Mitochondrial Mess

New work by a host of Center scientists gives the cell bodies a key role in ALS damage.

Mitochondria in trouble: Arrows show injured mitochondria caught within a large vacuole of their own making.

Like finding out an old bowl you’d been tossing your house keys into for years is really a valuable antique, so it is, often, with discoveries that turn into therapy. They pop up when scientists focus on something long in the background—when they begin to see the always-there in a different way.

Recently, Packard Center work on mitochondria has taken on that feel. A few years ago, nobody paid any mind—ALS-wise—to the tiny cell structures. “Journal editors would say ‘Your work is fine, but there’s no interest,'” shrugs Center researcher Zuoshang Xu. Now scientists believe mitochondria lie at the heart of what actually kills cells in the disease.

As discovery on the organelles builds on discovery—Center researchers are among the most-published in the field—you can feel the tension as their finds begin to fit together. “Everybody’s work here is making sense of this,” says Don Cleveland, a Packard scientist at the University of California. “Not a result has gone to waste.”

Moreover, studies linking the course of ALS with what goes wrong in mitochondria suggest that whatever damage the bodies trigger in motor neurons becomes irreversible later in the illness than suspected. “This gives us hope,” says Xu, “that the right therapy might still be helpful even as muscles continue to weaken.” Also, by nailing down the role of mitochondria in the disease, Center scientists know they’re revealing novel targets for therapy.

As in much ALS research, mitochondria studies lean on animal models of the disease. Like patients with one inherited type of ALS, mouse and rat models carry an abnormal version of a human gene that codes for SOD1—a widespread cell protein. The animals develop a

Zuoshang Xu, Giovanni Manfredi, Don Cleveland— they assemble the pieces.

neuromuscular illness almost identical to human ALS, though much more rapid. And it’s that close overlap between animal and all human ALS, not just the inherited sort, that bolsters the idea they share the crucial downhill pathways.

The mitochondria story began when researchers found the mere presence of altered SOD1 protein could lead to animal paralysis. In a study out in 1995, Philip Wong, Don Cleveland and other soon-to-be Center scientists showed that only animals carrying mutant human SOD1 sickened. Neighbor mice with normal human SOD1 looked fine.

And what also became obvious—early on—was the appearance of vacuoles in motor neurons. Parts of neurons were filled with the membrane-bound bubbles. Even more odd, the vacuoles contained mitochondria in various stages of damage. “We certainly didn’t expect that,” says Wong. “We’d thought mitochondria had nothing to do, directly, with SOD1 and the damage it prompts.”

In 1998, Xu’s group also noticed the vacuoles in animal model studies at his University of Massachusetts lab. He’d aimed to clarify the stages cells go through in the course of ALS—stages that might mirror changes in muscle strength.

“At first,” he says, “you’d see unusual-looking mitochondria. But at the first outward sign of ALS—the onset of muscle weakness—you couldn’t miss the mitochondria-associated vacuoles in motor neurons,” says Xu. “And that got us thinking mitochondrial damage somehow tripped the onset of outright illness. Similar vacuoles appear in Alzheimer’s and Huntington’s disease.”

With that, the scientists took a closer look at the organelles: Could they still function in spite of obvious changes? Chiefly, mitochondria act as a cell’s power plants, housing the enzymes and machinery for whip-fast chemical reactions that produce energy. Yet Center investigator Giovanni Manfredi, at Cornell University, reported not only that spinal cord mitochondria of model mice suffered internal molecular damage but that their energy production had fallen—all around the time vacuoles appeared. And Xu’s team noted a corresponding drop in enzymes mitochondria needed to make energy.

Then last year came the first news of an actual physical link between SOD1 and mitochondria. Following the lead of Johns Hopkins/Packard researcher Val Culotta, who’d spotted SOD1 in yeast mitochondria, Manfredi and Xu announced they’d found abnormal versions of that molecule inside spinal cord mitochondria of model mice. This year, work published by Xu’s lab revealed abundant mutant SOD1 attached to the vacuole membranes. “It certainly looks like flawed SOD1 homes in on mitochondria and does damage,” says Xu.

Now, awaiting publication, comes what’s surely a pivotal study by Cleveland, Wong, Center director Jeffrey Rothstein and others. It draws from the earlier work to propose a likely scenario—how, together, SOD1 gone wrong and mitochondria make a cell’s worst partnership.

Recently, Cleveland’s team purified mitochondria from tissues in animal models—from liver, muscle, brain and spinal cord. Analyzing each sample led them to conclude, he says, “that basically, mutant SOD1 gums up mitochondria in the spinal cord, but not in other places.”

Like Xu’s, his team also found mutant SOD1 associated with mitochondria. In some instances, Cleveland explains, it slips inside them. But—and this is new—it’s far more common to find masses of the mutant protein gumming up the surface of the tiny bodies.

“The glitch lies in the entry mechanism,” say Cleveland. Mitochondria, scientists are finding, maintain a sophisticated import system to decide what gets inside and what doesn’t. Though still sketchy, this may be a case where mutant SOD1 passes muster enough to get into the sentry station but not inside the fort. So it accumulates outside. “And troubles multiply,” says Cleveland. “Excess SOD1 could clog mitochondrial pores, disrupting the healthy flow of molecules.” That’s important because—in addition to energy-production—mitochondria, when injured, can trip hard-wired cell programs for cell death. Already, Center and other scientists have signs that mitochondria are doing just that in ALS.

“Surprisingly,” adds Cleveland, “none of this goes on in other cells. You can have a model animal whose motor neuron mitochondria are going downhill fast, but whose liver mitochondria are perfectly fine. Same mutant SOD1. Same mouse.” Exploring why this is so is “a crucial objective,” he says, “for scientists like us, bent on therapy.”

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