Abstract
Tumor formation is generally linked to the acquisition of two or more driver genes that cause normal cells to progress from proliferation to abnormal expansion and malignancy. In order to understand genetic alterations involved in this process, we compared the transcriptomes of an isogenic set of breast epithelial cell lines that are non-transformed or contain a single or double knock-in (DKI) of PIK3CA (H1047R) or KRAS (G12V). Gene set enrichment analysis revealed that DKI cells were enriched over single mutant cells for genes that characterize a MYC target gene signature. This gene signature was mediated in part by the bromodomain-containing protein 9 (BRD9) that was found in the SWI-SNF chromatin-remodeling complex, bound to the MYC super-enhancer locus. Small molecule inhibition of BRD9 reduced MYC transcript levels. Critically, only DKI cells had the capacity for anchorage-independent growth in semi-solid medium, and CRISPR-Cas9 manipulations showed that PIK3CA and BRD9 expression were essential for this phenotype. In contrast, KRAS was necessary for DKI cell migration, and BRD9 overexpression induced the growth of KRAS single mutant cells in semi-solid medium. These results provide new insight into the earliest transforming events driven by oncoprotein cooperation and suggest BRD9 is an important mediator of mutant PIK3CA/KRAS-driven oncogenic transformation.
Highlights
The tumorigenic potential of an oncogene is defined by its cooperation with other driver mutations that result in complex changes to a cell’s organization [1,2]
Only double knock-in (DKI) cells had the capacity for anchorage-independent growth in semi-solid medium, and CRISPR-Cas9 manipulations showed that PIK3CA and bromodomain-containing protein 9 (BRD9) expression were essential for this phenotype
In order to understand the effects of oncogenic PIK3CA in the context of a second driver mutation, we employed a set of MCF-10A cell lines that differ by a single amino acid substitution in phosphatidylinositol 3-kinase (PI3K)
Summary
The tumorigenic potential of an oncogene is defined by its cooperation with other driver mutations that result in complex changes to a cell’s organization [1,2]. PIK3CA, the gene encoding p110α, the catalytic subunit of phosphatidylinositol 3-kinase (PI3K), is one of the most frequently mutated genes in human cancer [3]. Activating mutations in PIK3CA can co-occur with other driver mutations [6]. An example of this is the presence of both mutant PIK3CA and mutant KRAS (GTPase-encoding oncogene, first identified in the Kirsten rat-sarcoma virus). Activation of either protein alone results in increased signaling through both the PI3K and mitogen-activated protein kinase (MAPK) pathways [7]. Data from human tumor sequencing efforts have shown that KRAS and PIK3CA mutations can co-occur, but their
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