Project description:p53 is a pivotal tumor suppressor and a major barrier against cancer. We now report that silencing of the Hippo pathway tumor suppressors LATS1 and LATS2 in non-transformed mammary epithelial cells reduces p53 phosphorylation and increases its association with the p52 NF-?B subunit. Moreover, it partly shifts p53âs conformation and transcriptional output towards a state resembling cancer-associated p53 mutants, and endow p53 with the ability to promote cell migration. Notably, LATS1 and LATS2 are frequently downregulated in breast cancer; we propose that such downregulation might benefit cancer by converting p53 from a tumor suppressor into a tumor facilitator. MCF10A cells transfected with siRNA against LATS1/2 alone, p53 alone or LATS1/2 and p53 together. Two independent MCF10A batches provided biological replicates
Project description:We performed ChIP-seq analysis of p53-null HCT116 cells stably expressed wild-type p53 or mutant p53 followed by ATO treatment (PANDAs) to identify p53 binding characteristics by wild-type p53 and PANDAs.
Project description:Linking clinical multi-omics analyses with mechanistic studies may improve the understanding of rare cancers. We leveraged two precision oncology programs to investigate rhabdomyosarcoma with FUS/EWSR1-TFCP2 fusions, an orphan malignancy without effective therapies. All tumors exhibited outlier ALK expression, partly accompanied by intragenic deletions and aberrant splicing. This resulted in the expression of ALK variants, i.e. short transcripts (ST), which we named ALK-ST1 (consisting of exons 1-2:18–29), ALK-ST2 (1:18–29), ALK-ST3 (18–29), and ALK-ST4 (1–5:12–17). To systematically investigate the oncogenic capacity of these ALK variants, we stably expressed them in p53-deficient MCF10A human mammary epithelial cells and performed different transformation assays. These experiments demonstrated that ALK-ST1, ALK-ST2, and ALK-ST3 are oncogenic variants, while ALK-ST4 could not transform MCF10A cells. We confirmed protein expression of ALK-ST1, ALK-ST2, and ALK-ST3 by western blotting, which was not possible for ALK-ST4 due to lack of a specific antibody that binds to the N-terminus of ALK that is lost in ALK-ST4. We therefore performed mass spectrometry-based label-free quantitative proteomics on lysates from MCF10A cells stably expressing empty vector (EV), wildtype ALK (ALK-WT), or ALK-ST4 to confirm its expression.