Building a Better Gene

Researcher Dongsheng Duan says adding a piece to miniaturized dystrophin genes makes them more effective

by Quest Staff on March 15, 2009 - 9:00pm

Displacement of a protein called neuronal nitric oxide synthase (nNOS) from the membrane that surrounds each skeletal muscle fiber appears to be a much more important contributor to exercise intolerance and even cardiac degeneration in some forms of muscular dystrophy than previously recognized.

Several dystrophies – Duchennne (DMD), Becker (BMD), some types of limb-girdle, and some types of congenital MD — result from mutations in genes for proteins that are normally located in or near this membrane. Without these proteins, the membrane is often more fragile and more porous (permeable) than it would be otherwise, which is a problem for muscle function and structure.

Muscle-Fiber Membrane Proteins
Neuronal nitric oxide synthase (nNOS) needs a specific section of the dystrophin protein for its proper positioning. Researchers say adding repeats 16 and 17 to a dystrophin minigene will provide it.

Although membrane fragility and permeability are no doubt major contributors to several muscular dystrophies, scientists now believe another side effect of membrane disruption — the untethering of nNOS from its usual location near the membrane — is also significant.

Until recently, it's been thought that neuronal NOS is attached to the muscle-fiber membrane via proteins called syntrophins. However, recent studies suggest that syntrophins alone cannot accomplish the task.

A multicenter team coordinated by MDA grantee Dongsheng Duan at the University of Missouri-Columbia reported its findings on this subject in the March 2, 2009, issue of the Journal of Clinical Investigation.

The team, which also included MDA grantee Jeffrey Chamberlain at the University of Washington-Seattle, found that a specific piece of dystrophin, the protein that's missing in DMD and truncated in BMD, is essential to help tether neuronal NOS to the muscle-cell membrane.

The region of dystrophin that's required to bring neuronal NOS to the membrane is called "spectrin-like repeats 16 and 17." When neuronal NOS is properly tethered, it leads to production of the blood-vessel dilator nitric oxide, which is required for adequate blood flow to muscle during exercise.

Miniaturization of the dystrophin gene has been necessary in order to fit the very large gene into small viral shells for delivery to muscle. Unfortunately, all the miniaturized dystrophin genes now in development for gene therapy lack repeats 16 and 17.

However, Duan and his colleagues have synthesized a series of new miniaturized dystrophin genes that contain repeats 16 and 17.

"These new genes are as effective as the ones in use now when you measure muscle force," Duan said. "However, when the mice run on a treadmill, only the new genes, with repeats 16 and 17, improved performance."

The other good news is that therapies to offset the loss of nNOS, such as the blood-vessel dilator sildenafil (Viagra), have already shown promise in dystrophin-deficient mice (see Viagra Shows Promise).

In the same issue of JCI, MDA research grantee Elizabeth McNally and colleague Ahlke Heydemann, both at the University of Chicago, comment on the significance of the recent findings. They note, "As gene therapy approaches for DMD evolve, nNOS restoration mediated by dystrophin spectrin-like repeats 16 and 17 will need to be incorporated in order to correct muscle fatigue."

Duan said his group's findings "bring hope for better amelioration of Duchenne muscular dystrophy with gene therapy."

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