Molecular dynamics simulations disclose early stages of the photo-activation of cryptochrome 4

Daniel R. Kattnig, Claus Nielsen, Ilia A. Solov'yov

Preprint posted on May 17, 2018

Birds use a magnetic compass for orientation during their amazing migratory journeys. This study highlights the involvement of candidate regions of the putative Cry4 receptor molecule in magnetoreception related signal transduction.

Selected by Miriam Liedvogel


Birds are able to use the Earth’s magnetic field as reference cue for orientation during their amazing migratory journeys. From behavioural experiments we know what characteristics of the magnetic field the birds are using, but we still don’t understand how birds are sensing the magnetic field.

The magnetic compass in migratory birds has been shown to be light-dependent, and theoretical physicists have suggested the reaction could be based on a radical pair reaction, where the ratio of singlet and triplet transient states of a radical pair could be indicative for the orientation of that molecule within the magnetic field. Cryptochromes have been suggested as promising candidates as they are the only known photosensitve proteins in the vertebrate eye that have the potential to form radical pairs. In theory, cryptochromes (Cry) in the bird’s eye undergo a specific chemical reaction that is governed by the direction of the Earth’s magnetic field, and could provide a signal for orientation.

Cryptochromes come in different flavours and are members of a multigene family of blue light photoreceptors. In birds, four different cryptochromes have been identified (Cry1a and Cry1b that are different splicing variants, Cry2 and Cry4) – but which of them could potentially act as the magnetosensor? Cryptochrome 4 stands out as the most promising candidate and receives most attention in this skating exhibition, because it comes with specific features that distinguish it clearly from other family members.

Cry4 has been detected in the retina of several bird species and is particularly suited as (unlike the other mentioned family members) expression levels are constant throughout the day and independent of a circadian rhythm. Importantly, expression levels are higher during the migratory season compared to periods of the year when these birds do not engange in oriented long-distance flights, which clearly supports its role as putative magnetosensor. Cry4 shows high affinity to bind the photacitve flavin cofactor (FAD) and undergoes light-dependent structural changes in the C-terminal end, which are investigated focally here. The protein structure of this promising magnetoreceptor molecule remains to be solved, but a homology model for Cry4 from the European robin (ErCry4), an iconic and well described study species in the field of bird migration, has been established.

Key findings

Illustration of the secondary structure of ErCry4 with mobile sites indicated in red.

Here the authors use this homology model to simulate structural reorganisations that accompany the photoreduction of the flavin cofactor and are able to demonstrate that photo-activation of the hypothesized magnetoreceptor molecule induces large-scale conformational changes on very short timescales. Excitingly, these molecular dynamics simulations disclose early stages of the photo-activation of cryptochrome 4. Specifically, the photoreduction leads to the release of the C-terminal region, accompanied by structural rearrangements close to the binding site for the photacitve flavin cofactor binding site.

The authors highlight strengths and shortcomings of different modelling approaches and graphical representations that are used to identify common motifs of these mobile sites. Using a graph-based approach to describe the conformational dynamics of the potential magnetoreceptor protein upon photoactivation, the authors are able to show that these rearrangements appear to expose potential phosphorylation sites that have the potential to be functionally coupled to photoactivation. Although based on simulation data only so far, this provides the exciting possibility that these phosphorylation sites could have a function in modulating/deactivating the photo-transduction cascade.

The authors speculate this could be realised in a manner similar to the deactivation of rhodopsin (another retinal photopigment), where photoexcitation triggers conformational changes that lead to phosphorylation and subsequent regulation of downstream transduction steps in the visual process.

Summary and future prospects This paper reveals the identification of residual sites in the hypothesized magnetoreceptor protein Cry4 of migratory robin that undergo conformational changes triggered by photoactivation. Specifically, the authors identify the most promising sites to be located at the C-terminal tail of the protein or motifs closely coupled to that region. Importantly, these results identify candidate regions that are potentially involved in signal transduction related to magnetoreception and pave the avenue for experimentally testing these major regulators in subsequent experiments, possibly by targeted molecular manipulation of the focal sites. These exciting findings clearly call for support by experimental evidence from analogous studies on the crystal structures once available.


What I like about this work Understanding how birds perceive the Earth’s magnetic field and use this information to orient during their fascinating long migratory journeys is one of the key unresolved mysteries in sensory biology. This study capitalises on the recently published homology model of the candidate receptor molecule Cry 4 from European robins to identify key regulatory candidate regions and predict their involvement in magnetic compass orientation related signal transduction. Understanding the phenomenon of magnetoreception undoubtably needs a cross-disciplinary and highly integrative approach. This study provides a perfect example how theoretical prediction provide the necessary avenue for experimentalists and behavioural biologists to join in and design the diagnostic tests to challenge these findings in analogous studies and/or with complementary approaches.

And one question to the authors – and yes, I am asking for gut feeling here The transient radical pair is the one that is the one that is affected by and thus the one translating magnetic information into an oriented behavior. Here the the focus is on a radical pair state that is photoinduced from the dark state protein with the fully oxidised FAD cofactor. What is your feeling here – do you think this is “it”, i.e.the hero radical pair that runs for magnetoreceptor, or is it the fully reduced and then putatively reoxidise Cry4?

Tags: animal orientation, bird migration, magnetoreceptor

Posted on: 4th June 2018 , updated on: 6th June 2018

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  • Author's response

    Daniel R. Kattnig shared

    This is a delicate question as it capitalizes on a vigorously discussed but currently undecidable aspect of magnetoreception in cryptochromes. The two main competitors are undoubtedly: [FAD W+], for which magnetic field effects have been documented in vitro, and [FADH O2], which is often disguised as [FADH Z] in order to hide concerns implicated by O2‘s fast spin relaxation precluding magnetic sensitivity in weak magnetic fields. The strongest support in favor of the reoxidation hypothesis, [FADH O2], is provided by an interesting study by Wiltschko et al. using flickering light to demonstrate that the magnetosensitive process could occur in the dark [1]. While stipulated by these experiments, the [FADH O2]-hypothesis is not without inconsistencies: The spin relaxation in O2 is too fast, and, contrary to popular assumption, does not explain the Zeeman resonance effect postulated to interfere with the animals’ magnetoreceptor unless the radicals are too far apart to act as a magnetosensitive radical pair [2]. Furthermore, the sensitivity to low light suggest that the radical pair is generated in a photo-reduction process [3]. Concerning fast spin relaxation, we have recently suggested a way of how this issue could be alleviated by a more complicated reaction scheme involving three instead of two radicals [4]. However, while promising, this research into radical triads (instead of pairs) is still in its infancy. Yet, my gut feeling is that these processes could be vastly more sensitive sensors than any radical pair. In fact, in this scenario O2 and a radical derived from the (photo-)oxidation of a protein residue such as tyrosine or tryptophan could be simultaneously relevant. Thus, could both schools be right in the end? It is not utterly inconceivable.

    I expect that the resolution of the crystal structure of the avian cryptochromes, in particular Cry4, and the identification of interaction partners will eventually shed light on the question of the identity of the radical pair/triad. Furthermore, key experiments like that in [1] will have to be independently repeated and extended (e.g. by using radio frequency perturbation in various frequency bands). I expect that in vitro magnetosensitivity will be found for the photo-reduction process in Cry4 similar to that established for the cryptochrome of D. melanogaster. Any magnetic field effect on the re-oxidation is yet to be found, but is admittedly much harder to establish unequivocally. It is my feeling that this question will not be easy to resolve and we will have to resort to refined theoretical calculations and experiments to uncover the actual sensory process. As for the current work, we do not think that the [FAD W+] vs. [FADH O2] question is relevant as for DmCry similar activation patterns have been established for the photo-induced radical pairs and the chemical generated flavin semiquinone.

    In any case, I rest assured that we are far from a conclusive picture and I am looking forward to new discoveries and directions.


    1. Wiltschko, R.; Ahmad, M.; Nießner, C.; Gehring, D.; Wiltschko, W., Light-dependent magnetoreception in birds: The crucial step occurs in the dark. J. R. Soc., Interface 2016, 13 (118), 20151010.
    2. Hiscock, H. G.; Mouritsen, H.; Manolopoulos, D. E.; Hore, P. J., Disruption of magnetic compass orientation in migratory birds by radiofrequency electromagnetic fields. Biophys. J. 2017, 113, 1475-1484
    3. Vanderstraeten, J.; Gailly, P.; Malkemper, E. P., Low-Light Dependence of the Magnetic Field Effect on Cryptochromes: Possible Relevance to Plant Ecology. Frontiers in Plant Science 2018, 9.
    4. Kattnig, D. R., Radical-Pair-Based Magnetoreception Amplified by Radical Scavenging: Resilience to Spin Relaxation. J. Phys. Chem. B 2017, 121 (44), 10215-10227.

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