A blind circadian clock in cavefish reveals that opsins mediate peripheral clock photoreception.

This paper is a tour de force! It shows that opsins can function as photoreceptors for peripheral circadian clocks in zebrafish. An imaginative comparative approach, using blind cavefish in parallel with zebrafish, was critical to nail down this important finding.

The blind cavefish Phreatichthys andruzzii, which has evolved in a lightless environment for a few million years, has lost its eyes and pigmentation. It possesses a blind, food-entrainable clock that utilizes the standard clock genes, exhibits an infradian period in constant conditions and lacks temperature compensation. Molecular studies and the use of a cavefish cell line show that its peripheral clocks, containing the typical circadian genes, also do not respond to light; peripheral clocks in zebrafish are light-sensitive. Two opsins, melanopsin and TMT-opsin, which have been implicated in peripheral light sensing in zebrafish, are mutant and non-functional in cavefish. However, the cavefish cell line can be rendered light sensitive by transfection with zebrafish versions of either opsin. Further study shows that additional peripheral photoreceptors sensitive to red light await discovery in zebrafish. The food-entrainable clock of cavefish makes it an interesting one for further evaluation, as the nature of a light-independent, food-entrainable oscillator system is poorly understood in vertebrates. This paper emphasizes the importance of probing unconventional species to help solve biological problems.

If circadian clocks are so important, what happens in species that live in a time-free environment? In a wonderfully elegant, classical 'compare and contrast' exercise, using zebrafish and a blind cavefish, Cavallari et al. show the effect upon the clock of ca. two million years spent in complete darkness, under the Somalian desert. Life in the extreme, molecules, evolution, behaviour and clocks -- what more can you ask for to whet your biological appetite?

The significance of this work is that it examines directly the consequences for circadian clock function of living and evolving in a time-free environment. Even though in vivo the blind cavefish, Phreatichthys andruzzii, can not exhibit daily rhythms of behaviour and gene expression when exposed to a 24 hours light::dark cycle, it can do so when a different time cue, daily feeding, is applied. Thus, the clock is still there and can be activated, but what the fish clearly lacks is vision -- or rather the means for an artificially applied light-dark cycle to engage the cogs of the latent clockwork. In a wonderful piece of DNA cut and pasting, clock vision is restored in cavefish cells by replacing their deficient, mutated opsins with functional sequences from zebrafish. Surprisingly, it seems as though everything about the clock downstream from its sensory input works fine. One might conclude, therefore, that the clock is so hardwired into the genome that the absence of adaptive advantage for two million years was insufficient disuse for it to become vestigial.

There is, however, an intriguing deficiency. A canonical property of circadian clocks is that they are temperature compensated, i.e. they maintain a nearly 24-hour period across a wide temperature range and so Q10 is ca. 1.0. A clock that ran faster in the heat would not keep time effectively and so, across all phyla, evolution has furnished clocks with biochemical mechanisms to keep a steady pace whatever the thermal environment might throw at them. Intriguingly, the competent clock of the cavefish, after two million years without temperature challenges, has weakened temperature compensation, such that Q10 is ca. 1.35. This slackening of the internal biochemistry is intriguing not only from an evolutionary perspective but also because it may provide a new lead into determining how temperature is compensated in more conventional species.

Overall then, the paper shows once again what a rich seam of discovery is provided by examining life in the extreme when one wishes to understand events closer to the norm -- in this case clocks and their control by light. Moreover, the old-style 'compare and contrast' approach remains the bedrock of great biology.