Researchers film fundamental life process at cellular level

Pre-60S subunit (red), passes through a nuclearpore. The "gel plugs", or the paths of the subunit, are marked in green. Credit: Dr. Jan Ruland, AG Kubitscheck/Uni Bonn
Complex molecular machines are responsible for the assembly of all proteins within a cell. These ribosome precursors are made in the cell nucleus. They then enter the cells through what are called nuclear pores. This fundamental process has been filmed in living cells by researchers at the University of Bonn, ETH Zurich and others. These experiments have improved our understanding of the creation of ribosomes. Nature Communications published the study.

The cell nucleus acts as a vault. It is found in every cell and protects the DNA. This contains all of the instructions that will make the cells' proteins. The cell orders the transcript of the matching DNA fragment in the nucleus when it needs a particular protein. This copy is sent to the nucleus, where it reaches the complex molecular machines called ribosomes. They then follow the steps to make the desired protein.

The vast majority of cell molecules are made outside the nucleus. This does not apply to ribosomes as their many components are assembled in the nucleus. The pre-60S and pre-40S subunits are formed. They are then passed through the nuclear pores to the cell where they are assembled in the final step to create the ribosome.

Now, the University of Bonn's team and ETH Zurich have filmed the process of exporting the larger pre-60S unit. "To accomplish this, we stained nuclear pores with a green dye while the pre-60S unit was in red."

Prof. Ulrich Kubitscheck, of the Institute for Physical and Theoretical Chemistry of the University of Bonn, explains. The recording was made using a special microscope that the researchers had modified specifically for this purpose.

A subunit of pre-60S (red) passes through the nuclear pore. The "gel plugs", are marked in green. Credit: Dr. Jan Ruland, AG Kubitscheck/University of Bonn

Gel plugs are pore-stoppers

Dr. Jan Ruland is the study's principal author. He completed his doctorate at Kubitscheck's research lab. This is not trivial. The nucleus is almost completely covered by several thousand pores. Each one has a diameter of approximately one ten-thousandthof a millimeter. This success is due to advances in microscopy technology and more than ten years worth of research in which researchers have continually optimized their method.

Transport through pores is complex. They are covered with a gel that blocks larger molecules from passing. Without helpers, the subunits of ribosomes would not be able to leave the nucleus. These molecules are called export receptors. They can "swim through" the gel plug. Kubitscheck's colleague, Dr. Jan Peter Siebrasse, explains that the protein gripper pulls out the unit of ribosomes from each pore.

Only 25 milliseconds is required for passage

This seems to be the "bottleneck", in the transport process. "We were able show that the pre-60S units accumulate exactly at the point when the protein grabper reaches into a pore," said Siebrasse. Siebrasse is also a member of Transdisciplinary Research Area "Building Blocks of Matter" and Fundamental Interactions (TRA Matter). The export proceeds quickly, however. Study participants estimate that between 35 and 50 subunits can pass through one pore at a time.

However, the film evaluations show that export doesn't always work. Only in three cases where a pre-60S unit was in contact with a pore did it actually leave the nucleus. Kubitscheck speculates that the remaining cases were aborted because other molecules were being moved out of the nucleus simultaneously.

This study gives more insight into the formation and function of ribosomes.

The method can also be used to investigate other transport processes. Kubitscheck says, "We've exhausted all technical options." "It's not surprising that reviewers called our study a reference we hope other groups will benefit from," Kubitscheck said.

Continue reading: Learn more about how ribosomes work in human cells

Jan Andreas Ruland and colleagues, Nuclear export of pre-60S ribosomal protein through single nuclear pores observed over time, Nature Communications (2021). Information from Nature Communications Jan Andreas Ruland and colleagues, Nuclear export of pre-60S subunit via single nuclear pores observed over time, (2021). DOI: 10.1038/s41467-021-26323-7