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Advanced analysis of a cryptochrome mutation's effects on the robustness and phase of molecular cycles in isolated peripheral tissues of Drosophila.

Levine JD, Funes P, Dowse HB, Hall JC - BMC Neurosci (2002)

Bottom Line: Here, we use these tools to analyze our earlier results as well as additional data obtained using the same experimental designs.In these conditions, the cry(b) mutation significantly decreases the number of rhythmic specimens in each case except the wing.Furthermore, peak phase of luciferase-reported period and timeless expression within cry+ samples is indistinguishable in some tissues, yet significantly different in others.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biology and NSF Center for Biological Timing, Brandeis University, Waltham, MA 02454, USA. jlev@brandeis.edu

ABSTRACT

Background: Previously, we reported effects of the cry(b) mutation on circadian rhythms in period and timeless gene expression within isolated peripheral Drosophila tissues. We relied on luciferase activity driven by the respective regulatory genomic elements to provide real-time reporting of cycling gene expression. Subsequently, we developed a tool kit for the analysis of behavioral and molecular cycles. Here, we use these tools to analyze our earlier results as well as additional data obtained using the same experimental designs.

Results: Isolated antennal pairs, heads, bodies, wings and forelegs were evaluated under light-dark cycles. In these conditions, the cry(b) mutation significantly decreases the number of rhythmic specimens in each case except the wing. Moreover, among those specimens with detectable rhythmicity, mutant rhythms are significantly weaker than cry+ controls. In addition, cry(b) alters the phase of period gene expression in these tissues. Furthermore, peak phase of luciferase-reported period and timeless expression within cry+ samples is indistinguishable in some tissues, yet significantly different in others. We also analyze rhythms produced by antennal pairs in constant conditions.

Conclusions: These analyses further show that circadian clock mechanisms in Drosophila may vary in a tissue-specific manner, including how the cry gene regulates circadian gene expression.

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Circular phase analysis of luciferase expression in isolated body parts continued. All of the details of this figures are the same as in Figure 7. d) isolated wings. e.) isolated forelegs. Number of rhythmic samples for each group are given in Table 1.
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Figure 8: Circular phase analysis of luciferase expression in isolated body parts continued. All of the details of this figures are the same as in Figure 7. d) isolated wings. e.) isolated forelegs. Number of rhythmic samples for each group are given in Table 1.

Mentions: We measured the time of peak luciferase activity for each of the five daily cycles included in the monitoring and analysis of a given specimen (see Materials and Methods). A mean peak time (per day) was thereby calculated for each specimen, with such a value taken as the representative phase for that specimen. One goal of quantifying the molecular phases was to compare the normal tim-luc time-course values to the BG-luc ones for each tissue in a cry+ genetic background. For this, we used circular statistics [27,32]. Briefly, the phase of each specimen is plotted as a time point (Figures 7,8). An average vector is calculated based on the distribution of phase points around the unit circle. The angle of the vector corresponds to the mean phase for the group of points, and the magnitude of the vector represents the variability in the phase estimates [32]. The Watson-Williams-Stevens statistic was applied to evaluate whether two such vectors are significantly different from one another [32]. Significant differences were evident in the phase of these two clock genes' cyclical expression under a light-dark cycle, for all tissues except isolated antennal pairs and the heads (see Figure 7) and 8.


Advanced analysis of a cryptochrome mutation's effects on the robustness and phase of molecular cycles in isolated peripheral tissues of Drosophila.

Levine JD, Funes P, Dowse HB, Hall JC - BMC Neurosci (2002)

Circular phase analysis of luciferase expression in isolated body parts continued. All of the details of this figures are the same as in Figure 7. d) isolated wings. e.) isolated forelegs. Number of rhythmic samples for each group are given in Table 1.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC103668&req=5

Figure 8: Circular phase analysis of luciferase expression in isolated body parts continued. All of the details of this figures are the same as in Figure 7. d) isolated wings. e.) isolated forelegs. Number of rhythmic samples for each group are given in Table 1.
Mentions: We measured the time of peak luciferase activity for each of the five daily cycles included in the monitoring and analysis of a given specimen (see Materials and Methods). A mean peak time (per day) was thereby calculated for each specimen, with such a value taken as the representative phase for that specimen. One goal of quantifying the molecular phases was to compare the normal tim-luc time-course values to the BG-luc ones for each tissue in a cry+ genetic background. For this, we used circular statistics [27,32]. Briefly, the phase of each specimen is plotted as a time point (Figures 7,8). An average vector is calculated based on the distribution of phase points around the unit circle. The angle of the vector corresponds to the mean phase for the group of points, and the magnitude of the vector represents the variability in the phase estimates [32]. The Watson-Williams-Stevens statistic was applied to evaluate whether two such vectors are significantly different from one another [32]. Significant differences were evident in the phase of these two clock genes' cyclical expression under a light-dark cycle, for all tissues except isolated antennal pairs and the heads (see Figure 7) and 8.

Bottom Line: Here, we use these tools to analyze our earlier results as well as additional data obtained using the same experimental designs.In these conditions, the cry(b) mutation significantly decreases the number of rhythmic specimens in each case except the wing.Furthermore, peak phase of luciferase-reported period and timeless expression within cry+ samples is indistinguishable in some tissues, yet significantly different in others.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biology and NSF Center for Biological Timing, Brandeis University, Waltham, MA 02454, USA. jlev@brandeis.edu

ABSTRACT

Background: Previously, we reported effects of the cry(b) mutation on circadian rhythms in period and timeless gene expression within isolated peripheral Drosophila tissues. We relied on luciferase activity driven by the respective regulatory genomic elements to provide real-time reporting of cycling gene expression. Subsequently, we developed a tool kit for the analysis of behavioral and molecular cycles. Here, we use these tools to analyze our earlier results as well as additional data obtained using the same experimental designs.

Results: Isolated antennal pairs, heads, bodies, wings and forelegs were evaluated under light-dark cycles. In these conditions, the cry(b) mutation significantly decreases the number of rhythmic specimens in each case except the wing. Moreover, among those specimens with detectable rhythmicity, mutant rhythms are significantly weaker than cry+ controls. In addition, cry(b) alters the phase of period gene expression in these tissues. Furthermore, peak phase of luciferase-reported period and timeless expression within cry+ samples is indistinguishable in some tissues, yet significantly different in others. We also analyze rhythms produced by antennal pairs in constant conditions.

Conclusions: These analyses further show that circadian clock mechanisms in Drosophila may vary in a tissue-specific manner, including how the cry gene regulates circadian gene expression.

Show MeSH