Pilarsky, C. | Search bioRxiv

bioRxiv2020

Exploration of endogenous miRNA-200b/c activity and regulation through a functional dual fluorescence reporter

Since their discovery, microRNAs (miRNA)s have been widely studied in almost every aspect of biology and medicine, leading to the identification of important gene regulation circuits and cellular mechanisms. However, investigations are generally focused on the analysis of their downstream targets and biological functions in overexpression and knockdown approaches, while miRNAs endogenous levels and activity remain poorly understood. Here, we used the cellular plasticity-regulating process of epithelial-to-mesenchymal transition (EMT) as a model to show the efficacy of a fluorescent sensor to separate cells with distinct EMT signatures, based on miR-200b/c activity. The system was further combined with a CRISPR-Cas9 screening platform to unbiasedly identify miR-200b/c upstream regulating genes. The sensor allows to infer miRNAs fundamental biological properties, as profiling of sorted cells indicated miR-200b/c as a molecular switch between EMT differentiation and proliferation, and suggested a role for metabolic enzymes in miR-200/EMT regulation. Analysis of miRNAs endogenous levels and activity could lead to a better understanding of their biological role in physiology and disease.

molecular biology↗

bioRxiv2019

The presence of copy number variants in specific topologically associating domains has prognostic value in many cancer types

The human genome is organized into topologically associating domains (TADs), which represent contiguous regions with a higher frequency of intra-interactions as opposed to inter-interactions. TADs contribute to gene expression regulation by restricting interactions between regulatory elements, and their disruption by genomic rearrangements can result in altered gene expression and, ultimately, in cancer. Here, we provide a proof-of-principle that mutations within TADs can be used to predict the survival of cancer patients. For this purpose, we first constructed a set of 1,467 TADs representing the three-dimensional organization of genome across 24 normal human tissues. We then used Cox regression analysis to assess the prognostic value of the TADs in different cancer types, and identified a total of 35 TADs that were prognostic for at least one of nine cancer types. Interestingly, only 46% of the prognostic TADs comprised one or more genes with a known causal association with cancer. Moreover, for those TADs encompassing such a gene, the prognostic effect of the TAD was only directed related to the presence/absence of mutations in the gene in 13% of the cases. These observations indicate that the predictive power of a large proportion of the prognostic TADs is independent of whether pan-cancer genes are mutated or not. Furthermore, 34% of the 35 prognostic TADs showed strong structural perturbations in the cancer genome, which might mediate cancer development and progression. This study has important implications for the interpretation of cancer-related non-coding mutations and offer insights to new strategies for personalizing cancer medicine.

cancer biology↗