Ionotropic receptors (IRs) contribute to temperature synchronization in Drosophila melanogaster.
Abstract
Like most organisms, Drosophila melanogaster can synchronize its
physiological and behavioural processes by possessing internal circadian clock
that regulates. Naturally fluctuating timing cues, like light and temperature
(also known as Zeitgebers), synchronize these endogenous and self‐sustained
clocks with external time. In Drosophila, synchronization of the circadian clock
by light has been studied in detail, but much less is known about the molecular
mechanisms underlying temperature entrainment.
Previous data from our lab shows that Nocte, a Chordotonal organ (Ch organ)
located protein, is required for normal temperature entrainment in Drosophila.
However, neither the function of Nocte in temperature entrainment nor the
molecular underlying mechanisms are clear. To address these issues, a
proteomics strategy of combing co‐immunoprecipitation and MS/MS
sequencing was applied to isolate potential interactors of Nocte. IR25a was
one of the most promising candidates, which was later confirmed by
behavioural tests using RNA interference: Reducing IR25a expression in Chorgan resulted in abnormal behaviour during temperature cycles, similar to
what had been described for Nocte mutant.
To further confirm the interaction between Nocte and IR25a, I showed that
IR25a physically interacts with Nocte in vivo. Moreover, using an IR25a‐gal4
line, I was able to show that IR25a is expressed in subsets of chordotonal
organs (Ch organ) including Johnston’s Organs (JO), where Nocte is also highly
expressed. These results, along with the behavioural data mentioned above
are consistent with the proteomics results and suggest that Nocte and IR25a
physically and functionally interact.IR25a mutants were employed to further investigate the function of IR25a in
temperature entrainment. First of all, I found that both central and peripheral
clocks in wild type flies can be synchronized to temperature cycles with only
two degree differences (12h: 12h, 27 °C: 25 °C). In contrast, synchronization of
locomotor activity rhythms in the IR25a null mutants to the same temperature
cycles and other TC’s with 2°C amplitude was eliminated. Under the same
conditions, the oscillations of the core clock proteins TIMLESS (TIM) and
PERIOD (PER) that normally occur in fly heads were completely abolished inIR25a null mutants, suggesting that IR25a is required for temperature
entrainment of peripheral clocks. In the central brain pacemaker neurons, the
oscillations of TIM in dorsal and lateral neurons were also affected by the
IR25a mutants. On the contrary, IR25a is not required for light entrainment
and temperature compensation, suggesting that IR25a is specifically involved
in temperature synchronization. Moreover, temperature entrainment of the
IR25a null mutants can be partially restored by applying larger temperature
intervals (29°C: 25°C) indicating that IR25amay function as amplitude detector
independent of absolute temperature values. Finally, neuronal activity in
IR25a+ neurons is crucial for the synchronization of circadian clocks to low
amplitude temperature cycles.Re‐constitution of functional olfactory receptors required the assembly of
IR25a with IR76a and IR76b. Interestingly, IR76a and IR76b are neither
required for temperature entrainment at the behavioural level nor expressed
in the Ch organs. To check if other potential IRs interacting with IR25a exist, I
screened the expression pattern of most divergent IRs using IR‐gal4/UAS‐GFP
flies. IR56a was isolated as a potential partner of IR25a because it is also
expressed in the femur chordotonal organs. To investigate the function of
IR56a in temperature entrainment, I generated a null mutant of IR56a.Surprisingly, this gene is not required for synchronizing clocks to a temperature
cycle (27°C: 25°C) at the behavioural level. However, the behavioural and
molecular phenotypes of IR56a mutant under different temperature cycles
need to be further characterized.
Authors
Chen, ChenghaoCollections
- Theses [4223]