The eye disorder CFEOM3
The nerve fibers responsible for eyelid opening and eye movement do not navigate properly, resulting in incomplete and/or erroneous connections to the eye muscles--and ultimately, droopy eyelids and restricted eye movement. Left, normal eye. Right, CFEOM3 eye. (Adapted from Tischfield MA et al. 2010 Jan 8; Cell 140(1): 74-87.)
by Michelle Pflumm, PhD
A gene whose mutation leads to droopy eyelids and restricted eye movement turns out to have a lot to do with how the nervous system as a whole establishes and maintains nerve connections. And it works in an unusual way, essentially washing out a highway that critical proteins in nerve cells, or neurons, need to travel.
The gene, called TUBB3, was identified as the critical gene altered in the rare eye disease CFEOM3 in the lab of Elizabeth Engle, MD, professor of Neurology and Ophthalmology at Boston Children's Hospital. The so-called tubulin protein encoded by TUBB3 is active only in neurons, and is a critical component of microtubules, hollow tubes that structurally support the cell and serve as cellular highways. Kinesins travel these highways, carrying vital cargo--including proteins critical for axon growth and guidance. Without these proteins, nerve fibers cannot navigate to their proper targets in the developing nervous system.
Engle's international, multi-institutional team pinpointed eight independent TUBB3 mutations associated with CFEOM3. Some patients with these mutations also had varying degrees and combinations of intellectual, behavioral and social deficits, facial paralysis and progressive motor and sensory nerve degeneration.
"The human disease phenotype really depends on where the mutation is in the microtubule," says Engle. "That is what is so surprising."
Looking at the impact of these mutations in mice, Engle's team found evidence that TUBB3 plays a critical role in axon guidance. The nerve fibers in the brains of TUBB3 mutant mice failed to grow toward their intended targets.
TUBB3 mutations result in defects in axon pathfinding
Above, a slice of a mouse brain containing either normal or altered TUBB3 protein. In the TUBB3 mutant mice, nerve fibers (denoted by the arrows) do not extend beyond the midline to either side of the brain. Image courtesy of graduate student and first author Max Tischfield. (Adapted from Tischfield MA et al. 2010 Jan 8; Cell 140(1): 74-87.
Taking a closer look at the affected microtubules, the researchers found that some of the TUBB3 mutations specifically knocked out the ability of kinesins to hold onto the microtubules, rendering them unable to deliver their cargo.
"The kinesins just so happen to 'walk' specifically on two of the residues mutated in these disorders," explains Engle, "exactly where their feet go down onto the microtubule."
TUBB3 mutations slow certain traffic on microtubules to a crawl
Kinesins are unable to grab onto TUBB3-altered microtubules, rendering them unable to deliver vital cargo (purple and yellow dots), including proteins needed for axon pathfinding. (Adapted from Tischfield MA et al. 2010 Jan 8; Cell 140(1): 74-87.)
In addition, the researchers found that these structures were unable to rapidly grow and shrink--and therefore unable to rapidly respond to external cues. This is critical for axon pathfinding.
Now, in collaboration with Judith Steen PhD, of the Department of Neurobiology, Engle's team is using quantitative proteomics techniques to identify other critical components that are missing on TUBB3 microtubules. Engle hopes the identified proteins will provide more insight into the signaling mechanisms necessary for axon guidance and help explain the varied symptoms in CFEOM3 patients with different TUBB3 mutations.
Two other rare brain diseases were recently traced to altered forms of other neuronal types of tubulin. Collectively, these studies suggest that different kinds of microtubules work together to wire the brain properly. By understanding their differences, we can understand how our brains develop and ultimately, how we can most effectively treat these diseases.
"The more we understand the specific disorders," says Engle, "the easier it will be to provide targeted therapy."