Research objectives and content
The existence of five highly conserved subunits that are common to all three RNA polymerases is one + the less well understood features of eukaryotic transcription. My project focusses on ABC27 (or Rpb5 is the only common subunit that is not functionally interchangeable between human and yeast. Recent evidence suggests that it communicates with transcriptional transactivators and thus directly contributes the regulation of gene expression. We propose to obtain a functional human ABC27 protein in yeast, by directed mutagenesis, by construction of chimeric proteins between human and yeast, and by direct searching for yeast suppressor mutants in which the human form of the subunit would be functional in vivo. Recent unpublished data (based on a two hybrid assay) strongly suggest that ABC27 interacts with conserved domain located on the largest subunit of the three yeast RNA polymerases. We shall examine there is a direct correlation between the ability to support growth and this interaction. This genetic a molecular approaches are meant to clarify the role played by ABC27/Rpb5 in eukariotic transcription complexes and, by extension, may help understanding the general role(s) played by the common subunits.
Training content (objective, benefit and expected impact)
the host institution is among the leading european laboratories in the field of eucaryotic transcription a yeast molecular biology. It will therefore provide high level training in a field that is not yet well develop in Spain. Working in the group of Dr. P. Thuriaux, will specifically introduce me to the molecular genetic of an eukaryotic organism, thereby completing my previous formation in the field of prokariotic molecular biology and genetics.All techniques and skill acquired will be of great interest to my future retour to Spain. Links with industry / industrial relevance (22)
The human form of RPB5 is thought to directly interact with the HBx protein of the human hepatitis B virus. Understanding the precise role of that subunit might therefore open new therapeutic perspectives for Hepatitis B, even though this is not the primary objective of our research. Similarly, the African Swine Fever Virus (a major veterinary plague) encodes an Rpb5-like protein that is in all likelyhood a component of its transcription machine. One outcome of our project will be to identify the most critical residues of ABC27, which are also present in the viral protein: this may conceivably be relevant to viral therapy.