The human pol II-transcribed snRNA genes; a model for gene-type specific transcription.

The genes contained in our chromosomes are composed of DNA and direct the production of all the components necessary for the cells within our bodies to survive and function correctly. This is known as 'expression' of the gene. To ensure that the appropriate products are made at the right time and in the right place, gene expression is regulated at many levels. Failure of these controls can result in disease due to the appearance of a product in the wrong place and/or at the wrong time. The first step in gene expression is to copy the DNA of a gene into RNA, which is a similar type of molecule. This process is known as transcription and often the RNA product must be modified or processed before it can be translated into protein. This processing can also be controlled to determine when and where gene products are made. We are studying the expression of a distinct set of genes, the snRNA genes, whose products help to process the RNAs that code for protein (known as messenger RNAs) and therefore perform fundamental tasks during expression of most of our genes. The snRNA genes have their own specialised expression mechanisms that are not yet fully understood. Understanding exactly how expression of snRNA genes is controlled is critical to understanding how our cells function in health and disease. For example, cancer cells need to make more of the products of snRNA genes to rapidly divide. We have developed a range of "state of the art" techniques to answer the outstanding questions about regulation of expression of these genes. The answers to our questions will also help to understand how expression of protein-coding genes is controlled.

The human snRNA genes transcribed by pol II constitute a major class of independently- and ubiquitously-transcribed genes in the human genome. Their products are critical for the correct processing of protein-coding and ribosomal gene transcripts and the genes have specialised promoters and snRNA-gene specific RNA processing elements. The role of general transcription factors, CTD phosphorylation and the Integrator complex in expression of snRNA genes has been reasonably well-characterized in the last three decades. The recent elucidation of the roles of DSIF, CTCF and NELF in expression of snRNA genes suggests how 3' end processing of snRNA transcripts is linked to transcription termination. Also, it has been shown that mRNA 3'end processing such as Pcf11 and Ssu72 are involved in termination of transcription of snRNA genes. We can now appreciate that there are many transcription factors shared by protein-coding genes and snRNA genes, although some play distinct roles in expression of the two gene types.
Despite all this new information, there are still some important aspects of the expression of pol II-transcribed snRNA genes that remain to be elucidated. For example, how is initiation of transcription of these genes so tightly coupled to 3' end formation of the transcripts? Why do snRNA genes require a specific elongation complex that is recruited by a non-coding RNA? Does the Mediator complex on the snRNA genes differ from that on protein-coding genes? And why does the Herpesevirus transactivator protein VP16 only activate transcription of protein-coding genes and not snRNA genes when activation is thought to occur through interaction with several shared members of the pre-initiation complex.
Towards a complete understanding of the mechanisms regulating transcription of the human pol II-transcribed snRNA genes we will address these questions using a range of complementary "state of the art" approaches including genomics, proteomics and CRISPR/Cas9 technology.

Who might benefit and how will they benefit?
1) the scientists directly involved will benefit, as taking the research through to a successful conclusion will result in publications and training important for career advancement and the possibility of commercialization of discoveries. For example, Dr Zaborowska will add important skills in dCas9-mediated genomic pulldown and ChIP-SICAP, bioinformatic analysis of ChIP-seq, mNET-seq and proteomic data to her current portfolio of expertise, through hands on training. Dr Zaborowska will publish the results of the research, which will greatly help her transition to an independent group leader. In addition, her scientific communication skills will be honed by presenting the results of our research to both academic audiences and the general public (e.g. Oxford's Café Scientifique and local schools). Appropriate training for both audiences will be provided by the University of Oxford Science Outreach Programme, in house Science Communication Courses and Dr Zaborowska will apply to take part in the Genetics Society "Communicating Your Science" Workshop in April 2019 to help her further develop her outreach and public engagement skills. I will benefit from the set up of new technology in the laboratory and Professor Kiss, Professor Malik and I will benefit from the knowledge gained, which will inform our future research directions. Dr Zaborowska and I will benefit from interacting closely with other top flight scientists with a primary interest in the regulation of gene expression at the level of transcription and RNA processing.
Professor Kiss and Professor Malik will benefit from being involved in studying new aspects of the function of 7SK snRNA and the role of Mediator in control of gene expression respectively, which dovetails with their own research. They will also benefit by making important new contacts in Oxford and I will benefit from making new contacts in Toulouse and New York through collaboration with these two groups.
Thus, the proposed research will underpin career advancement through the acquisition of new skills, new contacts and publications. In addition, the discoveries will benefit those scientists involved by enhancing future research possibilities.
2) scientists in the UK and further afield researching the control of gene expression at the level of transcription and co-transcriptional RNA processing will benefit as our discoveries will inform their future research strategies. In addition, UK scientists in particular will benefit as we will introduce new expertise in recently-developed cutting edge molecular biology technology to the UK.
3)the students and postdoctoral fellows in my laboratory, who will have access to a wider range of cutting edge technologies.
4) the wider scientific community will benefit from a clearer understanding of the regulation of expression of human genes. The proposal addresses the BBSRC strategic priorities: "Data driven biology", Technology development for the biosciences" and "Systems approaches to the biosciences" and will produce a wealth of new gene expression data for the wider scientific community to access and exploit.
5)Dr Zaborowska and I will continue our extensive outreach activities to ensure that the public benefits from our involvement in scientific research through science education of both lay people and science students. We will also publicise our results through the Divisional Communications Office, the University Press and Information Office, the Departmental Twitter feed, our website, Oxford's Café Scientifique and the BBSRC media office in addition to scientific publications.