ONCOMIRNA-BIOGENESIS

Biogenesis of Oncogenic MicroRNAs : from the structure of the microRNA processing complexes to the inhibition of the maturation of human oncogenes

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 Obiettivo del progetto (Objective)

'Micro-RNAs are a recently identified class of small non-coding RNAs that have been shown to regulate the expression of up to 30% of human genes. The contribution of miRNAs in gene regulation is gaining considerable attention, particularly given the growing links between miRNA mis-function or expression and disease. Certain miRNAs have been demonstrated to display oncogenic-like phenotypic effects and their overexpression is heavily implicated in the development of cancers. MiRNAs mature via a two-step pathway involving two processing complexes: first Drosha-DCGR8 and second Dicer-PACT-TRBP. The two ribonucleases Drosha and Dicer are recruited by multi-domain RNA-binding accessory proteins that interact with specific, but currently unknown, structural features present in immature miRNAs. In this project I propose to perform an ambitious and comprehensive multidisciplinary analysis of biomolecular interaction networks involved in human miRNA biogenesis. An automated molecular biology approach will be employed to rapidly clone, express and screen soluble domains derived from each modular processing protein. I will use the latest fast NMR methods to rapidly assess protein and RNA folds, and protein and RNA interaction sites. These results will define the roles of each biogenesis protein, describe RNA recognition mechanisms and reveal the first atomic resolution picture of miRNA processing complexes. This structural evaluation will form the basis for the design and testing of inhibitors of miRNA biogenesis. Over-expression of oncogenic miRNAs correlates well with cancer. I propose to evaluate the possibility of using anti-sense oligonucleotides to block the regions of the immature miRNA recognized by the biogenesis machinery. The efficacy of these molecules will be explored in vivo with the view to developing innovative ways of probing miRNA function or potential new cancer therapies.'

Introduzione (Teaser)

Micro-RNAs are a class of small non-coding ribonucleic acids thought to play a role in regulating the expression of some 30/;% of human genes. There is also increasing research evidence that their misfunction is linked to disease and even cancer development.

Descrizione progetto (Article)

Micro-RNAs (miRNAs) mature in two phases along a pathway involving Drosha-DGCR8 and Dicer-PACT-TRBP processing complexes. These ribonucleases are recruited by multi-domain RNA-binding accessory proteins that interact with structural features specific to immature miRNAs.

the 'Biogenesis of oncogenic microRNAs: from the structure of the microRNA processing complexes to the inhibition of the maturation of human oncogenes' (oncoMiRNA-biogenesis) project is working on a multidisciplinary analysis of biomolecular interaction networks involved in the production processes of human miRNA. Aiming to clone, express and screen soluble domains taken from modular processing proteins, project partners will also use nuclear magnetic resonance (NMR) methods to assess protein and RNA folds, and protein-RNA interaction sites.

Experiments focus on defining the role of biogenesis proteins, describing RNA recognition mechanisms and arriving at the first atomic resolution image of miRNA processing complexes. Such an approach is expected to lay the foundation for the design and testing of inhibitors of miRNA biogenesis with a view to uncovering the means of probing miRNA function or potential new cancer therapies.

To date, the EU-funded project has identified potential domains constructs from multi-domain proteins involved in miRNA production and provided an initial design of constructs from miRNA biogenesis accessory proteins. Partners have also performed automated cloning and expression testing, achieved biophysical and NMR spectroscopy analysis of features of miRNA biogenesis processes, and optimised miRNA in vitro production.

other project accomplishments include analysis of certain inter- and intra-molecular interactions, producing mg quantities of RNA for NMR analysis and developing isotope-labelling strategies for NMR-based analysis of biomolecular interfaces.

the oncoMiRNA-biogenesis partners expect that ongoing work will result in achieving structural analysis of miRNA biogenesis machinery and mapping protein-protein interactions and protein-RNA interactions by NMR spectroscopy and other bio techniques. They also aim to develop a high-resolution model for describing critical biomolecular interfaces and interactions playing a role in miRNA biogenesis.