Fig 5. Both miR-279 and miR-996 contribute to maintenance of circadian rhythm.

(A-F) Typical activity profiles of individual flies of various mir-279/996 genotypes. Animals were entrained in 12hr-light/12hr-dark (LD) cycles for four days, and then kept in constant darkness (DD) for seven more days. In LD cycles, white bars represent the light phase (day) and black bars represent the dark phase (night). During DD cycles, grey and black bars represent the subjective day and night time, respectively. (A) mir-279[ex117] heterozygotes behave normally in that they maintain circadian activity in the dark, although the strong morning and evening activity peaks and mid-day siesta are not as well maintained. (B-C) In mir-279[ex117] homozygotes, the majority of animals gradually lost behavioral rhythmicity after transferring to constant darkness (B), but about 1/3 of animals could maintain circadian activity in constant darkness (C). Note that all mir-279[ex117/ex117] animals exhibited generally less activity than heterozygotes. The activity and circadian defects in mir-279[ex117/ex117] animals were rescued by single copies of the wild-type 16.6kb mir-279/996 transgene (D) or the 2x-mir-279-only (E) or 2x-mir-996-only (F) transgenes. (G-J) Averaged activity profiles of various mir-279/996 genotypes. (G) mir-279/996[ex15C] heterozygotes exhibit robust behavioral rhythmicity after transferring to constant darkness, but mir-279[ex117] homozygotes do not. In the mir-279/996[ex15C] homozygous background (which is normally mostly lethal by ~4 days), expression of only a single 1x-mir-279 (I) or single 1x-mir-996 (J) transgene can restore normal rhythmic behavior in constant darkness. n = ~32 for each genotype; the number of flies assayed for each genotype are indicated in Table 1.