Researchers from the Marine Biological Laboratory at Woods Hole and the University of Massachusetts Chan Medical School have discovered the first gene, Bmal1, that is essential for controlling circatidal behavior in the crab Parhyale hawaiensis. Animals can adapt to the rising and falling waves in coastal locations according to circadian cycles.
Published in Current Biology, the study by neurobiologists Patrick Emery, Ph.D., Joshua Rosenthal, Ph.D., and colleagues demonstrates the first molecular link between circatidal and circadian clocks, while establishing P. hawaiensis as a powerful new animal system for studying the genetics underlying circatidal rhythms.
“Biological clocks are critical for organisms including humans to optimize their physiology and adapt their behavior to environmental cycles,” said Dr. Emery, vice chair and professor of neurobiology at UMass Chan Medical School and a Marine Biological Laboratory Whitman Center investigator. “By understanding how these behaviors are genetically hardwired into organisms, we can map the sensory systems and neural circuits that impact physiology and behavior.”
Every 12.4 hours, there are tides. One of the two daily tides is brought on by the moon’s gravitational pull on Earth, and the other is brought on by the centrifugal force produced by the moon and Earth’s rotation in space. The habits of marine animals that inhabit tidal zones have evolved to cope with these abrupt transitions from dry to aquatic settings every 12.4 hours.
Over the years, the Patel lab created valuable resources in this organism, such as a sequenced genome and methods to knock-out genes using CRISPR. Although the original intention wasn’t to use Parhyale for the study of circatidal rhythms, it has turned out to be excellent for this purpose. We predict that this organism will catalyze a lot of future research in this area.
Dr. Joshua Rosenthal
Although circatidal rhythms were first noticed in the Roscoff worm (Symsagittifera roscoffensis) in the early 20th century and have been thoroughly investigated in crabs, mussels, and other marine species since the 1950s, the molecular and genetic basis of the circatidal clock and its connection to the circadian clock remain a mystery.
“The lack of an animal model amendable to genetic knockdown and transgenic manipulations has prevented scientists from definitively investigating the molecular origins of the circatidal clock and its relation to circadian clock genes,” said Emery. “Only a handful of studies about circatidal genetics exist and these are unable to either rule in or rule out a role for circadian clock genes in circatidal behaviors in animals.”
Erica Kwiatkowski, an MD/Ph.D. student in the Emery lab at UMass Chan, in collaboration with the lab of Dr. Rosenthal, senior scientist at the Marine Biological Laboratory, identified the small amphipod crustacean P. hawaiensis as a promising model.
The researchers created an artificial tidal habitat in the lab for the one-centimeter-long species using artificial saltwater that was pumped in and out of an aquarium every 12.4 hours to mimic P. hawaiensis’ natural home.
Kwiatkowski and colleagues exposed the amphipods to 10 cycles (the equivalent of five days) in the artificial tidal environment. The researchers took P. hawaiensis from the simulated tidal environment and placed it in a habitat with a constant water level once it had become accustomed to these settings.
The swimming activity of the animals in individual test tubes was monitored by infrared beams. Strikingly, every 12.4 hours, the majority of the animals (80 percent) increased their swimming activity in anticipation of high tides and then reduced activity in anticipation of low tide, even though they no longer were exposed to changing water levels. This demonstrated the existence of a circatidal clock controlling locomotor behavior in P. hawaiensis.
The lab of Marine Biological Laboratory director, Nipam Patel, Ph.D., has developed P. hawaiensis as a model organism for studying genes controlling numerous aspects of embryo development, including limb patterning.
“Over the years, the Patel lab created valuable resources in this organism, such as a sequenced genome and methods to knock-out genes using CRISPR. Although the original intention wasn’t to use Parhyale for the study of circatidal rhythms, it has turned out to be excellent for this purpose. We predict that this organism will catalyze a lot of future research in this area,” Rosenthal said.
Once the strong presence of a circatidal rhythm in P. hawaiensis was established, Kwiatkowski and colleagues used CRISPR/Cas9-guided gene knockdown to hunt for genes connected to the circatidal behavior. By knocking down individual genes, scientists can observe the effect that a lost gene has on a biological process.
Kwiatkowski and colleagues used the genes that regulate mammalian circadian rhythms as a reference to search for circatidal genes and discovered that shutting down the circadian gene Bmal1 altered P. hawaiensis behavior so that the animal no longer exhibited circatidal swimming habits. Instead, the animals exhibited arhythmic behavior unconnected to tidal flows.
“Bmal1 is a critical component for the maintenance of circatidal behavior in P. hawaiensis,” said Kwiatkowski. “This is the first evidence that a gene involved in circadian rhythms is also involved in circatidal rhythms. This establishes a molecular link between the two systems.”
Emery and colleagues will now look into the precise function Bmal1 performs in regulating circatidal behavior, as well as any potential involvement of additional genes.
