Epigenetic changes induced by in utero dietary challenge result in phenotypic variability in successive generations of mice.

Transmission of epigenetic information between generations occurs in nematodes, flies and plants, mediated by specialised small RNA pathways, modified histones and DNA methylation. Similar processes in mammals can also affect phenotype through intergenerational or trans-generational mechanisms. Here we generate a luciferase knock-in reporter mouse for the imprinted Dlk1 locus to visualise and track epigenetic fidelity across generations.

Forward and Reverse Genetics of B Cell Malignancies: From Insertional Mutagenesis to CRISPR-Cas.

Cancer genome sequencing has identified dozens of mutations with a putative role in lymphomagenesis and leukemogenesis. Validation of driver mutations responsible for B cell neoplasms is complicated by the volume of mutations worthy of investigation and by the complex ways that multiple mutations arising from different stages of B cell development can cooperate. Forward and reverse genetic strategies in mice can provide complementary validation of human driver genes and in some cases comparative genomics of these models with human tumors has directed the identification of new drivers in human malignancies.

Galactose-modified duocarmycin prodrugs as senolytics.

Senescence is a stable growth arrest that impairs the replication of damaged, old or preneoplastic cells, therefore contributing to tissue homeostasis. Senescent cells accumulate during ageing and are associated with cancer, fibrosis and many age-related pathologies. Recent evidence suggests that the selective elimination of senescent cells can be effective on the treatment of many of these senescence-associated diseases.

LUMI-PCR: an Illumina platform ligation-mediated PCR protocol for integration site cloning, provides molecular quantitation of integration sites.

Background: Ligation-mediated PCR protocols have diverse uses including the identification of integration sites of insertional mutagens, integrating vectors and naturally occurring mobile genetic elements. For approaches that employ NGS sequencing, the relative abundance of integrations within a complex mixture is typically determined through the use of read counts or unique fragment lengths from a ligation of sheared DNA; however, these estimates may be skewed by PCR amplification biases and saturation of sequencing coverage.

Results: Here we describe a modification of our previous splinkerette based ligation-mediated PCR using a novel Illumina-compatible adapter design that prevents amplification of non-target DNA and incorporates unique molecular identifiers.

Cardiac glycosides are broad-spectrum senolytics.

Senescence is a cellular stress response that results in the stable arrest of old, damaged or preneoplastic cells. Oncogene-induced senescence is tumor suppressive but can also exacerbate tumorigenesis through the secretion of pro-inflammatory factors from senescent cells. Drugs that selectively kill senescent cells, termed senolytics, have proved beneficial in animal models of many age-associated diseases.

Author Correction: Subclonal mutation selection in mouse lymphomagenesis identifies known cancer loci and suggests novel candidates.

The original version of this Article contained an error in the hyperlink for the online repository http://mulvdb.org which was incorrectly given as http://mulv.lms.

Subclonal mutation selection in mouse lymphomagenesis identifies known cancer loci and suggests novel candidates.

Determining whether recurrent but rare cancer mutations are bona fide driver mutations remains a bottleneck in cancer research. Here we present the most comprehensive analysis of murine leukemia virus-driven lymphomagenesis produced to date, sequencing 700,000 mutations from >500 malignancies collected at time points throughout tumor development. This scale of data allows novel statistical approaches for identifying selected mutations and yields a high-resolution, genome-wide map of the selective forces surrounding cancer gene loci.

Visualizing Changes in Cdkn1c Expression Links Early-Life Adversity to Imprint Mis-regulation in Adults.

Imprinted genes are regulated according to parental origin and can influence embryonic growth and metabolism and confer disease susceptibility. Here, we designed sensitive allele-specific reporters to non-invasively monitor imprinted Cdkn1c expression in mice and showed that expression was modulated by environmental factors encountered in utero. Acute exposure to chromatin-modifying drugs resulted in de-repression of paternally inherited (silent) Cdkn1c alleles in embryos that was temporary and resolved after birth.

Dual EZH2 and EHMT2 histone methyltransferase inhibition increases biological efficacy in breast cancer cells.

Background: Many cancers show aberrant silencing of gene expression and overexpression of histone methyltransferases. The histone methyltransferases (HKMT) EZH2 and EHMT2 maintain the repressive chromatin histone methylation marks H3K27me and H3K9me, respectively, which are associated with transcriptional silencing. Although selective HKMT inhibitors reduce levels of individual repressive marks, removal of H3K27me3 by specific EZH2 inhibitors, for instance, may not be sufficient for inducing the expression of genes with multiple repressive marks.

GFAP-Cre-mediated transgenic activation of Bmi1 results in pituitary tumors.

Bmi1 is a member of the polycomb repressive complex 1 and plays different roles during embryonic development, depending on the developmental context. Bmi1 over expression is observed in many types of cancer, including tumors of astroglial and neural origin. Although genetic depletion of Bmi1 has been described to result in tumor inhibitory effects partly through INK4A/Arf mediated senescence and apoptosis and also through INK4A/Arf independent effects, it has not been proven that Bmi1 can be causally involved in the formation of these tumors.

Insertional mutagenesis identifies multiple networks of cooperating genes driving intestinal tumorigenesis.

The evolution of colorectal cancer suggests the involvement of many genes. To identify new drivers of intestinal cancer, we performed insertional mutagenesis using the Sleeping Beauty transposon system in mice carrying germline or somatic Apc mutations. By analyzing common insertion sites (CISs) isolated from 446 tumors, we identified many hundreds of candidate cancer drivers.

Mutant nucleophosmin and cooperating pathways drive leukemia initiation and progression in mice.

Acute myeloid leukemia (AML) is a molecularly diverse malignancy with a poor prognosis whose largest subgroup is characterized by somatic mutations in NPM1, which encodes nucleophosmin. These mutations, termed NPM1c, result in cytoplasmic dislocation of nucleophosmin and are associated with distinctive transcriptional signatures, yet their role in leukemogenesis remains obscure. Here we report that activation of a humanized Npm1c knock-in allele in mouse hemopoietic stem cells causes Hox gene overexpression, enhanced self renewal and expanded myelopoiesis.

Novel candidate cancer genes identified by a large-scale cross-species comparative oncogenomics approach.

Comparative genomic hybridization (CGH) can reveal important disease genes but the large regions identified could sometimes contain hundreds of genes. Here we combine high-resolution CGH analysis of 598 human cancer cell lines with insertion sites isolated from 1,005 mouse tumors induced with the murine leukemia virus (MuLV). This cross-species oncogenomic analysis revealed candidate tumor suppressor genes and oncogenes mutated in both human and mouse tumors, making them strong candidates for novel cancer genes.

Insertional mutagenesis in mice deficient for p15Ink4b, p16Ink4a, p21Cip1, and p27Kip1 reveals cancer gene interactions and correlations with tumor phenotypes.

The cyclin dependent kinase (CDK) inhibitors p15, p16, p21, and p27 are frequently deleted, silenced, or downregulated in many malignancies. Inactivation of CDK inhibitors predisposes mice to tumor development, showing that these genes function as tumor suppressors. Here, we describe high-throughput murine leukemia virus insertional mutagenesis screens in mice that are deficient for one or two CDK inhibitors.

A high-throughput splinkerette-PCR method for the isolation and sequencing of retroviral insertion sites.

Insertional mutagens such as viruses and transposons are a useful tool for performing forward genetic screens in mice to discover cancer genes. These screens are most effective when performed using hundreds of mice; however, until recently, the cost-effective isolation and sequencing of insertion sites has been a major limitation to performing screens on this scale. Here we present a method for the high-throughput isolation of insertion sites using a highly efficient splinkerette-PCR method coupled with capillary or 454 sequencing.

Instant conditional transgenesis in the mouse hematopoietic compartment.

Adoptive transfer of retrovirally transduced stem cells has recently been described for instant transgenesis in the hematopoietic compartment of mice. This method circumvents the need to manipulate the germline. However, cell type specific gene expression in this 'retrogenic' mouse model has remained tedious.

Large-scale mutagenesis in p19(ARF)- and p53-deficient mice identifies cancer genes and their collaborative networks.

p53 and p19(ARF) are tumor suppressors frequently mutated in human tumors. In a high-throughput screen in mice for mutations collaborating with either p53 or p19(ARF) deficiency, we identified 10,806 retroviral insertion sites, implicating over 300 loci in tumorigenesis. This dataset reveals 20 genes that are specifically mutated in either p19(ARF)-deficient, p53-deficient or wild-type mice (including Flt3, mmu-mir-106a-363, Smg6, and Ccnd3), as well as networks of significant collaborative and mutually exclusive interactions between cancer genes.

Co-occurrence analysis of insertional mutagenesis data reveals cooperating oncogenes.

Motivation: Cancers are caused by an accumulation of multiple independent mutations that collectively deregulate cellular pathways, e.g. such as those regulating cell division and cell-death.

Detecting statistically significant common insertion sites in retroviral insertional mutagenesis screens.

Retroviral insertional mutagenesis screens, which identify genes involved in tumor development in mice, have yielded a substantial number of retroviral integration sites, and this number is expected to grow substantially due to the introduction of high-throughput screening techniques. The data of various retroviral insertional mutagenesis screens are compiled in the publicly available Retroviral Tagged Cancer Gene Database (RTCGD). Integrally analyzing these screens for the presence of common insertion sites (CISs, i.