We have the first video of a plant cell wall being built

Plant cells are surrounded by an intricately structured protective coat called the cell wall. It’s built of cellulose microfibrils intertwined with polysaccharides like hemicellulose or pectin. We have known what plant cells look like without their walls, and we know what they look like when the walls are fully assembled, but we’ve never seen the wall-building process in action. “We knew the starting point and the finishing point, but had no idea what happens in between,” says Eric Lam, a plant biologist at Rutgers University. He’s a co-author of the study that caught wall-building plant cells in action for the first time. And once we saw how the cell wall building worked, it looked nothing like how we drew that in biology handbooks.

Camera-shy builders

Plant cells without walls, known as protoplasts, are very fragile, and it has been difficult to keep them alive under a microscope for the several hours needed for them to build walls. Plant cells are also very light-sensitive, and most microscopy techniques require pointing a strong light source at them to get good imagery.

Then there was the issue of tracking their progress. “Cellulose is not fluorescent, so you can’t see it with traditional microscopy,” says Shishir Chundawat, a biologist at Rutgers. “That was one of the biggest issues in the past.” The only way you can see it is if you attach a fluorescent marker to it. Unfortunately, the markers typically used to label cellulose were either bound to other compounds or were toxic to the plant cells. Given their fragility and light sensitivity, the cells simply couldn’t survive very long with toxic markers as well.

So, Lam’s team developed their own, custom-built imaging platform. The cells that star in the team’s wall-building video were protoplasts of Arabidopsis, a small flowering plant belonging to the same family as cabbage and mustard. As their shooting technique of choice, the team selected total internal reflection fluorescent microscopy (TIRFM), which illuminates only a tiny section of the sample at a time, reducing phototoxicity to manageable levels. For labelling, the team linked parts of enzymes that stick to cellulose to a non-toxic green fluorescent marker that could be used in live cells.

The setup also included a programmable LED light bulb that automatically switched off after each imaging cycle and a temperature control system that kept the cells at a steady 18° C (64° F). Once the setup was ready, the team fired it up and left the protoplasts in there for 24 hours. “This allowed us to have all the components to see the process and understand how it really works,” Lam says. And how it really works proved quite surprising.

Ordering chaos

“The idea we had about this process was that cells were continuously extruding these long polymer fibers that got assembled to form cell walls,” Chundawat told Ars. “It didn’t work like that—it was a lot more complex.” At first, very short cellulose fibers were being formed that were moved across the cell’s surface seemingly at random, as though looking for other short fibers. When those short fibers meet and engage with each other, they combine into the longer fibers that protrude from the cell like waving tentacles. Only after they bump into other fibers do they start to intertwine and assemble into the organized structure of the cell wall. “It was a two-step process. That was something we did not expect,” Chundawat says.