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Genetic analysis of circadian responses to low frequency electromagnetic fields in Drosophila melanogaster.

Fedele G, Edwards MD, Bhutani S, Hares JM, Murbach M, Green EW, Dissel S, Hastings MH, Rosato E, Kyriacou CP - PLoS Genet. (2014)

Bottom Line: Most strikingly, an isolated CRY C-terminus that does not encode the Tryptophan triad nor the FAD binding domain is nevertheless able to mediate a modest EMF-induced period change.In contrast, when we examined circadian molecular cycles in wild-type mouse suprachiasmatic nuclei slices under blue light, there was no field effect.Our results are therefore not consistent with the classical Trp triad-mediated RPM and suggest that CRYs act as blue-light/EMF sensors depending on trans-acting factors that are present in particular cellular environments.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetics, University of Leicester, Leicester, United Kingdom.

ABSTRACT
The blue-light sensitive photoreceptor cryptochrome (CRY) may act as a magneto-receptor through formation of radical pairs involving a triad of tryptophans. Previous genetic analyses of behavioral responses of Drosophila to electromagnetic fields using conditioning, circadian and geotaxis assays have lent some support to the radical pair model (RPM). Here, we describe a new method that generates consistent and reliable circadian responses to electromagnetic fields that differ substantially from those already reported. We used the Schuderer apparatus to isolate Drosophila from local environmental variables, and observe extremely low frequency (3 to 50 Hz) field-induced changes in two locomotor phenotypes, circadian period and activity levels. These field-induced phenotypes are CRY- and blue-light dependent, and are correlated with enhanced CRY stability. Mutational analysis of the terminal tryptophan of the triad hypothesised to be indispensable to the electron transfer required by the RPM reveals that this residue is not necessary for field responses. We observe that deletion of the CRY C-terminus dramatically attenuates the EMF-induced period changes, whereas the N-terminus underlies the hyperactivity. Most strikingly, an isolated CRY C-terminus that does not encode the Tryptophan triad nor the FAD binding domain is nevertheless able to mediate a modest EMF-induced period change. Finally, we observe that hCRY2, but not hCRY1, transformants can detect EMFs, suggesting that hCRY2 is blue light-responsive. In contrast, when we examined circadian molecular cycles in wild-type mouse suprachiasmatic nuclei slices under blue light, there was no field effect. Our results are therefore not consistent with the classical Trp triad-mediated RPM and suggest that CRYs act as blue-light/EMF sensors depending on trans-acting factors that are present in particular cellular environments.

No MeSH data available.


Related in: MedlinePlus

EMFs increase activity levels in wild-type flies.(A–C) Hyperactivity in EMF-exposed CS under static, 50 and 3 Hz field respectively at 300 µT. (C–E) Hyperactivity in CS flies under 300, 90 and 1000 µT field respectively at 3 Hz. N's are the same as in Figure 1. Mean activity events per 30 min time bin (± sem). For average activity and N refer to Table S2 (post-hoc *p<0.05, **p<0.01, ***p<0.001).
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pgen-1004804-g004: EMFs increase activity levels in wild-type flies.(A–C) Hyperactivity in EMF-exposed CS under static, 50 and 3 Hz field respectively at 300 µT. (C–E) Hyperactivity in CS flies under 300, 90 and 1000 µT field respectively at 3 Hz. N's are the same as in Figure 1. Mean activity events per 30 min time bin (± sem). For average activity and N refer to Table S2 (post-hoc *p<0.05, **p<0.01, ***p<0.001).

Mentions: When we scrutinised further our locomotor activity records we observed that exposure to low frequency EMF not only shortened circadian period but it also caused significant hyperactivity in wild-type flies. Comparison of static to 3 and 50 Hz at 300 µT fields revealed significant Frequency (F(2,294) = 42.35, p∼0), sham/EMF F(1,294) = 6.75, p<0.01), pre-exposure/exposure (F(1,294) = 7.98, p<0.01) and pre-exposure x EMF/sham exposure interaction (F(1,294) = 7.93, p<0.001), but no significant three-way interaction (F(2,294) = 0.17, p = 0.83) illustrating that all frequencies gave a similar pattern of EMF mediated hyperactivity (Figure 4A–C, Table S2). When we compared 90, 300 and 1000 µT at 3 Hz we did not observe a significant Intensity effect (F(2,272) = 2.14, p = 0.1), but sham/EMF (F(1,272) = 4.66 p<0.05), pre-exposure/exposure (F(1,272) = 8.133, p<0.05) and pre-exposure x EMF/sham exposure interactions (F(1,2272 = 3.71, p = 0.05) were all significant (Figure 4C–E, Table S2). Post-hoc tests revealed a significant hyperactivity in EMF exposed flies compared to sham at 90 and 300 µT, but not at 1 mT, but this difference was not sufficient to generate a significant three-way interaction (F(2,272) = 0.71, p = 0.5).


Genetic analysis of circadian responses to low frequency electromagnetic fields in Drosophila melanogaster.

Fedele G, Edwards MD, Bhutani S, Hares JM, Murbach M, Green EW, Dissel S, Hastings MH, Rosato E, Kyriacou CP - PLoS Genet. (2014)

EMFs increase activity levels in wild-type flies.(A–C) Hyperactivity in EMF-exposed CS under static, 50 and 3 Hz field respectively at 300 µT. (C–E) Hyperactivity in CS flies under 300, 90 and 1000 µT field respectively at 3 Hz. N's are the same as in Figure 1. Mean activity events per 30 min time bin (± sem). For average activity and N refer to Table S2 (post-hoc *p<0.05, **p<0.01, ***p<0.001).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4256086&req=5

pgen-1004804-g004: EMFs increase activity levels in wild-type flies.(A–C) Hyperactivity in EMF-exposed CS under static, 50 and 3 Hz field respectively at 300 µT. (C–E) Hyperactivity in CS flies under 300, 90 and 1000 µT field respectively at 3 Hz. N's are the same as in Figure 1. Mean activity events per 30 min time bin (± sem). For average activity and N refer to Table S2 (post-hoc *p<0.05, **p<0.01, ***p<0.001).
Mentions: When we scrutinised further our locomotor activity records we observed that exposure to low frequency EMF not only shortened circadian period but it also caused significant hyperactivity in wild-type flies. Comparison of static to 3 and 50 Hz at 300 µT fields revealed significant Frequency (F(2,294) = 42.35, p∼0), sham/EMF F(1,294) = 6.75, p<0.01), pre-exposure/exposure (F(1,294) = 7.98, p<0.01) and pre-exposure x EMF/sham exposure interaction (F(1,294) = 7.93, p<0.001), but no significant three-way interaction (F(2,294) = 0.17, p = 0.83) illustrating that all frequencies gave a similar pattern of EMF mediated hyperactivity (Figure 4A–C, Table S2). When we compared 90, 300 and 1000 µT at 3 Hz we did not observe a significant Intensity effect (F(2,272) = 2.14, p = 0.1), but sham/EMF (F(1,272) = 4.66 p<0.05), pre-exposure/exposure (F(1,272) = 8.133, p<0.05) and pre-exposure x EMF/sham exposure interactions (F(1,2272 = 3.71, p = 0.05) were all significant (Figure 4C–E, Table S2). Post-hoc tests revealed a significant hyperactivity in EMF exposed flies compared to sham at 90 and 300 µT, but not at 1 mT, but this difference was not sufficient to generate a significant three-way interaction (F(2,272) = 0.71, p = 0.5).

Bottom Line: Most strikingly, an isolated CRY C-terminus that does not encode the Tryptophan triad nor the FAD binding domain is nevertheless able to mediate a modest EMF-induced period change.In contrast, when we examined circadian molecular cycles in wild-type mouse suprachiasmatic nuclei slices under blue light, there was no field effect.Our results are therefore not consistent with the classical Trp triad-mediated RPM and suggest that CRYs act as blue-light/EMF sensors depending on trans-acting factors that are present in particular cellular environments.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetics, University of Leicester, Leicester, United Kingdom.

ABSTRACT
The blue-light sensitive photoreceptor cryptochrome (CRY) may act as a magneto-receptor through formation of radical pairs involving a triad of tryptophans. Previous genetic analyses of behavioral responses of Drosophila to electromagnetic fields using conditioning, circadian and geotaxis assays have lent some support to the radical pair model (RPM). Here, we describe a new method that generates consistent and reliable circadian responses to electromagnetic fields that differ substantially from those already reported. We used the Schuderer apparatus to isolate Drosophila from local environmental variables, and observe extremely low frequency (3 to 50 Hz) field-induced changes in two locomotor phenotypes, circadian period and activity levels. These field-induced phenotypes are CRY- and blue-light dependent, and are correlated with enhanced CRY stability. Mutational analysis of the terminal tryptophan of the triad hypothesised to be indispensable to the electron transfer required by the RPM reveals that this residue is not necessary for field responses. We observe that deletion of the CRY C-terminus dramatically attenuates the EMF-induced period changes, whereas the N-terminus underlies the hyperactivity. Most strikingly, an isolated CRY C-terminus that does not encode the Tryptophan triad nor the FAD binding domain is nevertheless able to mediate a modest EMF-induced period change. Finally, we observe that hCRY2, but not hCRY1, transformants can detect EMFs, suggesting that hCRY2 is blue light-responsive. In contrast, when we examined circadian molecular cycles in wild-type mouse suprachiasmatic nuclei slices under blue light, there was no field effect. Our results are therefore not consistent with the classical Trp triad-mediated RPM and suggest that CRYs act as blue-light/EMF sensors depending on trans-acting factors that are present in particular cellular environments.

No MeSH data available.


Related in: MedlinePlus