The mammalian HPRT gene is a housekeeping gene. It is expressed constitutively at a low level in all cell types with the exception of the brain where expression is elevated. It has been shown in human brains that HPRT expression is particularly high in the basal ganglia. Total HPRT deficiency in man is the cause of Lesch-Nyhan syndrome, a severe neurological disorder. Partial HPRT deficiency leads to gouty arthritis. In the mouse it has been shown that the elevated level of HPRT activity in the brain is accompanied by an elevation in the steady state level of HPRT mRNA. The aim of the work presented in this thesis was to elucidate the mechanism of this elevation. The role of both 5' and 3' sequences has been explored. Expression of the mouse HPRT gene in cultured cells has identified promoter elements essential for a basal level of transcription. This work has been extended to animals by using gene targeting in embryonic stem cells. A HPRT mutant gene has been corrected by gene targeting, introducing a 35 bp core promoter which is capable of directing expression in cell culture. This has enabled the minimal elements of the promoter sequence to be identified which can direct brain specific elevation <i>in vivo</i>. The role of the 3' untranslated region (UTR) in determining tissue specific elevation of expression has also been investigated. The possibility that differential polyadenylation might be responsible has been excluded. Gene targeting has been used to substitute the native 3' UTR with the 3' UTR of the human growth hormone gene. This has demonstrated the potential of gene targeting as a way of manipulating the mammalian genome to study the expression of endogenous genes.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:645020 |
| Date | January 1993 |
| Creators | Costello, Patrick Sean |
| Publisher | University of Edinburgh |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://hdl.handle.net/1842/13476 |
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