Dr Tobias von der Haar
Reader in Systems Biology/Director of Research
School of Biosciences
- 01227 (82)3535
1995 - Undergraduate Studies, University of Bielefeld, Germany
1998 - PhD, jointly at the German National Biotechnology Centre (GBF), Braunschweig, Germany; and at UMIST, Manchester, UK.
Postdoctoral Work at UMIST, Manchester
Postdoctoral Work at the University of Kent
Wellcome Trust Research Career Development Fellowship at the University of Kent
2009-present Lecturer in Systems Biology, University of Kentback to top
Also view these in the Kent Academic Repository
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Regulation of gene expression at a translational level
The processes that allow a cell to live rely on a particular mixture of proteins, the proteome, being present in the cell at any given time. The individual proteins making up the proteome are constantly diluted by cell divisions and lost because of protein turnover. In order to maintain a functional proteome and stay alive, cells must therefore constantly produce new proteins, with required rates of synthesis that may differ significantly between gene products. Moreover, the synthesis of an entirely different set of proteins may be required when environmental conditions change.
Eukaryotic gene expression relies on an ordered sequence of molecular events: First, a stretch of DNA containing a gene is transcribed into a messenger RNA molecule, this is then processed to its final form and exported from the nucleus to the cytoplasm. Ribosomes bind to the cytoplasmic mRNA with the help of translation initiation factors, and then (with the help of translation elongation and termination factors and tRNAs) assemble the protein from amino acids according to the genetic information encoded in the mRNA. The mRNA itself is destroyed after a short while, typically after about 1000 proteins have been produced from it.
If the rate of production of one particular gene product needs to be altered by the cell, any of the steps of gene expression can be targeted: for example, the rate of transcription may be adjusted, or the proportion of RNAs that are exported from the nucleus, or the rate with which ribosomes produce protein from the cytoplasmic mRNA. We are particularly interested in the latter form of control; the production of varying amounts of protein from the same amount of mRNA.
The motivation for our interest in this question comes from the need to understand in detail the mechanism by which cells live, develop and adapt to changing environmental conditions, in order to understand what goes wrong when diseases prevent cells from achieving these feats.
The molecular mechanism of translation termination
Translation termination is the last step in the overall process of translation, in which the newly synthesized protein is released from the ribosome:mRNA complex. It occurs when any one of the three stop codons UAA, UAG or UGA enter the decoding centre in the ribosomal A-site. Unlike sense codons, stop codons are not decoded by transfer RNAS but are instead acted on by the translation release factors (RFs in bacteria, or eRFs in eukaryotes).
A currently established minimal model for the mechanism of translation termination includes the following steps:
- A ternary complex forms between the two types of release factor (eRF1 and eRF3) and GTP.
- The ternary complex enters the ribosomal A-site.
- If the codon in the A-site is a stop codon, the GTP in the complex is hydrolysed to GDP, resulting in a conformational re-arrangement of the release factors and/ or the ribosome, severing of the P-site tRNA-peptidyl bond, and release of the newly translated peptide from the ribosome.
This relatively simple model incorporates many of the published data that address the process of translation termination. However, in yeast mutations in many genes apart from the two eRF-encoding genes affect translation termination efficiency, and it is currently not clear how the proteins encoded by those genes are involved in the termination reaction. Conversely, mutations that affect translation termination almost always also affect the stability of mRNAs, consistent with known interactions between the two processes of termination and mRNA turnover. Again, it is not clear at a molecular level how these processes interact
We are using general molecular biology techniques to investigate both in vivo and in vitro the molecular details of the mechanism of translation termination.
Cellular consequences of impaired translation termination
Efficient translation termination is essential for the production of all proteins. However, because different stop codons have different termination efficiencies depending on their sequence context, reductions in translation termination efficiency can affect the production of one protein much more than the production of another. Relatively small reductions in translation termination efficiency will therefore have significant impact on the composition of the proteome. Moreover, because of the close connection between translation termination and mRNA turnover, this may be exacerbated by additional differential effects on mRNA stability.
Small imbalances in the proteome are difficult to measure directly, but may become clearly evident through cellular phenotypes: for example, a small reduction in the activity of a factor involved in the adaptation to osmotic stress may not be easily visible at the protein level, but may become obvious because the cells can adopt less well to high osmolarity conditions. We are using this approach of looking for translation termination-factor dependent phenotypes to identify processes that may be specifically targeted when translation termination activity becomes impaired, and to search for potential novel, non-translational roles of the termination factors in the cell.
Mathematical models of translation termination in vivo
Interactions between translation termination, mRNA decay and the composition of the proteome are very complex and only poorly understood. This complexity makes it currently impossible to predict the effect that changes in translation termination efficiency have on any particular protein. In order to fully understand this important part of eukaryotic biology, it would be desirable to have quantitative computer models available that can predict such effects exactly.
We are currently still a long way from being able to develop such models. While computational procedures and mathematical frameworks exist that would allow us to develop and handle such models, the main remaining problem is the availability of sufficiently accurate quantitative data from in vivo studies. We are currently developing experimental approaches with the aim of improving the generation of quantitative datasets, and are also starting initial modelling exercises that are mainly aimed at providing estimates of the general translational activity in the cell. These initial modelling approaches will also allow us to more easily identify gaps in the existing datasets that need addressing before accurate models of the process of translation termination can be generated.
Funding from: Wellcome Trust
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- BI518 Molecular Biology and Genetics