A picture is worth 1,000 words, and this holds true for the plethora of information researchers can obtain from pictures of our cells. The constant processes occurring inside cells are critical for helping our bodies function properly, and their dysregulation can lead to various diseases. One of the goals of cell biology is to take pictures of the cells' tiny inner workings to understand them better. In "Visualizing nuclear pore complex plasticity with pan-expansion microscopy", Dr. Kimberly Morgan, a postdoctoral fellow in the laboratory of Drs. Patrick Lusk and Megan King (aka the "LusKing" lab) brings a novel level of detail to nuclear pore complexes by looking at them with pan-expansion microscopy.
Nuclear Pore Complexes (NPCs) are channels inside cells that span the nuclear envelope. The nuclear envelope separates the nucleus, which houses our DNA, from the rest of the cell: the cytoplasm, but materials still need to get across it. For example, the messages inscribed in our DNA in the nucleus need to be translated into proteins that carry out the cells’ functions, which occurs in the cytoplasm. NPCs serve as regulators of this transit across the nuclear envelope. The thousands of NPCs per cell are all composed of proteins called nucleoporins, but there are variations in each NPC’s size, shape, and composition. These features can impact how quickly material moves through the pores. For example, traffic moves quickly under a tall, wide bridge on a multi-lane highway, but moves slower under a short bridge on a narrow country road. Variation in NPC characteristics similarly affect which material can pass through and how quickly. Although some of this variation is normal, certain changes to NPCs can impair their function and are associated with conditions like neurodegenerative diseases. Observing these structures in their native environment inside cells can help us understand how NPC variation contributes to healthy and diseased cell states.
Pan-expansion microscopy is a new approach to visualize extremely small details inside the cell. Much like how objects far away appear fuzzy through a camera, the tiniest objects in our cells are difficult to visualize clearly through a microscope. One way researchers have tried to solve this problem is by improving microscopes to see ever smaller objects. Alternatively, pan-expansion microscopy seeks to address this problem differently by making the actual object, in this case the cells and their contents, bigger. The week-long pan-expansion process “includes ‘fixing’ your cells, which creates a snapshot of what they look like at a certain point in time”, describes Dr. Morgan. “Then, you physically embed the cells in a series of swellable hydrogels so that [the cells] physically expand in size 16-fold.” Dr. Morgan learned this approach from the Pan-Expansion Microscopy Workshop led by the Bewersdorf lab in the Yale Department of Cell Biology, who developed the technique. She is the first researcher to look at NPCs with the novel level of detail provided by pan-expansion microscopy. Armed with this approach, she set out to determine how variation between NPCs may affect their function in normal and diseased cells.
After staining the DNA and proteins and taking pictures of her expanded cells on the microscope, Dr. Morgan developed a computational protocol to identify NPCs by looking for their classical 3-ring structure spanning the nuclear envelope (Figure 1). By observing thousands of NPCs in their native environment, she could look for variations and patterns among them. She noticed that NPCs have different diameters, or different sized channels. However, while the variation seemed random at first, Dr. Morgan noticed an emerging pattern: NPCs with similar sizes clustered together in certain regions. This suggested that perhaps the NPCs were under different levels of tension across the nuclear envelope. Cells, and the structures inside of them, can experience different levels of mechanical force when in environments of different stiffness or tension. The size of the NPCs can determine how much material they allow to pass through. Thus, changes in their diameter from the tension they experience suggests a new way that cells regulate traffic between the nucleus and cytoplasm.
In addition to visualizing NPCs in healthy cells, Dr. Morgan investigated how changes in NPCs drive neurodegenerative diseases, like Amyotrophic Lateral Sclerosis (ALS). Many ALS patients have a mutation, or change, in their DNA that causes certain nucleoporins to be lost from NPCs. These “injured” NPCs prevent proper transportation between the nucleus and cytoplasm in neurons. However, researchers could not fully understand this mechanism without looking at NPCs in greater detail. To address this problem, Dr. Morgan generated neurons from stem cells from an ALS patient with this mutation that affects nuclear pores. With pan-expansion microscopy, she observed that the specific nucleoporin affected in ALS was indeed lost from many NPCs in patient neurons. Surprisingly, her detailed approach also revealed that some NPCs retained the nucleoporin, but it had localized to a different position in the NPC structure. This change in location may be a previously undescribed step in ALS progression and reveals how her approach could identify more molecular changes in NPCs in other diseases.
Studying changes in NPCs has been a throughline through Dr. Morgan’s research career. Originally from Melbourne, Australia, Dr. Morgan completed her graduate work at the Walter and Eliza Hall Institute of Medical Research where she studied NPCs in cancer in a zebrafish model. She then joined the LusKing lab as a postdoctoral fellow in 2022. Her passion for discovering new information led Dr. Morgan to pursue science. “I became a scientist because I’m curious about how things work” says Dr. Morgan. “Even now when I image [cells] and discover something exciting, it’s really a privilege to get to see that and share it”. Outside of the lab, she continues to educate others as a volunteer at New Haven Reads, where she tutors a student weekly in reading comprehension. She is also an avid rock climber.
Read Dr. Morgan’s full paper here!