Lab rotation project description
The rotation will include learning key histological techniques including tissue preparation, histological staining and immunohistochemistry in order to start looking for proteins of interest within 1) the adult cardiovascular system, 2) the developing heart, 3) diseased versus healthy hearts and in 4) differing species. You will learn basic bioinformatics in order to pick antibodies and compare proteins between differing species. You will also learn microscopy and systematic random sampling techniques. If time permits you will also undertake western blotting in order to start quantitating these proteins of interest. You will attend group laboratory meetings and individual supervision meetings and undertake literature reviews in addition to other vital skills essential for your PhD.
Molecules, cells and organisms
School of Veterinary Medicine and Science
The presence of m6A in mRNA is a common modification across the eukaryotes, and its presence is necessary for developmental decisions, and responses to nutritional cues. Our previous work has demonstrated that the pathway for mRNA methylation cross-talks to the TOR pathway, a major, conserved signalling pathway between environmental nutritional changes and translation, in the cell.
Most metazoans, plants and yeast have two related methyltransferases, METTL3 and METTL14, of which the METTL3 homologues have been shown to be crucial for mRNA methylation. However, our unpublished work shows that domesticated chicken genome only contains the METTL14 homologue, and it appears that it is this enzyme that is required for the mRNA methylation in this case. In addition we have carried out many immunohistochemistry experiments on murine and chicken heart and vasculature from a variety of tissues exploring whether the key components of the RNA methylation complex are expressed. Based on these positive findings (which were also part of a successful Masters project) we propose that the complex plays vital roles in cardiogenesis, angiogenesis, and normal heart and blood vessel function. Therefore it could play a vital role in healthy aging and disease states.
The key objectives are to investigate whether mRNA methylation and the key pathway components are differentially expressed in relation to both abundance and localisation in 1) tissues within the cardiovascular system, 2) in the developing heart, 3) in diseased versus healthy hearts in 4) differing species.Preliminary evidence: Our collaborative team have already produced preliminary evidence in order to develop the concepts involved in this PhD. This work includes testing antibodies for immunohistochemistry and western blots on avian and mammalian cardiovascular system tissues, TLCs have been carried out to show basic expression mRNA methylation, bioinformatics and expression studies have already shown differing expression patterns within differing species, therefore this project builds upon that work and answers vital questions relating to mRNA methylation in the cardiovascular system. This work will coordinate with a programme of research being undertaken by our PhD students (Daisy Haigh and Corinne Woodcock) and compliments the work of a recently recruited NRF (Dr Nathan Archer), in addition to collaborations with Biosciences and two pathologists (Dr Simone de Brot and Dr Llorenc Grau) who collected some samples towards this project.
References:1. Nature. 2016 Dec 8;540(7632):301-304. doi: 10.1038/nature205772. New Phytol. 2017 Jul;215(1):157-172. doi: 10.1111/nph.14586
Triangle Project with:
Catrin Rutland - firstname.lastname@example.org - School of Veterinary Medicine and Science
Nigel Mongan - email@example.com - School of Veterinary Medicine and Science
Rupert Fray - firstname.lastname@example.org - School of Biosciences
Nathan Archer - email@example.com - School of Veterinary Medicine and Science