Abstract:
Many behavioral and physiological processes in the fruit fly Drosophila
melanogaster show robust circadian oscillation. Some of the best studied circadian
rhythms include those in adult emergence, activity/rest, olfaction, mating and egg
laying. Unlike most other circadian rhythms egg laying rhythm is unique and
relatively less understood; it persists under constant light (LL), and also in the absence
of the ventral lateral neurons which are known to be the circadian pacemakers for
several other rhythms. Further, the expression of core clock genes such as period
(per) and timeless (tim) in the ovaries do not show any oscillation. In this thesis, I
have discussed the findings of some of my studies aimed at probing behavioural,
neuronal and genetic mechanisms underlying egg laying rhythm in Drosophila.
We began by studying egg laying rhythm under temperature cycles with the
objective of assessing whether it can synchronize to temperature cycles. We studied
egg laying behavior in temperature cycles imposed under constant darkness (DD) and
LL conditions. Results of our study suggest that temperature cycles synchronize
circadian egg laying rhythm with the phase of oviposition peak occurring close to the
onset of low temperature phase of temperature cycle. Also, the percentage
entrainment of egg laying rhythm is significantly greater under temperature cycles
compared to light/dark (LD) cycles suggesting that temperature cycle is a stronger
Zeitgeber.
Next we tested whether electrical silencing of the pigment dispersing factor
(PDF) expressing LNv has any effect on the persistence and entrainability of egg
laying rhythm. For this we genetically manipulated the electrical properties of PDFvi
expressing ventral lateral neurons (LNv) in flies by using pdf-GAL4 driver to express
ion channels dORKΔ-C1 and Kir2.1 in a tissue-specific manner, and studied its effect
on the egg laying rhythm in DD and LD. We found that while electrical silencing of
LNv neurons abolished adult emergence and activity/rest rhythms, egg laying rhythm
continued unabated under DD. However, electrical silencing of the LNv neurons
significantly lengthens the circadian periodicity of egg laying rhythm. This suggest
that although the electrical output from the LNv neurons may not be required for the
persistence of circadian egg laying rhythm in DD, it is required for maintaining the
clock periodicity close to 24 hr.
We also studied the role of mating in the regulation of egg laying rhythm to
determine whether mating patterns have any effect on circadian egg laying rhythm.
We used per0w mutants (arrhythmic for mating behaviour) and its control w (rhythmic
for mating behavior) to assay egg laying behavior. Our results indicate that presence
of rhythmic female (w) invariably enhanced the percentage of flies that showed
rhythmic egg laying behavior in DD, and percentage of flies that entrained to LD
cycles in all the male-female combinations compared to the case when arrhythmic
females (per0w) were used. This suggests that the robustness in the persistence of egg
laying rhythm in DD and its entrainability in LD is primarily driven by females.
Finally, we studied the expression of logjam (loj), a gene essential for
oviposition in Drosophila. The objective of this study was to find out whether
expression pattern of this gene oscillates in ovaries. We used female CantonS flies
and performed quantitative real-time PCR to quantify the mRNA levels of loj using
primers which amplify a region of the gene which is common to all known transcripts
of loj. Results of our studies showed that the mRNA expression of loj differs in a time
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dependent manner; its expression level is significantly greater at Zeitgeber Time 0
(ZT0) compared to ZT12. This suggests that loj may have some role to play in the
regulation of circadian egg laying rhythm in Drosophila apart from its role in
oviposition behaviour.
Based on the results of our studies we conclude that egg laying rhythm in
Drosophila is a complex circadian phenomenon whose underlying molecular
mechanisms seem to be independent of the core clock genes and the circadian neural
network that have been implicated in the regulation of other behavioural rhythms. It is
likely that circadian egg laying rhythm in Drosophila is regulated by molecular
mechanisms involving post-translational regulations of core clock genes and/or novel
molecular mechanisms involving the gene loj. The circadian pacemaker for egg laying
rhythm could be the peripheral oscillators in the ovaries.