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Home | Pregnancy Timeline | News Alerts | News Archive May 15, 2013

 
Cilium molecule capture

This is a depiction of the researchers' experimental setup. Three fluorescent molecules of various sizes are attempting to enter the cilium, where they will be trapped when they snap together with the protein anchored in the membrane of the cilium.

Credit: Yu-Chun Lin (first author), courtesy of Nature Chemical Biology

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How cilia trap proteins inside a cell: Implications for hearing, vision loss and kidney disease

Experiments at Johns Hopkins have unearthed clues about which protein signaling molecules are allowed into the hollow, hair-like "antennae" called cilia, that alert cells to critical changes in their environments.

Researchers found that the size limit for entry into celia is much greater than previously thought, allowing most of a cell's proteins into cilia. The researchers believe that the specific collection of proteins in each cilium, customized to the needs of each cell type, is determined by whether and how cilia keep proteins inside once they enter –– not which ones they allow in initially.


"According to our experiments, 90 percent of the proteins in mammalian cells should be able to fit inside cilia based on their size. But most of them have never been found [inside the cilia], so we think that most proteins do wander in at some point, but only certain ones remain inside."

Takanari Inoue, Ph.D., assistant professor of cell biology, Johns Hopkins University School of Medicine


So-called "primary cilia" are attracting intense attention in recent research. It has been confirmed that primary cilia monitori the cell's exterior environment, conveying information to the rest of the cell using an arsenal of thin, antenna-like cilium. The results of this study, which help explain how that arsenal is developed, are published online in the journal Nature Chemical Biology.


Primary cilia are found protruding from most cells in a wide variety of organisms, and defects in cilia have been implicated in everything from polycystic kidney disease to vision and hearing loss: in the kidney, they monitor the flow of urine; in the eye, they sense the wavelength of light; in cartilage, they sense pressure; and in the heart, blood flow.

No matter where they are, Inoue says, their job is to translate such mechanical forces—or, in some cases, chemical ones—into molecular signals for the cell, so that it can respond appropriately to its environment.


The signaling molecules inside cilia are tailored to the required responses. For example, some are proteins that bind to DNA to modify gene activity, letting a cell respond to environmental cues by producing more of a particular protein.

The only way into the cilial column is through a hole at its base. "A cilium forms like a short drinking straw being pushed outwards from inside an inflated balloon," says Inoue. "What we don't know is whether there is some sort of cap over the hole, regulating what goes in and out. We found that more signaling molecules can enter the straw than we thought."

Inoue: "Primary cilia are really difficult to study, because they are so small and narrow, with each cilium just 1/10,000th the volume of the rest of the cell."


Previous research in other laboratories found a fixed pore exists at the base of a cilium that restricts entry to relatively small molecules.

By developing more sensitive experimental methods, the Inoue group was able to show that molecules almost 10 times larger than previously thought could enter the cilium pore opening.


Specifically, Inoue's group engineered an anchor-like molecule that embedded itself in the membranes of cilia. On the inside end of the anchor was half of a "molecular snapper." Within the watery interior of the cell, the team placed fluorescent molecules fitted with the other half of the molecular snapper. When these molecules entered cilia, they became trapped inside where the snappers clicked together, allowing the researchers to easily take images of them.


By repeating this experiment many times, the Inoue team was able to show that molecules of increasing size were able to enter the cilia. The only difference between different sized molecules was their rate of entry: smaller molecules entering more quickly than larger ones.


Figuring out how cilia select their captives is a question for another study, says Inoue.

Other authors of the report include Yu-Chun Lin, Benjamin Lin, Siew Cheng Phua, John Jiao and Andre Levchenko of the Johns Hopkins University School of Medicine; Pawel Niewiadomski and Rajat Rohatgi of the Stanford University School of Medicine; and Hideki Nakamura and Takafumi Inoue of Waseda University.

This work was supported by grants from the National Institute of General Medical Sciences (GM092930), the National Institute of Diabetes and Digestive and Kidney Diseases (P30DK090868), the National Cancer Institute (R00CA129174), the National Institute of Neurological Disorders and Stroke (R21NS074091) and the Pew Foundation.

Original article: http://www.eurekalert.org/pub_releases/2013-05/jhm-roc050913.php