Project description:We discovered a novel somatic gene fusion, CD74-NRG1, by transcriptome sequencing of 25 lung adenocarcinomas of never smokers. By screening 102 lung adenocarcinomas negative for known oncogenic alterations we found four additional fusion-positive tumors, all of which were of the invasive mucinous subtype. Mechanistically, CD74-NRG1 leads to extracellular expression of the EGF-like domain of NRG1 III-beta3, thereby providing the ligand for ERBB2-ERBB3 receptor complexes. Accordingly, ERBB2 and ERBB3 expression was high in the index case and expression of phospho-ERBB3 was specifically found in tumors bearing the fusion (p<0.0001). Ectopic expression of CD74-NRG1 in lung cancer cell lines expressing ERBB2 and ERBB3 activated ERBB3 and the PI3K-AKT pathway, and led to increased colony formation in soft agar. Thus, CD74-NRG1 gene fusions are activating genomic alterations in invasive mucinous adenocarcinomas and may offer a therapeutic opportunity for a lung tumor subtype with, so far, no effective treatment.
Project description:Accurate annotation of regulatory RNAs is a complex task but nevertheless essential as sRNA molecular and functional studies ensue from it. Several formerly considered small RNAs (sRNA) are now known to be parts of UTR transcripts. In light of experimental data, we review hundreds of Staphylococcus aureus putative regulatory RNAs. We pinpoint those that are likely acting in trans and are not expressed from the opposite strand of a coding gene. We conclude that HG003, a NCTC8325 derivative strain, has about 50 bona fide sRNAs, indicating that these RNAs are less numerous than commonly stated.
Project description:The post-translational modification of proteins by ubiquitination is a highly regulated process that involves a dynamic, three-step enzymatic cascade, where more than 600 E3 ligases play a critical role in recognizing specific substrates for modification. Separating bona fide targets of E3s from E3-interacting proteins remains a major challenge in the field. In this study, we present BioE3, a novel approach for identifying substrates of ubiquitin-like (UbL) E3 ligases of interest. Using BirA-E3 ligase fusion proteins and bioUbLs, the method facilitates site-specific biotinylation of UbL-modified substrates for proteomic identification. We demonstrate that the BioE3 system can identify both known and novel targets of two RING-type ubiquitin E3 ligases: RNF4, known to be involved in DNA damage response and the regulation of PML nuclear bodies, and MIB1, implicated in endocytosis, autophagy, and centrosomal protein homeostasis. We further show the versatility of BioE3 by identifying targets of an organelle-specific E3 (MARCH5) and a relatively uncharacterized E3 (RNF214). Furthermore, we show that BioE3 works with HECT-type E3 ligases and identify novel targets of NEDD4 involved in vesicular trafficking. BioE3 is a powerful tool that enables identification of bona fide substrates of UbL E3 ligases and how they change with chemical perturbations, which may be useful for the emerging applications in targeted protein degradation (TPD). BioE3 may also be applicable for UbLs beyond Ub and SUMO, as well as other E3 ligase classes. The resulting knowledge can shed light on the regulation of cellular processes by the complex UbL network, advancing our understanding of fundamental biological mechanisms.
Project description:Dynamic interactions between RNA and RNA-binding proteins (RBPs) regulate a broad spectrum of bacterial functions. Here, we have characterised how the RBPome is dynamically rewired during the E. coli life cycle. We applied Orthogonal Organic Phase Separation (OOPS) coupled with an RNase digestion step to shortlist bona fide bacterial RBPs and assessed whether proteins bound RNA differentially express at different cell proliferation stages.
Project description:Hyperglycemia is a risk factor for breast cancer-related morbidity and mortality. Hyperglycemia induces Neuregulin 1 (Nrg1) overexpression in breast cancer, which subsequently promotes tumor progression. However, molecular mechanisms underlying hyperglycemia-induced Nrg1 overexpression remain poorly understood. Here, We used DNA-protein pull-down using Nrg1 enhancer sequence as bait followed by LC/MS and identified RBPJ as a key component of the Nrg1 enhanceosome. We show that hyperglycemia causes active histone modifications at the Nrg1 enhancer, forming enhanceosome complexes where RBPJ, P300, and SETD1A are recruited to upregulate Nrg1 expression.