# Hsp90 buffers behavioral variability by regulating Pdf transcription in clock neurons of Drosophila melanogaster

## TL;DR

The protein HSP83 in fruit flies helps regulate circadian behavior by controlling the production of a key neuropeptide, which may allow flies to adapt to stressful conditions by expanding their active time.

## Contribution

This study reveals a novel role for HSP83 in buffering behavioral variability by regulating the transcription of the circadian neuropeptide PDF in clock neurons.

## Key findings

- HSP83 mitigates inter-individual behavioral variability in a subset of clock neurons.
- Reduced HSP83 function leads to decreased levels of the circadian neuropeptide PDF.
- Hsp83 mutants show disrupted synchronized oscillations of the clock protein PERIOD.

## Abstract

Circadian rhythms are prevalent on Earth and temporally organize behaviour and physiology of organisms to occur in species-specific ‘temporal niches’. However, species differ in how strictly individuals are controlled by their circadian clock, suggesting that it may offer a selective advantage for an individual to extend its temporal niche under certain circumstances, for example during stressful environmental conditions. A potential mechanism controlling temporal niche adherence involves the evolutionary capacitor and chaperon protein HSP90, known to assist the proper folding of important signalling molecules. If HSP90 becomes rate limiting (e.g., under environmental stress) hidden genetic variation will be expressed, producing novel and potentially beneficial phenotypes for the individual. While this role of HSP90 is well established for morphological traits, we show here that it extends to regulation of temporal behavioural patterns. We show that within a small subset of clock neurons in the fly brain, HSP83, the fly homologue of HSP90, mitigates inter-individual behavioural variability. We provide evidence for the requirement of HSP83 for efficient transcription of the gene encoding the circadian neuropeptide Pigment Dispersing Factor (PDF), and for correct PDF accumulation in central clock neurons. Strikingly, Hsp83 mutants affect synchronized oscillations of the clock protein PERIOD (PER) in subsets of circadian clock neurons in the same way as flies without PDF, further supporting a role of Hsp83 in regulating Pdf. Our findings therefore provide a mechanistic explanation for HSP83 function in regulation of behavioural variability, and offer an explanation for how to restrict temporal niche extension to stressful environmental conditions.

Circadian clocks are endogenous timers that temporally organize life functions to be synchronized with external time, determined by the predictable daily light-dark and temperature cycles. This allows organisms to anticipate environmental changes, for example by regulating an animal’s activity to occur a few hours before sunrise. Constricting activity to these so called ‘temporal niches’, may help to avoid predators, or to increase foraging success, and it is generally accepted that functional and synchronized circadian clocks improve fitness. However, under extreme or rapidly changing environmental conditions (e.g., drought, excessive heat), it may be disadvantageous if an individual’s activity is constricted to a narrow temporal niche, and exploring novel niches could be beneficial. We test the idea that the chaperon protein HSP83 contributes to temporal niche constriction. HSP83 helps to properly fold proteins, even if they contain mutations that would normally lead to misfolding and altered protein function. Under environmental stress, HSP83 becomes rate-limiting, resulting in new phenotypes, because the normally hidden mutations become visible. We show that reducing HSP83 function within circadian clock neurons increases behavioural variation, indicating a relaxation of the temporal niche. Reduced HSP83 function results in decreased levels of the important circadian neuropeptide Pigment Dispersing Factor (PDF), presumably contributing to the increased behavioural variation.