The Structure and Expression of the Murine Glutathione Peroxidase Gene

Chambers, Ian Paul (1987) The Structure and Expression of the Murine Glutathione Peroxidase Gene. PhD thesis, University of Glasgow.

Abstract

The literature relating to the cell biology of the murine haemopoietic system, particularly the erythropoietic lineage has been reviewed, as have current ideas relevant to the regulation of cell-specific gene transcription. A series of experiments analysing the structure and expression of the murine gene encoding a polypeptide of approximately 19kD present in reticulocytes is then presented. These show that the gene encodes the selenium-dependent enzyme glutathione peroxidase (GSHPx) which contains the unusual amino acid selenocysteine in its polypeptide backbone. This amino acid has been shown by sequencing of the GSHPx gene and mRNA to be encoded by the "termination" codon, UGA. The significance of this finding is discussed. The GSHPx mRNA is detectable in every cell type and tissue examined, although at varying levels. Thus in erythroid cells, liver and kidney, the GSHPx mRNA level is 20-50 fold higher than in other cells. These differences have been shown not to be due to nuclear retention of the mRNA or pre-mRNA in low expressing cells. Furthermore, the levels of pre-mRNA in a low and a high expressing cell line vary in the same way as the mRNA. It therefore seems that the different levels of GSHPx mRNA arise through differences either in the stability of the GSHPx mRNA or in the rate of transcription of the GSHPx gene in different tissues. Indirect support for the involvement of the latter mechanism has come from an analysis of the chromatin structure of the GSHPx gene in tissues expressing the mRNA at a high or low level. Thus a broad DNAse I hypersensitive site is seen downstream from the gene. In addition, two erythroid specific DNAse I hypersensitive sites are also located 3' to the GSHPx gene. Functional studies, involving short term transfection assays using fragments of the GSHPx gene linked to the chloramphenicol acetyltransferase (CAT) gene, showed that the region of DNA immediately upstream of the GSHPx gene transcription initiation site functions equally well as a promoter of transcription in both a low expressing and a high expressing cell line. In addition, a DNA fragment from the 3' end of the GSHPx gene, shown by sequence analysis to contain elements homologous to cis-acting viral and cellular DNA transcription regulatory sequences, did not have any effect upon the transcription from the GSHPx or other promoters when linked in cis to them.

• Item Type: Thesis (PhD)
• Qualification Level: Doctoral
• Keywords: Genetics, Cellular biology
• Date of Award: 1987
• Depositing User: Enlighten Team
• Unique ID: glathesis:1987-77533
• Copyright: Copyright of this thesis is held by the author.
• Date Deposited: 14 Jan 2020 11:53
• Last Modified: 14 Jan 2020 11:53
• URI: https://theses.gla.ac.uk/id/eprint/77533