Abstract
The Period (Per) genes are central components of the mammalian circadian oscillator and form part of a feedback loop, which inhibits their own transcription. Genetic variations in the human PER2 and PER3 genes have been associated with advanced and delayed sleep phase syndrome (ASPS and DSPS, respectively) and extreme diurnal preference. These discoveries directed the focus of this thesis towards other polymorphisms within the human PER 7, 2 and 3 genes that may associate with subject groups with extreme diurnal preference or circadian rhythm sleep disorders (CRSD). The single nucleotide polymorphism (SNP) database was used to identify SNPs with potential functional relevance and a collection of pooled genomic DNA samples was used to validate these SNPs before further investigation within individual subjects with extreme diurnal preference and DSPS. This strategy identified four polymorphisms, which associated significantly with extreme diurnal preference or DSPS. A synonymous polymorphism (T2434C) in PERI was found to associate with extreme morning preference. This polymorphism was hypothesised to be in linkage disequilibrium with another polymorphism, which affects PERI gene expression at the DNA, RNA or protein level because the T2434C SNP did not produce a change in the translated amino acid sequence. A polymorphism (C111G) located in the 5'-untranslated region of PER2 was found to associate with extreme morning preference. Computer prediction indicated that the C111G SNP might affect the secondary structure of the transcript, but a reporter assay found no significant difference in mRNA transiatability between the C and G allele. Four polymorphisms (G-320T, C-319A, G-294A and a repeat) were identified in the PER3 promoter region. The G-320T and C-319A were both found to associate with DSPS and haplotypes generated from the four polymorphisms were found to have varying effects on PER3 expression in reporter assays. These findings therefore suggest that polymorphisms within the PER genes associate with defined circadian phenotypes and CRSD. The continued screening for polymorphisms within circadian clock genes is therefore essential to elucidate the molecular mechanisms underlying circadian phenotypes and CRSD in humans.