primarily driven by the Hedgehog (HH) pathway. Advanced BCCs are a critical subset of BCCs that frequently acquire resistance to Smoothened (SMO) inhibitors and identifying pathways that bypass SMO could provide alternative treatments for patients with advanced or metastatic BCC. Here, we use a combination of RNA-sequencing analysis of advanced human BCC tumor-normal pairs and immunostaining of human and mouse BCC samples to identify a PI3K pathway expression signature in BCC. Pharmacological inhibition of PI3K activity in BCC cells significantly reduces cell proliferation and HH signaling. However, treatment of Ptch1fl/fl ; Gli1-CreERT2 mouse BCCs with the PI3K inhibitor BKM120 results in a reduction of tumor cell growth with no significant effect on HH signaling. Downstream PI3K components aPKC and Akt1 showed a reduction in active protein, whereas their substrate, cyclin-dependent kinase inhibitor p21, showed a concomitant increase in protein stability. Our results suggest that PI3K promotes BCC tumor growth by kinase-induced p21 degradation without altering HH signaling.