The circadian clock plays a vital role in monarch butterfly (Danaus plexippus) migration by providing the timing component of time-compensated sun compass orientation, a process that is important for successful navigation. We therefore evaluated the monarch clockwork by focusing on the functions of a Drosophila-like cryptochrome (cry), designated cry1, and a vertebrate-like cry, designated cry2, that are both expressed in the butterfly and by placing these genes in the context of other relevant clock genes in vivo. We found that similar temporal patterns of clock gene expression and protein levels occur in the heads, as occur in DpN1 cells, of a monarch cell line that contains a light-driven clock. CRY1 mediates TIMELESS degradation by light in DpN1 cells, and a light-induced TIMELESS decrease occurs in putative clock cells in the pars lateralis (PL) in the brain. Moreover, monarch cry1 transgenes partially rescue both biochemical and behavioral light-input defects in cryb mutant Drosophila. CRY2 is the major transcriptional repressor of CLOCK:CYCLE-mediated transcription in DpN1 cells, and endogenous CRY2 potently inhibits transcription without involvement of PERIOD. CRY2 is co-localized with clock proteins in the PL, and there it translocates to the nucleus at the appropriate time for transcriptional repression. We also discovered CRY2-positive neural projections that oscillate in the central complex. The results define a novel, CRY-centric clock mechanism in the monarch in which CRY1 likely functions as a blue-light photoreceptor for entrainment, whereas CRY2 functions within the clockwork as the transcriptional repressor of a negative transcriptional feedback loop. Our data further suggest that CRY2 may have a dual role in the monarch butterfly's brain—as a core clock element and as an output that regulates circadian activity in the central complex, the likely site of the sun compass.
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机译:通过提供经过时间补偿的太阳罗盘定向的时间部分,该生物钟在帝王蝶(Danaus plexippus)的迁移中起着至关重要的作用,这一过程对于成功导航至关重要。因此,我们通过集中在蝶类中表达的果蝇状隐花色素(cry)(称为cry1)和脊椎动物状啼声(称为cry2)的功能来评估帝王发条,并将这些基因置于体内其他相关的时钟基因。我们发现时钟基因表达和蛋白质水平的类似时间模式出现在包含光驱动时钟的君主细胞系的头部,就像在DpN1细胞中一样。 CRY1通过DpN1细胞中的光来介导TIMELESS降解,并且光诱导的TIMELESS减少发生在大脑侧外侧(PL)的假定时钟细胞中。此外,帝王cry1转基因可以部分挽救cry b sup>突变果蝇的生化和行为光输入缺陷。 CRY2是DpN1细胞中CLOCK:CYCLE介导的转录的主要转录阻遏物,内源性CRY2可以有效地抑制转录而无需PERIOD的参与。 CRY2与PL中的时钟蛋白共定位,并在适当的时间在转录抑制位点转移到细胞核。我们还发现了在中央复合体中振荡的CRY2阳性神经投射。结果在君主中定义了一种新颖的,以CRY为中心的时钟机制,其中CRY1可能充当夹带的蓝光感光体,而CRY2在时钟内部起负转录反馈环的转录阻遏物的作用。我们的数据进一步表明,CRY2在帝王蝶的大脑中可能具有双重作用-作为核心时钟元素和调节中央复合体(太阳罗盘的可能位置)中昼夜活动的输出。
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