Xu Y, Padiath QS, Shapiro RE, Jones CR, Wu SC, Saigoh N, Saigoh K, Ptácek LJ, Fu YH. Nature. 2005 Mar 31;434(7033):640-4.
To discover the genetic causes of Familial Advanced Sleep Phase Syndrome (FASPS), the authors interviewed a family whose members suffer from FASPS. The authors found an A to G change in the csk1d gene that lead to a threonine to alanine mutation, by genetic screening of family members. The affected residue (amino acid 44) is conserved across species, and its mutation implies a role for csnk1d in the circadian clock.
The authors then assessed the function of the mutated protein by expressing it in bacteria. The mutant protein had a maximum reaction rate 60% that of the wildtype protein, as well as a 12% reduction in its Michaelis constant, as assessed by kinase assays.
The mutation was then introduced to fruit flies, and resulted in a slightly longer circadian period, as measured from periodogram analysis of locomotor activity.
Additionally, transgenic mice were generated via bacterial artificial chromosome cloning, and validated using RT-PCR with human primers. The authors found that the mice with the mutant csnk1d had a circadian period shorter by an estimated 24 minutes, and took longer to re-entrain to an LD 12:12 light cycle, compared to wild type mice.