The physics of fat droplets reveal DNA danger

Fat is a normal and necessary part of the body. Fat cells store and release energy, as well as play significant roles in hormonal regulation and immunity.

In recent decades, a concerning rise in metabolic illnesses -- such as cardiovascular disease, high blood pressure and diabetes -- has focused scientific attention on the biology and chemistry of fat, resulting in a wealth of information about how fat cells work.

But fat cells and their metabolic activities are only part of the story.

Fat-filled lipid droplets, tiny spheres of fat many times smaller than fat cells, are a growing subject of scientific interest. Found inside many different cell types, these lipid particles have long been little understood. Studies have begun to illuminate these droplets' participation in metabolic functions and cellular protection, but we still know next to nothing about the physical nature of fat.

Now, researchers at the University of Pennsylvania School of Engineering and Applied Science have looked beyond biochemistry to publish groundbreaking work on the physics of these droplets, revealing them to be a potential threat to a cell's nucleus. In the August issue of the Journal of Cell Biology, they are the first to discover fat-filled lipid droplets' surprising capability to indent and puncture the nucleus, the organelle which contains and regulates a cell's DNA.

The stakes of their findings are high: a ruptured nucleus can lead to elevated DNA damage that is characteristic of many diseases, including cancer.

The study was led by Dennis E. Discher, Robert D. Bent Professor in the Department of Chemical and Biomolecular Engineering, Irena Ivanovska, Ph.D. Research Associate in Penn's Molecular and Cell Biophysics Lab, and Michael Tobin, Ph.D. Candidate in the Department of Bioengineering.

"Intuitively, people think of fat as soft," says Discher. "And on a cellular level it is. But at this small size of droplet -- measuring just a few microns rather than the hundreds of microns of a mature fat cell -- it stops being soft. Its shape has a much higher curvature, bending other objects very sharply. This changes its physics in the cell. It can deform. It can damage. It can rupture."

"Imagine," adds Ivanovska, "trying to pop a balloon with your fist. Impossible. You can deform the balloon, but you won't puncture it. Now imagine trying to pop it with a pen. That's the difference between a fat cell and a cell with small fat droplets in the body. It's a fundamental physical difference, not a metabolic one."

The team's research reframes scientific inquiry into fat, underlining that fat's role in the body is much more than just a number on the scales.

"This isn't fat canonically conceived," says Tobin. "This is about how fat works at scales smaller than a cell and poses physical risks to cellular components, even at the level of DNA."

The team's work builds on a decade of foundational research, including leading contributions by Ivanovska, into the behaviors of nuclear proteins that give the nucleus its protective structural qualities. These proteins are dynamic, shifting levels to respond to their mechanical environments and provide what the nucleus needs to maintain its integrity.

"There's a constant process of repair to DNA damage that goes on in cells," says Ivanovska. "For this to happen, the nucleus needs to have enough DNA repair proteins. If a nucleus is ruptured, these proteins scatter and cannot repair damage in a timely manner. This causes DNA damage accumulation and can potentially result in a cancer cell."

A cell lives in a dynamic physical and mechanical environment where things can and do go wrong. But it also has an army of molecular helpers always working to maintain and repair it.

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How the motion of DNA controls gene activity

Performing cutting-edge science requires thinking outside the box and bringing together different scientific disciplines. Sometimes this even means being in the right place at the right time. For David Brückner, postdoctoral researcher and NOMIS fellow at ISTA, all the above-mentioned things came into effect as he attended an on-campus lecture by Professor Thomas Gregor from Princeton University. Inspired by the talk, Brückner reached out with an idea: to physically interpret the specific data sets Gregor presented. Now, the results of their collaboration are published in Science. They highlight the stochastic (random) motion of two specific gene elements on a chromosome, which have to come into contact for the gene to become active in 3D space.

How DNA fits into a cell nucleus

Living organisms like humans are built on genes that are stored in the DNA -- our molecular blueprint. DNA is a polymer, a huge molecule of smaller individual parts (monomers). It is located in every cell's nucleus. "Depending on the organism, the DNA polymer can be up to meters long, yet the size of the nucleus is on the order of microns," Brückner explains. To fit into the tiny nucleus, DNA gets compacted by being coiled as if on a spool and further compressed into the well-known shape of chromosomes, which we all encountered in a biology textbook.

"Despite being heavily condensed, chromosomes are not static; they are jiggling around all the time," the physicist continues. These dynamics are very important. Whenever a specific gene has to be activated, two regions on the polymer called "enhancer" and "promoter" need to come into close contact and bind to each other. Only when this happens, a cellular machinery reads off the gene's information and forms the RNA molecule, which eventually gives rise to proteins that are essential for all the processes a living organism requires.

Depending on the organism, the enhancer and promoter can be quite far from each other on the chromosome. "With previously used methods, you could get a static view of the distance between these elements, but not how the system evolves over time," Brückner explains. Intrigued by this missing information, the scientists set out to get a dynamic look at how these elements are organized and how they move in 3D space in real time.

Visualizing gene regions

To achieve this goal, the experimental scientists from Princeton established a method to track those two DNA elements over a certain time period in a fly embryo. Through genetic manipulation, the DNA elements were fluorescently labeled, with the enhancer region illuminating in green and the promoter in blue. Using live imaging (time-lapse microscopy of living cells) the scientists were able to visualize the fluorescent spots in fly embryos to see how they were moving around to find each other.

Once the two spots came into proximity, the gene was activated and an additional red light turned on as the RNA was also tagged with red fluorophores. Brückner excitedly adds, "We got a visual readout of when the enhancer and promoter got in contact. That gave us a lot of information about their trajectories."

DNA is densely packed and exhibits fast motion

The challenge then was how to analyze this huge data set of stochastic motion. His background in theoretical physics allowed Brückner to extract statistics to understand the typical behavior of the system. He applied two simplified, different physical models to cut through the data.

One was the Rouse model. It assumes that every monomer of the polymer is an elastic spring. It predicts a loose structure and fast diffusion -- a random movement, where occasionally the gene regions encounter each other. The other model is called the "fractal globule." It predicts a very compact structure and therefore slow diffusion. "Surprisingly, we found in the data that the system is described by a combination of these two models -- a highly dense structure you would expect based on the fractal globule model, and diffusion which is described by the statistics from the Rouse model," Brückner explains.

Due to the combination of dense packing and fast motion, the binding of these two gene regions depends much less on their distance along the chromosome than previously anticipated. "If such a system is in a fluid and dynamic state all the time, long-distance communication is much better than we might have thought," Brückner adds.

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