If you wear your skeleton on the outside, there's only one option when you need to grow; either stay cramped or lose the shell. In the weeks before a crayfish sheds its exoskeleton, its body moves a massive amount of calcium out of the cuticle and stores it as small chalky pebbles in part of the stomach. Once the old skeleton has been shed, those pebbles, known as gastroliths, drop into the stomach acid. The calcium is carried right back and deposited in the new cuticle, and is topped up with extra calcium extracted from the environment. Without their skeleton, crayfish are soft like jelly. But the calcium piles on so fast you can feel a papery shell develop within hours of having lost the last. This large-scale calcium transport across cells is an astounding feat of physiology, especially when you consider the fine balance of calcium ions maintained in cells. Michele Wheatly and Yongping Gao, at Wright State University in Dayton, Ohio, have described and sequenced one important gene involved in the process, which produces a protein that pumps calcium out of cells. They have shown that crayfish cells increase production of this protein before and after shedding(p. 2991) and they are beginning to understand how regulation of this and other proteins controls cellular calcium levels. Free calcium ions are crucial intermediates in signalling processes inside every cell. To keep communication lines free of interference, cells have to maintain low background calcium levels in the cytoplasm, using a complex system of calcium homeostasis. In some circumstances, such as when mammals produce milk or crustaceans moult, large amounts of calcium have to move through cells; then, the regulation system is put to the test and begins to reveal its secrets. `Problems of calcium homeostasis are implicated in all kinds of medical conditions, including cardiovascular disease and Alzheimer's,' says Wheatly. `The proteins involved are coded by ancient genes, so what happens in crayfish can tell us a lot about how [the proteins] work in humans.' Crayfish living in freshwater occupy an extreme environment in calcium terms, compared to their marine counterparts. Levels of calcium available to them from the surrounding water are low, so calcium recycling is especially important.
Wheatly and Gao studied one of four Plasma Membrane Calcium ATPases (PMCA3)present in most animals. This protein sits across the external membrane of cells and exports calcium. They found the protein's gene in the freshwater crayfish genome using stretches of sequence from human and rodent versions. After sequencing the gene, they watched its behaviour during the moult cycle,by testing dissected crayfish tissues for the RNA version of the gene, and the protein itself. As they expected, in cells transporting the calcium, more PMCA3 was produced before the moult, and just after the moult, than inbetween moulting. What they did not expect was that this pattern was also found, to a lesser extent, in crayfish tissues not involved in the mass calcium transport,such as muscles. Wheatly's team are now working on the genetic regulation of other proteins that move calcium from the cell cytoplasm into the folds of internal membranes known as sarco-endoplasmic reticulum, effectively hiding it. Regulation of the two proteins seems to be linked - as one goes up the other goes down. `The way the system is regulated is almost like a dance,'Wheatly enthuses.
