A Blog Around The Clock

I had no time to read this in detail and write a really decent overview here, perhaps I will do it later, but for now, here are the links and key excerpts from a pair of exciting new papers in PLoS Biology and PLoS ONE, which describe the patterns of expression of a second type of cryptochrome gene in Monarch butterflies.

This cryptochrome (Cry) is more similar to the vertebrate Cry than the insect Cry, also present in this butterfly. The temporal and spatial patterns of expression of the two types of Cry suggest that they may be involved in the transfer of time-information from the circadian clock to the brain center involved in spatial orientation during long-distance migration.

The PLoS Biology paper looks at these patterns of expression, while the PLoS ONE paper identifies a whole host of genes potentially implicated in migratory behavior, including the Cry2. Here is the PLoS Biology paper:

Cryptochromes Define a Novel Circadian Clock Mechanism in Monarch Butterflies That May Underlie Sun Compass Navigation:

During their spectacular fall migration, eastern North American monarch butterflies (Danaus plexippus) use a time-compensated sun compass to help them navigate to their overwintering sites in central Mexico. The circadian clock plays a critical role in monarch butterfly migration by providing the timing component to time-compensated sun compass orientation. Here we characterize a novel molecular clock mechanism in monarchs by focusing on the functions of two CRYPTOCHROME (CRY) proteins. In the monarch clock, CRY1, a Drosophila-like protein, functions as a blue-light photoreceptor for photic entrainment, whereas CRY2, a vertebrate-like protein, functions within the clockwork as the major transcriptional repressor of the self-sustaining feedback loop. An oscillating CRY2-positive neural pathway was also discovered in the monarch brain that may communicate circadian information directly from the circadian clock to the central complex, which is the likely site of the sun compass. The monarch clock may be the prototype of a clock mechanism shared by other invertebrates that express both CRY proteins, and its elucidation will help crack the code of sun compass orientation.

Here is the editorial synopsis:

In Monarchs, Cry2 Is King of the Clock:

Back in the brain, the authors showed that Cry2 was also found in a few dozen cells in brain regions previously linked to time-keeping in the butterfly, and this Cry2 underwent circadian oscillation in these cells, but not in many other cells that were not involved in time keeping. By taking samples periodically over many hours, they found that nuclear localization of Cry2 coincided with maximal transcriptional repression of the clockwork, in keeping with its central role of regulating the feedback cycle. This is a novel demonstration of nuclear translocation of a clock protein outside flies.

Finally, the authors investigated Cry2's activity in the central complex, the brain structure that is believed to house the navigational compass of the monarch. Monarchs integrate information on the position of the sun and the direction of polarized light to find their way from all over North America to the Mexican highlands, where they spend the winter. Cry2, but not the other clock proteins, was detected in parts of the central complex where it undergoes strong circadian cycling. Some cells containing Cry2 linked up with the clock cells, while others projected toward the optic lobe and elsewhere in the brain.

Along with highlighting the central importance of Cry2 in the inner workings of the monarch's clock, the results in this study suggest that part of the remarkable navigational ability of the butterfly relies on its ability to integrate temporal information from the clock with spatial information from its visual system. This allows the monarch to correct its course as light shifts across the sky over the course of the day. Other cues used for charting its path remain to be elucidated.

This is the PLoS ONE paper:

Chasing Migration Genes: A Brain Expressed Sequence Tag Resource for Summer and Migratory Monarch Butterflies (Danaus plexippus):

North American monarch butterflies (Danaus plexippus) undergo a spectacular fall migration. In contrast to summer butterflies, migrants are juvenile hormone (JH) deficient, which leads to reproductive diapause and increased longevity. Migrants also utilize time-compensated sun compass orientation to help them navigate to their overwintering grounds. Here, we describe a brain expressed sequence tag (EST) resource to identify genes involved in migratory behaviors. A brain EST library was constructed from summer and migrating butterflies. Of 9,484 unique sequences, 6068 had positive hits with the non-redundant protein database; the EST database likely represents ~52% of the gene-encoding potential of the monarch genome. The brain transcriptome was cataloged using Gene Ontology and compared to Drosophila. Monarch genes were well represented, including those implicated in behavior. Three genes involved in increased JH activity (allatotropin, juvenile hormone acid methyltransfersase, and takeout) were upregulated in summer butterflies, compared to migrants. The locomotion-relevant turtle gene was marginally upregulated in migrants, while the foraging and single-minded genes were not differentially regulated. Many of the genes important for the monarch circadian clock mechanism (involved in sun compass orientation) were in the EST resource, including the newly identified cryptochrome 2. The EST database also revealed a novel Na+/K+ ATPase allele predicted to be more resistant to the toxic effects of milkweed than that reported previously. Potential genetic markers were identified from 3,486 EST contigs and included 1599 double-hit single nucleotide polymorphisms (SNPs) and 98 microsatellite polymorphisms. These data provide a template of the brain transcriptome for the monarch butterfly. Our "snap-shot" analysis of the differential regulation of candidate genes between summer and migratory butterflies suggests that unbiased, comprehensive transcriptional profiling will inform the molecular basis of migration. The identified SNPs and microsatellite polymorphisms can be used as genetic markers to address questions of population and subspecies structure.

Here is an article written after the press release, which, as such articles usually do, greatly overstates the extent of the findings:

Clocking monarch migration:

In previous work, Reppert and his team showed that pigment-producing genes in the monarch eye communicate with the butterfly's circadian clock. As part of the new study, Reppert and his team also found, in an area of the monarch brain called the central complex, a definitive molecular and cellular link between the circadian clock and the monarch's ability to navigate using the sun. Briscoe said that Reppert's study was "really going to overturn a lot of views we had about the specific components of circadian clocks."

The spatial and temporal patterns of expression make Cry2 the most serious candidate for the connection between the clock and the Sun-compass orientation mechanism. Much work, both at the molecular and at higher levels of organization needs to be done to figure out the exact mechanism by which this animal, during migration, compensates for the Sun's movement across the sky during the day, and thus does not stray off course. Cry2 appears to be a good molecular "handle" for such studies.

For background, see my older post on the initial discovery of Cry2 in Monarch butterflies by the same team.