Comparison of chromosomal DNA composing timeless in Drosophila melanogaster and D. virilis suggests a new conserved structure for the TIMELESS protein

Two proteins, TIM and PER, physically interact to control circadian cycles of tim and per transcription in Drosophila melanogaster. In the present study the structure of TIM protein expressed by D. virilis was determined by isolation and sequence analysis of genomic DNA (gDNA) corresponding to the D. virilis tim locus (vtim). Comparison of vtim and mtim gDNA revealed high conservation of the TIM protein. This contrasts with poor sequence conservation previously observed for the TIM partner protein PER in these species. Inspection of the vtim sequence suggests an alternative structure for most TIM proteins. Sequences forming an intron in a previously characterized D. melanogaster tim cDNA appear to be most often translated to produce a longer TIM protein in both species. The N-terminal sequence of vTIM and sequence analysis of genomic DNA from several strains of D. melanogaster suggest that only one of two possible translation initiation sites found in tim mRNA is sufficient to generate circadian rhythms in D. melanogaster. TIM translation may be affected by multiple AUG codons that appear to have been conserved in sequences composing the 5'-untranslated tim mRNA leader.