A Tide-Tracking Clock

Some of the most spectacular landscapes on Earth are found near the coastline: high waves crashing on steep cliffs, beautiful sand beaches, lush mangrove forests. These habitats are full of life. However, for the creatures living there, survival is not easy. Their environment is constantly and dramatically changing. The sun rises and sets daily, causing changes in light and temperature. In addition, tides rhythmically expose and submerge creatures who live on the coastline. Tides also bring waves that are followed by periods of calm. Imagine for a minute that you are a sand-hopper, a small beach crustacean who needs water to breathe oxygen through its gills. You will want to know not just when it will be day or night, but also when water will retreat or return, and when you might be hit by a wave or dragged away by currents. 

The day/night cycle lasts 24 hours and the tide cycle spans 12.4 hours. While scientists understand quite well how organisms keep track of time during the 24-hour day/night cycle, little is known about how animals track the 12.4-hour cycle of tides. We wanted to investigate how they do so.

  First, we need to discuss how organisms, including humans, keep track of the time of day.  Our life is organized around the day and night cycle. Most of us get up in the morning, eat, work or go to school, return home to eat again and socialize, and finally go to sleep. In fact, it is not just our behavior, but most of our body’s functions that are constantly adjusting with the time-of-day. These adjustments are directed by an internal mechanism that keeps track of time for us, called the circadian clock (from the Latin circa dies, which means “about a day”). The circadian rhythms of our body’s functions and behaviors are critical for our health. Disruption of the circadian clock increases health risks, including depression and cancer. Circadian clocks are also necessary for organisms in the wild to adapt to the daily cycles in their environment, such as light intensity and temperature.

  Proteins are critical for all our bodily functions and their regulation, and circadian clocks are no exception. Our circadian clock contains activator and repressor proteins that regulate each other. The activator proteins increase the amount of repressors in our cells, while the repressor proteins block the function of the activators. During the day, the activator proteins function at full strength and the amount of repressors increases. At night, the level of repressors is high. This causes the activator proteins to shut down and the amount of repressors begins to decline. The cycle starts again the next day, when the repressors have disappeared. The remarkable feature of this cycle is that once it gets going, it does not require the daily light/dark cycle to occur. For instance, if you slept deep in a cave, you would still wake up at the right time for a while. However, our internal clock does not exactly match day length. As you stay longer in the cave, your pattern of sleep and wake will thus drift out of alignment with the external day/night cycle. In real life, we are exposed to light and darkness daily, so our circadian clock is reset every day. This is why prolonged exposure to light in the evening causes our circadian clock to shift, resulting in difficulty falling asleep at night and waking up in the morning. Interestingly, the circadian clock of most animals is made up of very similar building blocks. In fact, the first circadian protein, called Period, was discovered in fruit flies. Our scientific understanding of our own circadian clock is thus based on genetic and molecular studies in this tiny insect, initiated in the late 1960s.

So then, how does our sand-hopper keep track of tides? It does so through a distinct clock called the circatidal clock. Like the circadian clock, the circatidal clock also impacts bodily functions and behavior. However, it does so within a 12.4-hour period to match the tidal cycle. Georges Bohn, a French naturalist, was the first to observe circatidal rhythms in 1903. He noticed that on beaches in northern France, a tiny, primitive creature, the Roscoff’s worm, would come out of the sand when the tide retreated, and sink back into the sand before the arrival of the next tide. The worm contains green algae, earning it the nickname: the mint-sauce worm. As a result of this cyclic movement, the sand became greenish at low tide. Bohn took these worms home, placed them in a glass vial filled with sand, and observed a green ring moving up and down the tube. The ring would be at the surface of the sand at the time of low tide, and deep in the sand at what should have been the high tide. Similar to our circadian rhythms of sleep and wake that keep going in constant darkness, the worms kept up their routine in the absence of tides, showing that it was driven by a biological clock, the circatidal clock. 

  How could the circatidal clock work? A likely possibility is that it is based on the same idea as the circadian clock: a rhythmic dance between repressors and activators. Some of those repressors and activators could even be shared by both clocks. To test this hypothesis, the best way is to modify the DNA of a marine animal so that it cannot make a circadian repressor or activator. Then, we can check if this animal has lost its circatidal rhythms. Fortunately for us, making such a modified animal is now relatively easy with a technology called CRISPR/Cas9-mediated gene editing. With this technology, we can cut DNA precisely to change a specific gene. 

We decided to cut the DNA required to make a circadian activator called Bmal1 in a species closely related to sand-hoppers: Parhyale hawaiensis. We chose this 1-cm, shrimp-like creature because it lives in environments with tides, such as mangroves or along the Hawaiian coastlines. We expected it to have circatidal rhythms of behavior, plus, it is very easy to raise in the lab. The first set of experiments we performed were aimed at determining if Parhyale hawaiensis exhibits circatidal rhythms of swimming behavior. We exposed these animals both to a 24-hour day/night cycle and to artificial tides to mimic what they experience in the wild. Every 12.4 hours, they experienced a “low tide” lasting 2 hours during which the aquarium was emptied of its water. This was followed by a 10.4-hour period of “high tide” where they were submerged in water. After 5 days, we removed the animals from their tanks and moved them into tubes filled with artificial sea water, in a dark incubator. We then used infrared beams to detect the movement of individual animals. As Bohn had observed with the mint-sauce worm 120 years ago, we observed a rhythmic behavior under constant conditions. Parhyale hawaiensis animals were highly active at times when the tide would have been high if they were still in the aquarium, and activity decreased at the time of artificial low tide. Interestingly, if these crustaceans were exposed only to a 24-hour light/dark cycle, but not to a 12.4-hour tidal cycle, their swimming behavior had a circadian rhythm, not a circatidal one. The behavior of Parhyale hawaiensis can thus be controlled either by its circadian clock or by its circatidal clock, depending on the cues the animal receives from its environment. 

 We then wanted to see how animals that could not make the key circadian activator Bmal1 behaved. As expected, after exposing these animals only to a light/dark cycle, their circadian rhythms were no longer present. Excitingly, after exposure to an artificial tidal cycle, their circatidal behavior was also absent. Therefore, Bmal1 is important not just for circadian rhythms, but it is also critical for circatidal behavior. Bmal1 is actually the very first protein to be identified as necessary for circatidal behavior, and our work opens a broad range of investigation that could uncover the molecular mechanisms of the circatidal clock. Our next objective is to test whether other circadian proteins are also necessary for circatidal behavior, or if the circatidal clock uses specific basic building blocks that are distinct from the circadian clock. 

  Why is it important to understand tidal timekeeping? First, it allows us to better understand marine animal behavior and bodily functions. This is important, as many species of fish, mollusks, and crustaceans live near the coastline, and their interactions are important for a healthy ecosystem. These animals are also important sources of food for people around the world. Understanding their rhythmic behavior, and how they adapt to their changing environment is important to preserve healthy populations and minimize disturbances to their life. This is particularly important given the current pressure of climate change, which is affecting many characteristics of tidal environments – such as water temperature, currents, and water height. These changes affect behavioral patterns and bodily functions of marine organisms.

  It is also important to understand how a clock can run on a 12.4-hour period, while another can run on a 24-hour period. We now know that the circatidal and circadian clocks share a common protein, Bmal1, but there must be important differences as well. Understanding these differences could help us learn how to intervene to adjust the period of a biological clock. This has medical implications. For example, some patients suffer from advanced or delayed sleep onset, because their circadian clock does not run at the correct pace. If we know how to adjust this clock, we might be able to correct some patients’ circadian clocks. Finally, the activity of some human genes cycle within a 12-hour period. Surprisingly, these 12-hour cycles are disrupted in patients with schizophrenia. Some recent studies have suggested a link between these human 12-hour rhythms and the 12.4-hour rhythms found in marine organisms. While more work is needed to understand how deep this connection runs, this is certainly further motivation for researchers to understand in depth how the marine circatidal clock functions. We predict that the small crustacean Parhyale hawaensis will play a central role in this effort.

Written By: Dr. Patrick Emery

Academic Editor: Chemist 

Non-Academic Editor: ICN Nurse

Original Paper

• Title: ​​Behavioral circatidal rhythms require Bmal1 in Parhyale hawaiensis

• Journal: Current Biology

• Date Published: 27 March 2023

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