Impact Of Cancer-associated Mutations Research Articles

Deciphering principles of nucleosome interactions and impact of cancer-associated mutations from comprehensive interaction network analysis.

Nucleosomes represent hubs in chromatin organization and gene regulation and interact with a plethora of chromatin factors through different modes. In addition, alterations in histone proteins such as cancer mutations and post-translational modifications have profound effects on histone/nucleosome interactions. To elucidate the principles of histone interactions and the effects of those alterations, we developed histone interactomes for comprehensive mapping of histone-histone interactions (HHIs), histone-DNA interactions (HDIs), histone-partner interactions (HPIs) and DNA-partner interactions (DPIs) of 37 organisms, which contains a total of 3808 HPIs from 2544 binding proteins and 339 HHIs, 100 HDIs and 142 DPIs across 110 histone variants. With the developed networks, we explored histone interactions at different levels of granularities (protein-, domain- and residue-level) and performed systematic analysis on histone interactions at a large scale. Our analyses have characterized the preferred binding hotspots on both nucleosomal/linker DNA and histone octamer and unraveled diverse binding modes between nucleosome and different classes of binding partners. Last, to understand the impact of histone cancer-associated mutations on histone/nucleosome interactions, we complied one comprehensive cancer mutation dataset including 7940 cancer-associated histone mutations and further mapped those mutations onto 419,125 histone interactions at the residue level. Our quantitative analyses point to histone cancer-associated mutations' strongly disruptive effects on HHIs, HDIs and HPIs. We have further predicted 57 recurrent histone cancer mutations that have large effects on histone/nucleosome interactions and may have driver status in oncogenesis.

Impact of cancer-associated mutations in CC chemokine receptor 2 on receptor function and antagonism

CC chemokine receptor 2 (CCR2), a G protein-coupled receptor, plays a role in many cancer-related processes such as metastasis formation and immunosuppression. Since ∼ 20 % of human cancers contain mutations in G protein-coupled receptors, ten cancer-associated CCR2 mutants obtained from the Genome Data Commons were investigated for their effect on receptor functionality and antagonist binding. Mutations were selected based on either their vicinity to CCR2′s orthosteric or allosteric binding sites or their presence in conserved amino acid motifs. One of the mutant receptors, namely S101P2.63 with a mutation near the orthosteric binding site, did not express on the cell surface. All other studied mutants showed a decrease in or a lack of G protein activation in response to the main endogenous CCR2 ligand CCL2, but no change in potency was observed. Furthermore, INCB3344 and LUF7482 were chosen as representative orthosteric and allosteric antagonists, respectively. No change in potency was observed in a functional assay, but mutations located at F1163.28 impacted orthosteric antagonist binding significantly, while allosteric antagonist binding was abolished for L134Q3.46 and D137N3.49 mutants. As CC chemokine receptor 2 is an attractive drug target in cancer, the negative effect of these mutations on receptor functionality and drugability should be considered in the drug discovery process.

Brca2-mutant zebrafish exhibit context- and tissue-dependent alterations in cell phenotypes and response to injury

Cancer cells frequently co-opt molecular programs that are normally activated in specific contexts, such as embryonic development and the response to injury. Determining the impact of cancer-associated mutations on cellular phenotypes within these discrete contexts can provide new insight into how such mutations lead to dysregulated cell behaviors and subsequent cancer onset. Here we assess the impact of heritable BRCA2 mutation on embryonic development and the injury response using a zebrafish model (Danio rerio). Unlike most mouse models for BRCA2 mutation, brca2-mutant zebrafish are fully viable and thus provide a unique tool for assessing both embryonic and adult phenotypes. We find that maternally provided brca2 is critical for normal oocyte development and embryonic survival in zebrafish, suggesting that embryonic lethality associated with BRCA2 mutation is likely to reflect defects in both meiotic and embryonic developmental programs. On the other hand, we find that adult brca2-mutant zebrafish exhibit aberrant proliferation of several cell types under basal conditions and in response to injury in tissues at high risk for cancer development. These divergent effects exemplify the often-paradoxical outcomes that occur in embryos (embryonic lethality) versus adult animals (cancer predisposition) with mutations in cancer susceptibility genes such as BRCA2. The altered cell behaviors identified in brca2-mutant embryonic and adult tissues, particularly in adult tissues at high risk for cancer, indicate that the effects of BRCA2 mutation on cellular phenotypes are both context- and tissue-dependent.

Impact of Cancer-Associated Mutations in CC Chemokine Receptor 2 on Receptor Function and Antagonism

Impact of Cancer-Associated Mutations in CC Chemokine Receptor 2 on Receptor Function and Antagonism

Impact of cancer-associated mutations in Hsh155/SF3b1 HEAT repeats 9-12 on pre-mRNA splicing in Saccharomyces cerevisiae.

Mutations in the splicing machinery have been implicated in a number of human diseases. Most notably, the U2 small nuclear ribonucleoprotein (snRNP) component SF3b1 has been found to be frequently mutated in blood cancers such as myelodysplastic syndromes (MDS). SF3b1 is a highly conserved HEAT repeat (HR)-containing protein and most of these blood cancer mutations cluster in a hot spot located in HR4-8. Recently, a second mutational hotspot has been identified in SF3b1 located in HR9-12 and is associated with acute myeloid leukemias, bladder urothelial carcinomas, and uterine corpus endometrial carcinomas. The consequences of these mutations on SF3b1 functions during splicing have not yet been tested. We incorporated the corresponding mutations into the yeast homolog of SF3b1 and tested their impact on splicing. We find that all of these HR9-12 mutations can support splicing in yeast, and this suggests that none of them are loss of function alleles in humans. The Hsh155V502F mutation alters splicing of several pre-mRNA reporters containing weak branch sites as well as a genetic interaction with Prp2 and physical interactions with Prp5 and Prp3. The ability of a single allele of Hsh155 to perturb interactions with multiple factors functioning at different stages of the splicing reaction suggests that some SF3b1-mutant disease phenotypes may have a complex origin on the spliceosome.

Abstract AP05: OVARIAN CANCER-ASSOCIATED RAD51D MUTATIONS WHICH IMPAIR ITS INTERACTION WITH XRCC2 RESULT IN DNA REPAIR DEFICIENCY

Abstract The proficiency of ovarian cancer cells to repair DNA double-strand breaks (DSBs) by homologous recombination (HR) is a key determinant in predicting response to targeted therapies such as PARP inhibitors (PARPi). The RAD51 paralogs act downstream of BRCA1/2 to facilitate HR. Numerous epidemiological studies have linked mutations in the RAD51 paralogs with hereditary ovarian cancer predisposition. Despite their substantial links to cancer predisposition and development, RAD51 paralog function during HR has remained elusive, in part due to limitations in studying recombination events downstream of RAD51 filament formation. Here we investigate the impact of cancer-associated mutations in the RAD51 paralog, RAD51D, using yeast 2/3-hybrid assays to screen for altered protein-protein interactions. Following the identification of mutations that disrupt the interaction between RAD51D and XRCC2 in yeast, we validated the interaction by co-immunoprecipitation in human cells. Importantly, we determined the impact of these mutations on HR-proficiency using a direct-repeat recombination assay. By characterizing the impact of cancer-associated mutations in the RAD51 paralogs on HR-proficiency, we aim to develop more effective predictive models for therapeutic sensitivity and resistance in patients who harbor similar mutations in these essential genes. Citation Format: Robert A. Baldock, Catherine A. Pressimone, Jared M. Baird, Anton Y. Khodakov, Yoav Karpenshif, Edwige B. Garcin, Stéphanie Gon, Mauro Modesti, Kara A. Bernstein. OVARIAN CANCER-ASSOCIATED RAD51D MUTATIONS WHICH IMPAIR ITS INTERACTION WITH XRCC2 RESULT IN DNA REPAIR DEFICIENCY [abstract]. In: Proceedings of the 12th Biennial Ovarian Cancer Research Symposium; Sep 13-15, 2018; Seattle, WA. Philadelphia (PA): AACR; Clin Cancer Res 2019;25(22 Suppl):Abstract nr AP05.

Abstract 1234: Impact of cancer-associated mutations on MDM2 function

Abstract MDM2 is an E3 ubiquitin ligase that functions as a negative regulator of the tumor suppressor p53. MDM2 can promote the ubiquitination and proteasomal degradation through 26S proteasome. In addition to p53, MDM2 can ubiquitinate and promote the degradation of other proteins including SLUG, PCNA, and TERT. Although MDM2 has a well accepted role as an oncogene in human cancer, it also exhibits the ability to suppress the SLUG-driven epithelial mesenchymal transition (EMT) that is a key component of the metastasis process. The fact that MDM2 is required to suppress the SLUG-driven epithelial mesenchymal transition (EMT) led us to speculate that mutations in the MDM2 gene might impair the proteasomal degradation of key regulatory proteins that control biological events and thereby contribute to tumorogenesis. Here, we studied twenty-one somatic point mutants in MDM2 that occur in human cancer. We find that seven of these MDM2 mutants have lost the capacity to degrade p53. Of these mutants, three have lost the capacity to polyubiquitinate and two do not interact with p53. Additionally, we found that two of the mutants exhibit enhanced ability to degrade p53. We also assessed whether these mutants impacted the ability of mdm2 to degrade SLUG. Although previous studies have shown that MDM2 needs to form a trimeric complex with p53 and SLUG in order for degradation of the latter, we find that MDM2 efficiently degraded SLUG in the absence of p53. Furthermore, MDM2 mutants that do not interact with p53 still retain the ability to degrade SLUG. Our study indicates that mutations in MDM2 can yield mutant proteins that exhibit distinct capacities to degrade its substrates. Further, in vitro and in vivo studies will address the impact of these mutations on the oncogenic properties of MDM2. Citation Format: Krishna M. Chauhan, Luis A. Martinez. Impact of cancer-associated mutations on MDM2 function. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1234. doi:10.1158/1538-7445.AM2015-1234

Origins and functional consequences of somatic mitochondrial DNA mutations in human cancer.

Recent sequencing studies have extensively explored the somatic alterations present in the nuclear genomes of cancers. Although mitochondria control energy metabolism and apoptosis, the origins and impact of cancer-associated mutations in mtDNA are unclear. In this study, we analyzed somatic alterations in mtDNA from 1675 tumors. We identified 1907 somatic substitutions, which exhibited dramatic replicative strand bias, predominantly C > T and A > G on the mitochondrial heavy strand. This strand-asymmetric signature differs from those found in nuclear cancer genomes but matches the inferred germline process shaping primate mtDNA sequence content. A number of mtDNA mutations showed considerable heterogeneity across tumor types. Missense mutations were selectively neutral and often gradually drifted towards homoplasmy over time. In contrast, mutations resulting in protein truncation undergo negative selection and were almost exclusively heteroplasmic. Our findings indicate that the endogenous mutational mechanism has far greater impact than any other external mutagens in mitochondria and is fundamentally linked to mtDNA replication.

Abstract PR11: Medulloblastoma-associated mutations in the DEAD box RNA helicase DDX3X impair protein translation

Abstract Whole genome sequencing technologies recently identified novel recurrent mutations in childhood medulloblastomas (MB), including missense mutations in the DEAD box RNA helicase DDX3X. In the present study, we used complementary genomic approaches to illuminate normal DDX3X function and the impact of cancer-associated mutations. We used CLIPseq to identify the RNA targets of endogenous DDX3X. This analysis revealed that DDX3X primarily binds mature mRNAs and, further, that the distribution of DDX3X binding sites in targets spans the full length of these mRNAs, with enrichment in the coding regions, suggesting a role for DDX3X in translation elongation. Consistent with this observation, we detected a fraction of wild type DDX3X in polysomes by sedimentation experiments. In contrast, MB-associated DDX3X mutants appear to associate poorly with polysomes, but induce spontaneous stress granule (SG) formation in cells, suggesting a role of mutant DDX3X in triggering translation silencing. In fact, the RNA binding capabilities of DDX3X are still preserved in the context of cancer-associated mutations further supporting a role of mutant DDX3X in repressing translation of specific RNAs by retaining them within SGs. Interestingly, gene ontology analyses find mRNAs encoding translation factors themselves to be highly enriched among DDX3X RNA targets, suggesting that perturbations of DDX3X functions could have far-reaching impact on cellular translation. Indeed, knocking down DDX3X with siRNA or overexpressing a mutant led to a significant impairment in global translation as measured by S35-methionine metabolic labeling. Furthermore, assessment of whole-genome protein translation by ribosomal profiling analyses in cells overexpressing cancer-associated mutant DDX3X confirmed a decreased translation rate of direct DDX3X mRNA targets in particular, but also numerous other mRNAs. Finally, we have generated an allelic series of transgenic fruit flies expressing wild type or mutant versions of human DDX3X or its Drosophila ortholog Belle, and have captured mutation-dependent phenotypes when these transgenes are expressed in specific tissues. Together, our data show that DDX3X regulates the translation of a key subset of mRNAs and thereby indirectly influences cellular translation globally, and that this function is perturbed by cancer-associated mutations. This abstract is also presented as poster C48. Citation Format: Yasmine A. Valentin-Vega, Matthew Parker, Michael Rusch, Demelza Smeeth, Nam Chul Kim, Jinghui Zhang, Richard J. Gilbertson, Joseph P. Taylor. Medulloblastoma-associated mutations in the DEAD box RNA helicase DDX3X impair protein translation. [abstract]. In: Proceedings of the Third AACR International Conference on Frontiers in Basic Cancer Research; Sep 18-22, 2013; National Harbor, MD. Philadelphia (PA): AACR; Cancer Res 2013;73(19 Suppl):Abstract nr PR11.