Aberrant activation of the Wnt/β-catenin signalling pathway is considered as a key driver in the initiation and progression of colorectal cancer (CRC). MLL1 (encoded by KMT2A) is a histone methyltransferase, and has been reported as an oncogenic regulator in human acute leukemia. However, it is not clear how MLL1 regulates the Wnt/β-catenin pathway to promote CRC development. Based on a previous study in our lab, MLL1 can drive expression of the Wnt target genes, but independent of its histone modification functions. Therefore, the aim of my project is to explore the molecular mechanisms of MLL1 involved in the β-catenin-regulated oncogenic Wnt pathway.
Affinity-purification followed by mass spectrometry (AP-MS) coupled with cellular fractionation was performed using β-catenin as a bait protein in CRC cells in the presence and absence of MLL1. T cell factor 1 (TCF1 encoded by TCF7) and lymphoid enhancer factor 1 (LEF1 encoded by LEF1) were identified as common regulators in both conditions, thus suggesting MLL1-regulated β-catenin activities are TCF-dependent. T cell factor 4 (TCF4 encoded by TCF7L2) only interacted with β-catenin upon MLL1 stimulation, revealing TCF4 could refine MLL1-dependent β-catenin activities to induce the expression of specific Wnt target genes. Additionally, western blots were conducted for either whole cell lysates or fractionated extracts. MLL1 deficiency in CRC cells did not significantly alter total β-catenin expression, but rather decreased expression of c-MYC, a well-known Wnt target gene. Furthermore, MLL1 loss significantly reduced β-catenin chromatin localization but not cytoplasmic or nucleoplasmic accumulation.
MLL1-dependent regulation may not be required for β-catenin expression, but appears to be required for its subcellular localization to activate the oncogenic Wnt/β-catenin signalling pathway. This evidence indicates that MLL1 may work as a scaffolding protein to recruit other partners, directing the binding of β-catenin to targeted genomic loci.