Gene expression profiling of preneoplastic K8-creERT2/PIK3CAH1047R mammary subsets
ABSTRACT: This study examined the effect of mutant PIK3CAH1047R expression in mammary subsets of preneoplastic mammary glands from K8-creERT2/PIK3CA H1047R mice Mammary cell subpopulations were isolated from K8-creERT2/PIK3CAH1047R and K8-creERT2 control animals 4 weeks after activation of PIK3CA H1047R transgene expression by Tamoxifen injection. Pooled mammary glands of 2-3 estrus-synchronized mice per genotype were sorted in 3 independent sortings and used for microarray analysis (20 samples in total).
The adult mouse mammary epithelium contains self-sustained cell lineages that form the inner luminal and outer basal cell layers, with stem and progenitor cells contributing to its proliferative and regenerative potential. A key issue in breast cancer biology is the effect of genomic lesions in specific mammary cell lineages on tumour heterogeneity and progression. The impact of transforming events on fate conversion in cancer cells of origin and thus their contribution to tumour heterogeneity r ...[more]
Project description:This study examined the effect of mutant PIK3CAH1047R expression in mammary subsets of preneoplastic mammary glands from Lgr5-creERT2/PIK3CA H1047R mice Mammary cell subpopulations were isolated from Lgr5-creERT2/PIK3CA H1047R and Lgr5-creERT2 control animals 4 weeks after activation of PIK3CA H1047R transgene expression by Tamoxifen injection. Pooled mammary glands of 2-3 estrus-synchronized mice per genotype were sorted in 3 independent sortings and used for microarray analysis (24 samples in total).
Project description:This study examined the gene expression profile of mammary tumors derived from Lgr5- and K8-positive cell-of-origins Mammary tumors and reference mammary glands from control mice were collected, cryo-homogenized, total RNA was isolated and analyzed by microarray (total 25 samples).
Project description:Mammary epithelium is hierarchically organized, with multipotent basal mammary stem cells (MaSCs) producing both luminal and basal cells during development or upon transplantation. Recent studies suggested that most breast cancers might originate from luminal cells, and oncogenic events, such as ectopic expression of PIK3CAH1047R, could induce multipotency in committed luminal cells. p53 is the most commonly mutated gene in human breast cancer; in particular, its inactivating mutations are found in most triple-negative breast cancers, raising a question as to whether p53-loss plays a key role in acquisition of MaSC-like properties by luminal cells. By in situ lineage tracing, we found that induced loss of p53 in Keratin 8 (K8)+ luminal cells led to their clonal expansion, in part due to increased proliferation, but did not directly affect their luminal identity. Expansion of luminal cells, in particular oestrogen receptor-positive luminal cells, was observed 3-4 weeks after induced p53-loss, which was accompanied by increased expression of cell cycle genes and downregulation of genes related to immune microenvironment, p53 downstream pathway and apoptosis control. All induced mice eventually developed mammary tumours with 9qA1 (Yap1) amplification and/or 6qA2 (Met) amplification. The resulting tumours exhibited a MaSC-like expression signature and most closely resembled Claudin-Low breast cancer. Overall, these data suggest that although p53 does not directly control the luminal fate, its loss facilitates acquisition of MaSC-like properties by luminal cells and predisposes them to development of mammary tumours with loss of luminal identity. Our data also suggest that Claudin-Low breast cancer can originate from luminal cells, possibly upon transition through a basal-like state. Overall design: Total RNAs from YFP+ mammary epithelial cells sorted from K8-CreER;Trp53L/L;Rosa26-Stop-YFP or K8-CreER;Rosa26-Stop-YFP females 4 weeks after tamoxifen induction, or from mammary tumors developed in Trp53L/L females or K8-CreER;Trp53L/L females 6-7 months after intraductal injection of Ad-K8-Cre adenovirus or tamoxifen induction, respectively, were prepared and subjected to microarray expression profiling.
Project description:RNA-seq of breast tumor from Pik3caH1047R/+ mammary gland transduced with single or pooled lentivirus. Overall design: Tumor from Pik3caH1047R/+ mammary gland transduced with single or pooled lentivirus.
Project description:The aim of the experiment was to produce a mammary stem cell gene signature based on very high purity isolation of mouse mammary stem cells. Gene expression in the stem cells would be compared to gene expression in other cell types from the mammary epithelium to identify genes which may be important for regulation of mammary stem cell function.