Cryptochrome 1 mediates light-dependent inclination magnetosensing in monarch butterflies

Abstract

Many animals use the Earth's geomagnetic field for orientation and navigation. Yet, the molecular and cellular underpinnings of the magnetic sense remain largely unknown. A biophysical model proposed that magnetoreception can be achieved through quantum effects of magnetically-sensitive radical pairs formed by the photoexcitation of cryptochrome (CRY) proteins. Studies in Drosophila are the only ones to date to have provided compelling evidence for the ultraviolet (UV)-A/blue light-sensitive type 1 CRY (CRY1) involvement in animal magnetoreception, and surprisingly extended this discovery to the light-insensitive mammalian-like type 2 CRYs (CRY2s) of both monarchs and humans. Here, we show that monarchs respond to a reversal of the inclination of the Earth's magnetic field in an UV-A/blue light and CRY1, but not CRY2, dependent manner. We further demonstrate that both antennae and eyes, which express CRY1, are magnetosensory organs. Our work argues that only light-sensitive CRYs function in animal light-dependent inclination-based magnetic sensing.

Fig. 1. A robust individual-level behavioral assay for light-dependent magnetoreception in the monarch butterfly.

a Monarchs are tethered and fixed in the center of a flight simulator surrounded by a 3D Helmholtz coil system, facing the geomagnetic South. Of the three pairs of coils (red, blue, and green), the plane of the green pair is parallel to geomagnetic North (N) and perpendicular to geomagnetic east (E). b Frontal view of the tethered monarch and the infrared beam system counting the number of wingbeats. c Irradiance curve of full-spectrum light conditions (~350–800 nm) during trials. d Left panels, North-East-down coordinate system of the ambient magnetic inclination (AMI) and the reversal of AMI (RAMI). B_F, B_H, B_Z, and I represent the geomagnetic vector, horizontal component, vertical component, and inclination of the magnetic field, respectively. Right panels, magnetic responses of wild-caught migratory (orange) and laboratory-raised (black) wild-type monarchs to a reversal of magnetic inclination (red line) under full spectrum light (white circle) (n = 32 for each group). Each box plot shows the number of wingbeats for every 10 s time bin (median, centerline; interquartile range (IQR), box; 1.5× IQR, whiskers). Each dot represents the response of an individual. Statistical significance between wild-caught migrants and laboratory-raised monarchs was tested for each time bin using a two-tailed Mann–Whitney U test at p < 0.05. No significance was found.

Fig. 2. DpCry1 is necessary for light-dependent magnetoreception in monarchs.

a Irradiance curves of full-spectrum (~350–800 nm; black line), UV-A/blue (~380–430 nm; violet line) and cyan/green (~480–580 nm; green line) light. b Magnetic responses of dpCry1^+/+ (black) and dpCry1^−/− (blue) monarchs to a reversal of magnetic inclination (RAMI; red line) under different lighting conditions (full spectrum, white circle; darkness, black circle; UV-A/blue, violet circle; cyan/green, green circle). Each box plot shows the number of wingbeats for every 10 s time bin (median, centerline; interquartile range (IQR), box; 1.5× IQR, whiskers). Each dot represents the response of an individual (n = 30 for each genotype). In each lighting condition, statistical significance between genotypes was tested for each time bin using a two-tailed Mann–Whitney U test at p < 0.05 (Full spectrum: ^**p = 0.003 for fifth 10 s time bin, ^***p = 4.757E−7 and p = 7.000E−6 for third and fourth 10 s time bins respectively; UV-A/blue: ^*p = 0.038 for fifth 10 s time bin, ^***p = 2.000E−6 and p = 2.000E−5 for third and fourth 10 s time bins, respectively).

Fig. 3. DpCry2 is dispensable for light-dependent magnetoreception in monarchs.

Magnetic responses of dpCry2^+/+ (black) and dpCry2^−/− (red) monarchs to a reversal of magnetic inclination (RAMI; red line) under different lighting conditions (full spectrum, white circle; darkness, black circle; UV-A/blue, violet circle; cyan/green, green circle). Each box plot shows the number of wingbeats for every 10 s time bin (median, centerline; interquartile range (IQR), box; 1.5× IQR, whiskers). Each dot represents the response of an individual (n = 30 and 25 for dpCry2^+/+ and dpCry2^−/−, respectively). In each lighting condition, statistical significance between genotypes was tested for each time bin using a two-tailed Mann–Whitney U test at p < 0.05. No significance was found.

Fig. 4. Monarch light-dependent magnetoreception involves both the antennae and the compound eyes.

Magnetic responses of wild-type monarchs with black (black bars) and clear (white bars) painted antennae a or eyes b to a reversal of magnetic inclination (RAMI; red line) under different lighting conditions (full spectrum, white circle; darkness, black circle; UV-A/blue, violet circle; cyan/green, green circle). c Magnetic responses of wild-type monarchs with one antenna and one eye painted black (black bars) to a RAMI (red line) under full-spectrum light. Monarchs with clear painted antennae and eyes (white bars) were used as controls. Each box plot shows the number of wingbeats for every 10 s time bin (median, centerline; interquartile range (IQR), box; 1.5× IQR, whiskers). Each dot represents the response of an individual (n = 18 for each painting group). Statistical significance between painting groups was tested for each time bin using a two-tailed Mann–Whitney U test at p < 0.05 (Full spectrum in (a): ^* p = 0.019 and p = 0.029 for third and fourth 10 s time bins respectively; UV-A/blue in (a): ^* p = 0.029 and p = 0.017 for third and fourth 10 s time bins, respectively; Full spectrum in (b): ^* p = 0.016 for fourth 10 s time bin and ^**p = 0.009 for third 10 s time bin; UV-A/blue in (b): ^* p = 0.011 and p = 0.027 for third and fourth 10 s time bins, respectively). Transcript (d) and protein (e) expression levels of dpCRY1 in antennae (Ant), compound eye photoreceptors (Eye PR), and optic lobe (OL) of adult monarchs (n = 6 per tissue in (d)). d Data are presented as mean values ± SEM.