Left-right asymmetry is known to be established during early embryogenesis by a small cluster of cells termed the left-right organizer. Within this organizer, motile cilia beat rapidly to create a leftward directional flow of extracellular fluid, which is the first outward sign of a left-right difference. This early flow has been shown to be critical to the distinction of right from left. However, how this flow is sensed and translated into left-right asymmetry has been unknown.

Researchers from Massachusetts General Hospital (MGH) have published a study showing that cilia in the organizer function as the creators of the flow—they also act as sensors for the biomechanical forces exerted by the flow to shape the left-right body plan of the developing embryo. Their findings “Cilia function as calcium-mediated mechanosensors that instruct left-right asymmetry” appear in Science.

Defects in left-right asymmetry are associated with numerous human disorders, including heterotaxy syndrome (a rare condition where many organs in the body can be formed abnormally), primary ciliary dyskinesia (genetic mutations that affect the tiny hairline cilia in the lungs, nose and ears, impairing their ability to remove germs and pollutants, and allowing mucus buildup and infection), and congenital heart disease.

“The knowledge gleaned from this study not only advances our understanding of the fundamental cellular processes that govern the development of the human body, [it] may also open new avenues for the development of novel diagnostics of these disorders,” says Shiaulou Yuan, PhD, an investigator in the Cardiovascular Research center at MGH. “Additionally, this work may pave the way for targeted therapies on cilia signaling and mechanosensing to improve outcomes.”

“The breaking of bilateral symmetry in most vertebrates is critically dependent upon the motile cilia of the embryonic left-right organizer (LRO), which generate a directional fluid flow; however, it remains unclear how this flow is sensed. Here, we demonstrated that immotile LRO cilia are mechanosensors for shear force using a methodological pipeline that combines optical tweezers, light sheet microscopy, and deep learning to permit in vivo analyses in zebrafish,” write the investigators.

Mechanical manipulation of cilia was critical to the study

“Mechanical manipulation of immotile LRO cilia activated intraciliary calcium transients that required the cation channel Polycystin-2. Furthermore, mechanical force applied to LRO cilia was sufficient to rescue and reverse cardiac situs in zebrafish that lack motile cilia. Thus, LRO cilia are mechanosensitive cellular levers that convert biomechanical forces into calcium signals to instruct left-right asymmetry.”

“Nearly 25 years of work by numerous groups have shown that cilia and flow in the organizer are absolutely essential for establishing body left-right asymmetry”, notes Yuan, who is also assistant professor of medicine at Harvard Medical School, and senior author of the study. “But we haven’t had the right tools or techniques to definitively study how this all works.”

To overcome this challenge, the researchers utilized zebrafish as a model for left-right development and employed a novel optical toolkit consisting of custom-built microscopy and machine learning analysis. The team deployed optical tweezers—a biophysical tool that uses light to hold and move microscopic objects similar to a tractor beam—that enabled precise delivery of mechanical force onto cilia in an intact, living animal for the first time.

Utilizing these tools, the researchers discovered that cilia are cell-surface mechanosensors that are important for left-right asymmetry of the developing body and organs such as the heart. By applying mechanical force onto cilia in the left-right organizer of zebrafish, they showed that a subset of organizer cilia sense and translate flow forces into calcium signals that control left-right development in zebrafish.

Yuan and his colleagues continue to investigate the molecular mechanisms that govern cilia force sensing. They are also working on developing new strategies to visualize and manipulate cilia signaling, with the long-term goal of developing novel tools for the treatment of cilia-associated disorders.

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