String Theory

New types of biological filaments are turning up in yeast, fly, bacterial cells and in rat neurons, and they may yield clues to how the cytoskeleton evolved from metabolically active enzymes.

Aug 1, 2011
Richard P. Grant

James Wilhelm and Chalongrat Noree in the lab.DANE SAMILO

Cells have three filament types that together make up the cytoskeleton—actin, intermediate filaments, and microtubules. These filaments consist of repeating protein units and orchestrate vital functions, including cellular motility, cell division, macromolecule transport, and maintenance of the structural integrity of cells and tissues. Now James Wilhelm’s group at the University of California, San Diego, has discovered four new types of protein filaments, and believes that there could be more.

In 2003, researchers at the University of California, San Francisco used a library that contained three-quarters of the yeast proteome tagged with green fluorescent protein (GFP) to visualize the localization of proteins in living cells. A few years later, Wilhelm realized that they hadn’t detected P bodies—cytoplasmic foci made up of mRNA degradation enzymes—in the screen. Wondering what else could have been missed, he asked graduate student Chalongrat Noree to comb the yeast protein library to try to find aggregates of GFP-tagged proteins large enough to see using fluorescence microscopy.

Wilhelm says he’s been intrigued by the thought that there could be more than three types of filaments in cells since his own days as a grad student. Actin, microtubules, and intermediate filaments are hard to miss, he says, but “how do we know we have all the filament-forming proteins in the cell?” When Noree probed the yeast proteome, he was shocked. He saw that some proteins were forming long filaments in the cells. And they weren’t traditional cytoskeletal elements, but enzymes involved in metabolism and biosynthesis, such as glutamate synthase.

“We were stunned at the number of things we were finding,” Wilhelm says. At the time, Noree identified nine different proteins that formed four different filaments, although the number of proteins detected in various intracellular structures has ballooned since their paper was published last August (J Cell Bio, 190:541-51, 2010).

Composite photo of the filaments in A) yeast, B) Drosophila, and C) neurons
Composite photo of the filaments in A) yeast, B) Drosophila, and C) neurons

Wilhelm says he practically had to wrench Noree from the microscope so they could write the paper. “I kept expecting things to come to a focus, but instead they kept fanning out, and [we were] finding increasing numbers of novel structures,” Wilhelm says. “It made putting a manuscript together increasingly difficult.” Mark Rose at Princeton University says that this demonstrates the power of large-scale genomic approaches, and that it’s “worthwhile and effective to do open-ended screens.” Wilhelm adds that high-throughput, automated systems aren’t always optimal, saying, “It’s really hard to beat the human eye” at recognizing and interpreting new structures.

Wilhelm’s lab concentrated on an enzyme necessary for ribonucleotide metabolism, cytidine triphosphate (CTP) synthase, because its regulation has been well studied in yeast and it’s an essential, conserved protein. They used polyclonal antibodies against CTP synthase to look for similar filaments in Drosophila. Remarkably, they found them in all cell types in the fly’s egg chamber, but only in some of the cells in the gut. Wilhelm argues that this shows that formation of CTP synthase filaments is tightly regulated in different cells. The fact that they don’t form in every cell type, or under all conditions, might also explain why nobody had seen them before.

Rose says that these filaments “suggest yet another way one could regulate metabolism.” Making a filament out of an enzyme will fix it in a particular shape, which could lock it into an active or inactive state. CTP synthase, for example, appears to be inactive in filament form. This provides a mechanism for controlling its activity that is independent of substrate or protein concentration, but does raise the question of what regulates filament formation, a question Wilhelm is now investigating.

Remarkably, a paper from Zemer Gitai’s Princeton University lab, also published last August, found that CTP synthase also forms filaments in the bacterium Caulobacter crescentus (Nature Cell Bio, 12:739-46, 2010). Those filaments seem to be essential for regulating the curved shape of C. crescentus, and their formation may play a role in metabolic regulation by locking the enzyme into an active or inactive state. This could be analogous to how the actin cytoskeleton is thought to have evolved: actin is very similar to hexokinase, the enzyme that catalyzes the first step in glycolysis. It’s an “idea that we’re really excited about,” Wilhelm says, “in that you develop this type of enzyme regulation, and it’s relatively easy to co-opt that to build the cytoskeleton.”

A Hidden Jewel refers to an article, published in a specialist journal, which has been evaluated in Faculty of 1000, a post-publication peer review service of the Science Navigation Group. Read the evaluation of Wilhelm’s article.