Showing papers by "Toby J. Gibson published in 2017"

Metabolic shifts in residual breast cancer drive tumor recurrence

Abstract: Tumor recurrence is the leading cause of breast cancer-related death. Recurrences are largely driven by cancer cells that survive therapeutic intervention. This poorly understood population is referred to as minimal residual disease. Here, using mouse models that faithfully recapitulate human disease together with organoid cultures, we have demonstrated that residual cells acquire a transcriptionally distinct state from normal epithelium and primary tumors. Gene expression changes and functional characterization revealed altered lipid metabolism and elevated ROS as hallmarks of the cells that survive tumor regression. These residual cells exhibited increased oxidative DNA damage, potentiating the acquisition of somatic mutations during hormonal-induced expansion of the mammary cell population. Inhibition of either cellular fatty acid synthesis or fatty acid transport into mitochondria reduced cellular ROS levels and DNA damage, linking these features to lipid metabolism. Direct perturbation of these hallmarks in vivo, either by scavenging ROS or by halting the cyclic mammary cell population expansion, attenuated tumor recurrence. Finally, these observations were mirrored in transcriptomic and histological signatures of residual cancer cells from neoadjuvant-treated breast cancer patients. These results highlight the potential of lipid metabolism and ROS as therapeutic targets for reducing tumor recurrence in breast cancer patients.

Degrons in cancer

Abstract: Degrons are the elements that are used by E3 ubiquitin ligases to target proteins for degradation. Most degrons are short linear motifs embedded within the sequences of modular proteins. As regulatory sites for protein abundance, they are important for many different cellular processes, such as progression through the cell cycle and monitoring cellular hypoxia. Degrons enable the elimination of proteins that are no longer required, preventing their possible dysfunction. Although the human genome encodes ~600 E3 ubiquitin ligases, only a fraction of these enzymes have well-defined target degrons. Thus, for most cellular proteins, the destruction mechanisms are poorly understood. This is important for many diseases, especially for cancer, a disease that involves the enhanced expression of oncogenes and the persistence of encoded oncoproteins coupled with reduced abundance of tumor suppressors. Loss-of-function mutations occur in the degrons of several oncoproteins, such as the transcription factors MYC and NRF2, and in various mitogenic receptors, such as NOTCH1 and several receptor tyrosine kinases. Mutations eliminating the function of the β-catenin degron are found in many cancers and are considered one of the most abundant mutations driving carcinogenesis. In this Review, we describe the current knowledge of degrons in cancer and suggest that increased research on the "dark degrome" (unknown degron-E3 relationships) would enhance progress in cancer research.

CTCF-Mediated Chromatin Loops between Promoter and Gene Body Regulate Alternative Splicing across Individuals

Abstract: The CCCTC-binding factor (CTCF) is known to establish long-range DNA contacts that alter the three-dimensional architecture of chromatin, but how the presence of CTCF influences nearby gene expression is still poorly understood. Here, we analyze CTCF chromatin immunoprecipitation sequencing, RNA sequencing, and Hi-C data, together with genotypes from a healthy human cohort, and measure statistical associations between inter-individual variability in CTCF binding and alternative exon usage. We demonstrate that CTCF-mediated chromatin loops between promoters and intragenic regions are prevalent and that when exons are in physical proximity with their promoters, CTCF binding correlates with exon inclusion in spliced mRNA. Genome-wide, CTCF-bound exons are enriched for genes involved in signaling and cellular stress-response pathways. Structural analysis of three specific examples, checkpoint kinase 2 (CHK2), CDC-like kinase 3 (CLK3), and euchromatic histone-lysine N-methyltransferase (EHMT1), suggests that CTCF-mediated exon inclusion is likely to downregulate enzyme activity by disrupting annotated protein domains. In total, our study suggests that alternative exon usage is regulated by CTCF-dependent chromatin structure.

CiliaCarta: an integrated and validated compendium of ciliary genes

Abstract: The cilium is an essential organelle at the surface of most mammalian cells whose dysfunction causes a wide range of genetic diseases collectively called ciliopathies. The current rate at which new ciliopathy genes are identified suggests that many ciliary components remain undiscovered. We generated and rigorously analyzed genomic, proteomic, transcriptomic and evolutionary data and systematically integrated these using Bayesian statistics into a predictive score for ciliary function. This resulted in 285 candidate ciliary genes. We found experimental evidence of ciliary associations for 24 out of 36 analyzed candidate proteins. In addition, we show that OSCP1, which has previously been implicated in two distinct non-ciliary functions, causes a cilium dysfunction phenotype when depleted in zebrafish. The candidate list forms the basis of CiliaCarta, a comprehensive ciliary compendium covering 836 genes. The resource can be used to objectively prioritize candidate genes in whole exome or genome sequencing of ciliopathy patients and can be accessed at http://bioinformatics.bio.uu.nl/john/syscilia/ciliacarta/.

Exploring Short Linear Motifs Using the ELM Database and Tools.

Abstract: The Eukaryotic Linear Motif (ELM) resource is dedicated to the characterization and prediction of short linear motifs (SLiMs). SLiMs are compact, degenerate peptide segments found in many proteins and essential to almost all cellular processes. However, despite their abundance, SLiMs remain largely uncharacterized. The ELM database is a collection of manually annotated SLiM instances curated from experimental literature. In this article we illustrate how to browse and search the database for curated SLiM data, and cover the different types of data integrated in the resource. We also cover how to use this resource in order to predict SLiMs in known as well as novel proteins, and how to interpret the results generated by the ELM prediction pipeline. The ELM database is a very rich resource, and in the following protocols we give helpful examples to demonstrate how this knowledge can be used to improve your own research. © 2017 by John Wiley & Sons, Inc.

Intermolecular base stacking mediates RNA-RNA interaction in a crystal structure of the RNA chaperone Hfq

Abstract: The RNA-chaperone Hfq catalyses the annealing of bacterial small RNAs (sRNAs) with target mRNAs to regulate gene expression in response to environmental stimuli. Hfq acts on a diverse set of sRNA-mRNA pairs using a variety of different molecular mechanisms. Here, we present an unusual crystal structure showing two Hfq-RNA complexes interacting via their bound RNA molecules. The structure contains two Hfq6:A18 RNA assemblies positioned face-to-face, with the RNA molecules turned towards each other and connected via interdigitating base stacking interactions at the center. Biochemical data further confirm the observed interaction, and indicate that RNA-mediated contacts occur between Hfq-RNA complexes with various (ARN)X motif containing RNA sequences in vitro, including the stress response regulator OxyS and its target, fhlA. A systematic computational survey also shows that phylogenetically conserved (ARN)X motifs are present in a subset of sRNAs, some of which share similar modular architectures. We hypothesise that Hfq can co-opt RNA-RNA base stacking, an unanticipated structural trick, to promote the interaction of (ARN)X motif containing sRNAs with target mRNAs on a “speed-dating” fashion, thereby supporting their regulatory function.

The Gene Ontology of eukaryotic cilia and flagella

Abstract: Recent research into ciliary structure and function provides important insights into inherited diseases termed ciliopathies and other cilia-related disorders. This wealth of knowledge needs to be translated into a computational representation to be fully exploitable by the research community. To this end, members of the Gene Ontology (GO) and SYSCILIA Consortia have worked together to improve representation of ciliary substructures and processes in GO. Members of the SYSCILIA and Gene Ontology Consortia suggested additions and changes to GO, to reflect new knowledge in the field. The project initially aimed to improve coverage of ciliary parts, and was then broadened to cilia-related biological processes. Discussions were documented in a public tracker. We engaged the broader cilia community via direct consultation and by referring to the literature. Ontology updates were implemented via ontology editing tools. So far, we have created or modified 127 GO terms representing parts and processes related to eukaryotic cilia/flagella or prokaryotic flagella. A growing number of biological pathways are known to involve cilia, and we continue to incorporate this knowledge in GO. The resulting expansion in GO allows more precise representation of experimentally derived knowledge, and SYSCILIA and GO biocurators have created 199 annotations to 50 human ciliary proteins. The revised ontology was also used to curate mouse proteins in a collaborative project. The revised GO and annotations, used in comparative ‘before and after’ analyses of representative ciliary datasets, improve enrichment results significantly. Our work has resulted in a broader and deeper coverage of ciliary composition and function. These improvements in ontology and protein annotation will benefit all users of GO enrichment analysis tools, as well as the ciliary research community, in areas ranging from microscopy image annotation to interpretation of high-throughput studies. We welcome feedback to further enhance the representation of cilia biology in GO.

A Tool to Build Up-To-Date Gene Annotations for Affymetrix Microarrays

Abstract: Genome-wide expression profiling and genotyping is widely applied in functional genomics research, ranging from stem cell studies to cancer, in drug response studies, and in clinical diagnostics. The Affymetrix GeneChip microarrays represent the most popular platform for such assays. Nevertheless, due to rapid and continuous improvement of the knowledge about the genome, the definition of many of the genes and transcripts change, and new genes are discovered. Thus the original probe information is out-dated for a number of Affymetrix platforms, and needs to be re-defined. It has been demonstrated, that accurate probe set definition improves both coverage of the gene expression analysis and its statistical power. Therefore we developed a method that incorporates the most recent genome annotations into the annotation of the microarray probe sets, using tools from the next generation sequencing. Additionally our method allows to quickly build project specific gene annotation models, as well as for comparison of microarray to RNAseq data.

Comprehensive gene ontology annotation of ciliary genes in the laboratory mouse

Abstract: Interest in primary cilia has increased dramatically over the last ten years as it has become clear that ciliopathies are an underlying cause of numerous human diseases including some types of retinitis pigmentosa and polycystic kidney disease. Once thought to be restricted to a few cell types, it is now clear that primary cilia are found on almost all vertebrate cells and are critical to Sonic hedgehog (Shh) signaling. Mouse models play a key role in developing our understanding of the role of primary cilia in control of Shh signaling in development throughout the embryo and in ongoing maintenance of structures such as photoreceptors. To maximize the utility of the wealth of experimental data generated by these mouse ciliopathy models, we have initiated a project to comprehensively annotate ciliary genes of the laboratory mouse using Gene Ontology (GO) terms to describe their molecular functions, biological roles, and cellular locations. We are guided by the SysCilia gold standard of known human ciliary components as a starting point, but will also include additional genes experimentally shown to be involved in ciliary function in the mouse. If needed, we will also update the Gene Ontology to add new terms representing recent advances in our understanding of ciliary biology. Comprehensive GO annotation of ciliary genes in the mouse will be a great resource to those doing high throughput studies or comparative genomic analysis across species, and may help us better understand the similarities and differences between mouse and human in the role of Sonic hedgehog signaling in development. This work is funded by HG 002273 to the Gene Ontology Consortium. GENEONTOLOGY Unifying Biology ID\t\r Mapping\t\r at\t\r hYp://www.uniprot.org/ 307\t\r mouse\t\r genes\t\r on\t\r cilia\t\r candidate\t\r list AddiHonal\t\r research\t\r for\t\r human\t\r gene\t\r with\t\r no\t\r mouse\t\r homologs\t\r via\t\r MouseMine IdenHficaHon\t\r of\t\r mouse\t\r homologs\t\r \t\r with\t\r MouseMine\t\r -­‐\t\r hYp://www.mousemine.org/