The molecular components of mammalian circadian clocks are elusive. The applicant has isolated a human gene termed RIGUI that encodes a bHLH/PAS protein 44 percent homologous to the period gene in Drosophila. The highly conserved mouse homolog (m-rigui) is expressed in a circadian pattern in the suprachiasmatic nucleus (SCN), the master regulator of circadian clocks in mammals. Circadian expression in the SCN continues in constant darkness, and a shift in the light/dark cycle evokes a proportional shift of m-rigui expression in the SCN. m-rigui transcripts also appear in a periodic pattern in Purkinje neurons, pars tuberalis, and retina, but with a timing of oscillation different from that seen in the SCN. Given the observation that the RIGUI gene has all the properties known about a core circadian component the principal investigator would like to define better its exact role in the circadian pathway.
The specific aims of this study are to: a) Identify specific brain and peripheral tissue where the various circadian oscillators reside; b) Determine whether melatonin plays a role in the expression of m-rigui in the hypophyseal pars-tuberalis; c) Investigate the response of m-rigui expression in the retina and suprachiasmatic nucleus to environmental light cues; d) Make the targeted disruption of the m-rigui locus by gene knock-out methodology and determine the effect of its loss on the circadian behavior of the animals; e) Identify genes encoding proteins that interact with RIGUI using the two-hybrid system; f) Determine the genomic structure of the human RIGUI locus and delineate its promoter and cis-acting elements. Understanding genetics and molecular basis of RIGUI function could lead to new paradigms on how genes control mammalian behavior.
