Deciphering the molecular alterations sustaining an inverted apicobasal polarity phenotype in colorectal cancer clusters (Tumour Spheres with Inverted Polarity, TSIPs)
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PROVIDER: PRJNA638924 | ENA |
REPOSITORIES: ENA
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Project description:Colorectal cancer (CRC) is the second deadliest form of cancer and most of patients die from their metastases. As previously shown by our team, in serrated CRC patients, metastases are due to spherical clusters of epithelial cancer cells displaying an inverted apicobasal polarity. We called them TSIPs for Tumor spheres with inverted polarity. In this study, we investigated the alterations sustaining this remarkable phenotype using PDXs and organoids techniques. In particular, we compared a model that respond normally to its microenvironment (mainly 3D collagen-I networks) to two models that keep their inverted polarity. A whole transcriptome investigation allowed us to better understand the mechanisms of such a topology.
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Project description:DNA replication errors are a major driver of evolution—from single nucleotide polymorphisms to large-scale copy number variations (CNVs). Here we test a specific replication-based model to explain the generation of interstitial, inverted triplications. While no genetic information is lost, the novel inversion junctions and increased copy number of the included sequences create the potential for adaptive phenotypes. The model—Origin-Dependent Inverted-Repeat Amplification (ODIRA)—proposes that a replication error at pre-existing short, interrupted, inverted repeats in genomic sequences generates an extrachromosomal, inverted dimeric, autonomously replicating intermediate; subsequent genomic integration of the dimer yields this class of CNV without loss of distal chromosomal sequences. We used a combination of in vitro and in vivo approaches to test the feasibility of the proposed replication error and its downstream consequences on chromosome structure in the yeast Saccharomyces cerevisiae. We show that the proposed replication error—the ligation of leading and lagging nascent strands to create “closed” forks—can occur in vitro at short, interrupted inverted repeats. The removal of molecules with two closed forks results in a hairpin-capped linear duplex that we show replicates in vivo to create an inverted, dimeric plasmid that subsequently integrates into the genome by homologous recombination, creating an inverted triplication. While other models have been proposed to explain inverted triplications and their derivatives, our model can also explain the generation of human, de novo, inverted amplicons that have a 2:1 mixture of sequences from both homologues of a single parent—a feature readily explained by a plasmid intermediate that arises from one homologue and integrates into the other homologue prior to meiosis. Our tests of key features of ODIRA lend support to this mechanism and suggest further avenues of enquiry to unravel the origins of interstitial, inverted CNVs pivotal in human health and evolution.
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