Bringer, Pauline, Jensen, Michael Rugaard and Gembarska, Agnieszka (2015) Identification of piggyBac transposon insertions in malignant tumors. Master degree Bio-engineering applied to health, University of Toulouse III.

Abstract

In the last decades, many important genes responsible for the genesis of various cancers have been discovered, their mutations precisely identified, and the pathways through which they act characterized.
Nevertheless the wide range of possible genetic alterations, their functional relevance, their downstream targets or effector pathways represent a major hindrance to the therapeutical success. Genetic instability and heterogeneity in cancer vary between the different types and stages, tissues and individuals. While there has been considerable progress in understanding the impact of genetic mechanisms in tumorigenesis, large efforts must be made to develop experimental strategies to understand these cellular processes. The current challenge in the oncology field is to develop efficient tools to try to identify gene responsible for tumorigenesis processes to treat human cancers.
Here we describe the use of piggyBac transposon system to identify driver genes in cancer. Transgenic mice carrying several transposon copies able to randomly hop around the genome (RosaPB/+; ATP1/+) were generated. ATP1 transposon can confer proliferative advantage by activating or repressing genes expression. To accelerate the disease onset, we added a sensitizing mutation (Arf-/-) and studied mice with the three genetic components. After aging of mice, tumors developed due to Arf-/- and PB insertional mutagenesis, and were harvested. To create a bank with a broad range of tumor models, harvested tumors were passaged and expanded in recipient mice. Interestingly, our ATP1 DNA copy numbers quantification often showed gain of ATP1 in tumors compared to normal tissue, suggesting PB insertional mutagenesis is favorable for tumors. Genes of interest were identified by deep sequencing of amplicons of DNA fragments that contain extremities of ATP1 sequences.
Statistical analysis of transposon integration sites in tumor genomes identified Braf gene as target in almost all RosaPB/+; ATP1/+; Arf-/- tumors, suggesting a genetic cooperation between Arf deletion and Braf activation. Other genes are now evaluated. Here we have developed a powerful in vivo genomic tool allowing the identification of genes driving cancer.

Item Type:	Article
Keywords:	gene discovery, transposon, piggyBac, engineered mouse models
Date Deposited:	26 Apr 2016 23:45
Last Modified:	26 Apr 2016 23:45
URI:	https://oak.novartis.com/id/eprint/26459