Unfortunately, TRPML1 isn't the easiest protein to study. Instead of residing in the cell's easily-accessed outer membrane, where many other proteins nestle, it hides in a tiny, interior pocket called lysosome. To probe the protein, Xu's group had to modify a technique known as the patch clamp, in which a micropipette and electrodes are attached to a cell membrane to record the activity of individual or multiple proteins that serve as channels for charged particles (ions) moving in and out of cells. With their modification, which they call the lysosome patch clamp, Xu's group was able to record TRPML1 activity in the tiny lysosome.
They found that TRPML1 was indeed capable of ferrying iron out of the lysosome. But was there any evidence that interfering with that ability might result in ML4 symptoms? To address that question, Xu's group studied defective TRPML1 proteins bearing the same mutations as those found in ML4 patients. Mutations associated with severe symptoms were the least adept at shuttling iron, while those associated with milder symptoms were more proficient, although still not fully functional.
Further experiments confirmed that when TRPML1 is defective, iron becomes trapped in the lysosome. One result of the buildup is formation of a brownish waste material, lipofuscin, known as the "aging pigment." In skin cells, lipofuscin is the culprit responsible for the dreaded liver spots that appear with increasing age, but in nerve, muscle and other cells, its accumulation has more serious consequences.
"How lipofuscin causes problems in neurons and muscles is not clear, but it's believed that this is garbage that, in time, compromises the normal function of the lysosome," Xu said. "And we know the lysosome is important for all kinds of cell biology, particularly the recycling of intracellular components, so if it's damaged,
|Contact: Nancy Ross-Flanigan|
University of Michigan