Functional analysis of a Drosophila clock gene

The period (per)gene of Drosophila encodes a fundamental part of the circadian clock. The PER protein interacts with other proteins in a negative feedback loop which includes cycling of PER and other clock proteins. These proteins subsequently act as transcriptional regulators of other proteins in the loop. It is assumed therefore that they also act at the transcriptional level to affect the production of downstream genes which then control rhythmic phenotypes such as circadian activity patterns, pupal eclosion and lovesong cycle.;The work presented here concentrates on a polymorphic '(Thr-Gly)' region of per, part of which encodes for a series of threonine-glycine (Thr-Gly) repeats. Clock genes from orders as diverse as mammals, fungi and cyanobacteria have identifiable (Thr-Gly) regions albeit encoding for smaller (Thr-Gly) repeats, therefore the possibility exists that the region has some fundamental importance to clock function.;The role of the repeat and its interaction with sequences immediately 5' to it, is investigated with D.psuedoobscural/D.melanogaster interspecific chimaeric per transgenes. These molecular manipulations reveal a 'coevolved' functional unit which consists of the repeat and the approximately 60 amino acid upstream flanking sequence. Using these interspecific chimaeric per genes also allowed the subsequent mapping of species-specific behaviour differences between D.pseudoobscura and D.melanogaster to discrete regions of the per gene. The role of the polymorphic (Thr-Gly) repeat in temperature compensation (the process whereby clock function is maintained over a range of temperatures) is investigated with a series of (Thr-Gly) invitro mutated transgenes, and the results suggest a selective explanation for the latitudinal cline in (Thr-Gly) length variations in European populations of D.melanogaster. These (Thr-Gly) variants are also shown to affect the behavioural patterns displayed in light/dark cycles.