Long non-coding RNAs and its biological functions

About the Project

In addition to ~20,000 protein coding genes, human genome contains ~30,000 genes that encode for functional RNAs. Apart from the rRNAs and tRNAs that are involved in translation, the famous example is the Xist RNA involved in silencing of one X chromosome in females. The functions of other non-coding RNAs (ncRNAs) are less understood and require more investigation. It was discovered that some of these genes (MALAT1, HOTAIR and other) are overexpressed in cancers, which highlights their functional importance in health and diseases. Significantly, it was proposed that long non-coding RNAs (lncRNAs) shape the nuclear architecture by forming nuclear bodies that regulate gene expression (NEAT1, MALAT1, Firre, PNCTR and other). lncRNA also have many functions in cytoplasm regulating translation, protein degradation, protein trafficking and other. To study the functions of lncRNAs is very important but also challenging because the methods are limited.

The project will have several objectives. First, the development/application of a method that will allow the visualisation of the RNA in cells. This will require gene editing using CRISPR/Cas9 approach to introduce an RNA aptamer at the 3’ end of RNA that binds a fluorophore molecule. When targeted the RNA can be visualised in cells using fluorescent microscopy with high resolution.

Second, the functional significance of the RNA will be exploited by either its elimination or overexpression. These will require gene editing and evaluation of the results by the analysis of cellular genome and transcriptome.

The third objective is of the main importance too because RNA is always found in association with proteins. Therefore, identifying the proteins which bind the RNA using mass-spectrometry is necessary to uncover its function in full.

Determination of structure of lncRNAs is very challenging but can be approached as well by using cryo-electron microscopy for the visualisation of isolated complexes.

Training opportunities:

You will master the gene editing techniques which are now essential in all molecular biology research. You should learn various types of microscopies – how they work and what are the advantages and limitations. Bioinformatics will be essential to learn for the analysis of gene expression. Analysis of proteins using mass spectrometry is almost obligatory in modern days research.

Outputs:

This study will advance our understanding of gene expression in humans and will shed light on the role of lncRNAs in it. It will help to identify new therapeutical targets and use RNA as a versatile, effective and safe treatment as compared to gene alterations and modifications.