Context-defined cancer co-dependency mapping identifies a functional interplay between PRC2 and MLL-MEN1 complex in lymphoma.

Interplay between chromatin-associated complexes and modifications critically contribute to the partitioning of epigenome into stable and functionally distinct domains. Yet there is a lack of systematic identification of chromatin crosstalk mechanisms, limiting our understanding of the dynamic transition between chromatin states during development and disease. Here we perform co-dependency mapping of genes using CRISPR-Cas9-mediated fitness screens in pan-cancer cell lines to quantify gene-gene functional relationships. We identify 145 co-dependency modules and further define the molecular context underlying the essentiality of these modules by incorporating mutational, epigenome, gene expression and drug sensitivity profiles of cell lines. These analyses assign new protein complex composition and function, and predict new functional interactions, including an unexpected co-dependency between two transcriptionally counteracting chromatin complexes - polycomb repressive complex 2 (PRC2) and MLL-MEN1 complex. We show that PRC2-mediated H3K27 tri-methylation regulates the genome-wide distribution of MLL1 and MEN1. In lymphoma cells with EZH2 gain-of-function mutations, the re-localization of MLL-MEN1 complex drives oncogenic gene expression and results in a hypersensitivity to pharmacologic inhibition of MEN1. Together, our findings provide a resource for discovery of trans-regulatory interactions as mechanisms of chromatin regulation and potential targets of synthetic lethality.

Agnostic Framework for the Classification/Identification of Organisms Based on RNA Post-Transcriptional Modifications.

We propose a novel approach for building a classification/identification framework based on the full complement of RNA post-transcriptional modifications (rPTMs) expressed by an organism at basal conditions. The approach relies on advanced mass spectrometry techniques to characterize the products of exonuclease digestion of total RNA extracts. Sample profiles comprising identities and relative abundances of all detected rPTM were used to train and test the capabilities of different machine learning (ML) algorithms. Each algorithm proved capable of identifying rigorous decision rules for differentiating closely related classes and correctly assigning unlabeled samples. The ML classifiers resolved different members of the Enterobacteriaceae family, alternative Escherichia coli serotypes, a series of Saccharomyces cerevisiae knockout mutants, and primary cells of the Homo sapiens central nervous system, which shared very similar genetic backgrounds. The excellent levels of accuracy and resolving power achieved by training on a limited number of classes were successfully replicated when the number of classes was significantly increased to escalate complexity. A dendrogram generated from ML-curated data exhibited a hierarchical organization that closely resembled those afforded by established taxonomic systems. Finer clustering patterns revealed the extensive effects induced by the deletion of a single pivotal gene. This information provided a putative roadmap for exploring the roles of rPTMs in their respective regulatory networks, which will be essential to decipher the epitranscriptomics code. The ubiquitous presence of RNA in virtually all living organisms promises to enable the broadest possible range of applications, with significant implications in the diagnosis of RNA-related diseases.

Cancer-Targeted Controlled Delivery of Chemotherapeutic Anthracycline Derivatives Using Apoferritin Nanocage Carriers.

The interactions of chemotherapeutic drugs with nanocage protein apoferritin (APO) are the key features in the effective encapsulation and release of highly toxic drugs in APO-based controlled drug delivery systems. The encapsulation enables mitigating the drugs' side effects, collateral damage to healthy cells, and adverse immune reactions. Herein, the interactions of anthracycline drugs with APO were studied to assess the effect of drug lipophilicity on their encapsulation excess n and in vitro activity. Anthracycline drugs, including doxorubicin (DOX), epirubicin (EPI), daunorubicin (DAU), and idarubicin (IDA), with lipophilicity P from 0.8 to 15, were investigated. We have found that in addition to hydrogen-bonded supramolecular ensemble formation with n = 24, there are two other competing contributions that enable increasing n under strong polar interactions (APO(DOX)) or under strong hydrophobic interactions (APO(IDA) of the highest efficacy). The encapsulation/release processes were investigated using UV-Vis, fluorescence, circular dichroism, and FTIR spectroscopies. The in vitro cytotoxicity/growth inhibition tests and flow cytometry corroborate high apoptotic activity of APO(drugs) against targeted MDA-MB-231 adenocarcinoma and HeLa cells, and low activity against healthy MCF10A cells, demonstrating targeting ability of nanodrugs. A model for molecular interactions between anthracyclines and APO nanocarriers was developed, and the relationships derived compared with experimental results.

Global Epitranscriptomics Profiling of RNA Post-Transcriptional Modifications as an Effective Tool for Investigating the Epitranscriptomics of Stress Response.

The simultaneous detection of all the post-transcriptional modifications (PTMs) that decorate cellular RNA can provide comprehensive information on the effects of changing environmental conditions on the entire epitranscriptome. To capture this type of information, we performed the analysis of ribonucleotide mixtures produced by hydrolysis of total RNA extracts from S. cerevisiae that was grown under hyperosmotic and heat shock conditions. Their global PTM profiles clearly indicated that the cellular responses to these types of stresses involved profound changes in the production of specific PTMs. The observed changes involved not only up-/down-regulation of typical PTMs, but also the outright induction of new ones that were absent under normal conditions, or the elimination of others that were normally present. Pointing toward the broad involvement of different classes of RNAs, many of the newly observed PTMs differed from those engaged in the known tRNA-based mechanism of translational recoding, which is induced by oxidative stress. Some of the expression effects were stress-specific, whereas others were not, thus suggesting that RNA PTMs may perform multifaceted activities in stress response, which are subjected to distinctive regulatory pathways. To explore their signaling networks, we implemented a strategy based on the systematic deletion of genes that connect established response genes with PTM biogenetic enzymes in a putative interactomic map. The results clearly identified PTMs that were under direct HOG control, a well-known protein kinase pathway involved in stress response in eukaryotes. Activation of this signaling pathway has been shown to result in the stabilization of numerous mRNAs and the induction of selected lncRNAs involved in chromatin remodeling. The fact that PTMs are capable of altering the activity of the parent RNAs suggest their possible participation in feedback mechanisms aimed at modulating the regulatory functions of such RNAs. This tantalizing hypothesis will be the object of future studies.