Conjugative transfer of broad host range plasmids to an acidobacterial strain, Edaphobacter aggregans

The Acidobacteria phylum is of high ecological interest. Its members are ubiquitous and particularlyabundant in soils but many are recalcitrant to cultivation in the laboratory. Thus, the ability of Acidobac-teria to capture and maintain plasmids remains largely unexplored. In this work we tested the transferand the stability of (i) the PromA plasmid pMOL98 and (ii) the IncQ plasmid pKT230 to the acidobacterialstrain Edaphobacter aggregans DSM 19364. To this end quantitative conjugation assays were performedand transconjugants were scored for plasmid-borne antibiotic selection markers. The tested plasmidswere transferred and maintained in the new host. Plasmid pMOL98 was more stable than pKT230 in Ed.aggregans in the absence of positive selection. Thus, from an ecological point of view, we have extendedthe host range of PromA and IncQ plasmids for the first time to an acidobacterial strain. Furthermore,we have uncovered the potential of Acidobacteria to capture as-yet-unknown plasmids and to foster thedevelopment of new cloning and expression systems for the exploitation of biotechnologically valuablesoil resources

Genome-wide Screens Implicate Loss of Cullin Ring Ligase 3 in Persistent Proliferation and Genome Instability in TP53-Deficient Cells

TP53 deficiency is the most common alteration in cancer; however, this alone is typically insufficient to drive tumorigenesis. To identify genes promoting tumorigenesis in combination with TP53 deficiency, we perform genome-wide CRISPR-Cas9 knockout screens coupled with proliferation and transformation assays in isogenic cell lines. Loss of several known tumor suppressors enhances cellular proliferation and transformation. Loss of neddylation pathway genes promotes uncontrolled proliferation exclusively in TP53-deficient cells. Combined loss of CUL3 and TP53 activates an oncogenic transcriptional program governed by the nuclear factor kappa B (NF-kappa B), AP-1, and transforming growth factor beta (TGF-beta) pathways. This program maintains persistent cellular proliferation, induces partial epithelial to mesenchymal transition, and increases DNA damage, genomic instability, and chromosomal rearrangements. Our findings reveal CUL3 loss as a key event stimulating persistent proliferation in TP53-deficient cells. These findings may be clinically relevant, since TP53-CUL3-deficient cells are highly sensitive to ataxia telangiectasia mutated (ATM) inhibition, exposing a vulnerability that could be exploited for cancer treatment

Pan-cancer analysis of somatic copy-number alterations implicates IRS4 and IGF2 in enhancer hijacking

Extensive prior research focused on somatic copy-number alterations (SCNAs) affecting cancer genes, yet the extent to which recurrent SCNAs exert their influence through rearrangement of cis-regulatory elements (CREs) remains unclear. Here we present a framework for inferring cancer-related gene overexpression resulting from CRE reorganization (e.g., enhancer hijacking) by integrating SCNAs, gene expression data and information on topologically associating domains (TADS). Analysis of 7,416 cancer genomes uncovered several pan-cancer candidate genes, including IRS4, SMARCA1 and TERT. We demonstrate that IRS4 overexpression in lung cancer is associated with recurrent deletions in cis, and we present evidence supporting a tumor promoting role. We additionally pursued cancer-type-specific analyses and uncovered IGF2 as a target for enhancer hijacking in colorectal cancer. Recurrent tandem duplications intersecting with a TAD boundary mediate de novo formation of a 3D contact domain comprising IGF2 and a lineage-specific super-enhancer, resulting in high-level gene activation. Our framework enables systematic inference of CRE rearrangements mediating dysregulation in cancer