Tiny pores in the cell nucleus protect and preserve genetic material, which is essential for healthy aging. A team from the Max Planck Institute of Biophysics in Frankfurt am Main and the Synthetic Biophysics of Protein Disorder group at Johannes Gutenberg University Mainz (JGU) filled a gap in our understanding of the structure and function of these nuclear pores.
The researchers discovered how inherently disordered proteins in the pore’s core can form a spaghetti-like movable barrier that allows critical cellular components through but inhibits viruses and other pathogens.
The nuclear membrane protects human cells’ genetic material inside the cell nucleus. The nucleus, as the cell’s control center, must be able to exchange vital messenger molecules, metabolites, or proteins with the remainder of the cell. As a result, around 2,000 pores are created into the nuclear membrane, each of which contains approximately 1,000 proteins.
For decades, researchers have been fascinated by the three-dimensional structure and function of these nuclear pores, which act as guardians of the genome, allowing substances required for cell control to pass while preventing pathogens or other DNA-damaging substances from entering.
As a result, the nuclear pores can be compared to molecular bouncers, with each checking thousands of visitors every second. Only those who have an entrance ticket are allowed to pass.
How do the nuclear pores manage this enormous task? About 300 proteins attached to the pore scaffold protrude like tentacles deep into the central opening.
Researchers had not know how these tentacles are structured or how they repel invaders until today. This is due to the fact that these proteins are disordered by nature and lack a clear three-dimensional structure. They are pliable and constantly moving, much like spaghetti in boiling water.
With our combination of methods, we can now study IDPs in more detail to find why they are indispensable for certain cellular functions, despite being error-prone. In fact, IDPs are found in almost all species, although they carry the risk of forming aggregates during the aging process which can lead to neurodegenerative diseases such as Alzheimer’s.
Edward Lemke
Combination of microscopy and computer simulations
Because these intrinsically disordered proteins (IDPs) are constantly changing their structure, scientists are having difficulty deciphering their three-dimensional architecture and function. Most experimental techniques for imaging proteins only work with a specified 3D structure. Because the organization of the IDPs in the opening could not be determined, the central region of the nuclear pore has been represented as a hole thus far.
The team led by Gerhard Hummer, Director at the Max Planck Institute of Biophysics, and Edward Lemke, Professor for Synthetic Biophysics at Johannes Gutenberg University Mainz and Adjunct Director at the Institute of Molecular Biology Mainz (IMB) has now used a novel combination of synthetic biology, multidimensional fluorescence microscopy and computer-based simulations to study nuclear pore IDPs in living cells.
“We used modern precision tools to mark several points of the spaghetti-like proteins with fluorescent dyes that we excite by light and visualize in the microscope,” explained Lemke. “Based on the glow patterns and duration, we were able to deduce how the proteins must be arranged.”
And Hummer added, “We then used molecular dynamics simulations to calculate how the IDPs are spatially organized in the pore, how they interact with each other and how they move. For the first time, we could visualize the gate to the control center of human cells.”
Dynamic protein network as transport barrier
Because they interact with each other and with the cargo, the tentacles in the transport pore exhibit completely different behavior than we previously observed. They travel indefinitely, much like the previously mentioned spaghetti in boiling water. So, instead of a hole, there is a shield of wiggly, spaghetti-like molecules in the center of the pore.
Viruses or bacteria are too big to get through this sieve. Other large cellular molecules required in the nucleus, on the other hand, can pass because they carry very specific signals. Pathogens, on the other hand, do not normally have an entry ticket.
“By disentangling the pore filling, we enter a new phase in nuclear transport research,” added Martin Beck, collaborator and colleague at the Max Planck Institute of Biophysics.
“Understanding how the pores transport or block cargo will help us identify errors. After all, some viruses manage to enter the cell nucleus despite the barrier,” Hummer summed up.
“With our combination of methods, we can now study IDPs in more detail to find why they are indispensable for certain cellular functions, despite being error-prone. In fact, IDPs are found in almost all species, although they carry the risk of forming aggregates during the aging process which can lead to neurodegenerative diseases such as Alzheimer’s,” said Lemke.
Researchers want to develop novel treatments or vaccinations that prevent viral infections and promote healthy aging by studying how IDPs work.
