A collaboration involving Lawrence Berkeley National Laboratory scientists published one of the latest studies in epigenetics using imaging from a powerful x-ray microscope — shedding new light on cell differentiation, the process through which stem cells develop to serve a specific function.
The research, which began in 2012 and was published last week in the journal Cell Report, was led by University of California, San Francisco anatomy professor Carolyn Larabell and Columbia University biochemistry and molecular biophysics professor Stavros Lomvardas, and included a team of chemists, physicists and biologists.
The study marks a major breakthrough in cell imaging techniques that reveal all the structures in a cell nucleus, as well as an opportunity for scientists to gain a better understanding of changes in cell structure during the process of differentiation, wherein some genes in the cell are “silenced,” or prevented from being expressed.
“All cells have the same DNA in them, but they end up doing different things,” said researcher and UCSF anatomy professor Mark Le Gros. “The stem cell kind of has everything there (but) decides to become a particular type of cell. (This) allows us to respond to our environment without having to change our genes.”
To observe the organic components in cells during differentiation, Larabell said, the team built a soft x-ray microscope which uses a unique range of low energy x-rays. Nitrogen and carbon — key elements in organic material — absorb this range of energy in an order of magnitude greater than water. Thus, the scientists were able to create images in which water becomes invisible compared to the cell structures.
Larabell said that by observing the contrast in these images, the researchers were able to tell how crowded a cell region is and to quantify the presence of chromatin, a DNA-containing material.
The method also provides a new three-dimensional view on the entire cell nucleus that had been previously unavailable to scientists using electron microscopy, which could only show dehydrated cell fragments, according to Larabell.
The U.S. Department of Energy funded the operation of the microscope used to observe the cells, while the biological project itself was supported by the National Institutes of Health.
The researchers found that the percentage of the nucleus occupied by heterochromatin, or silenced DNA, increases during the differentiation of neurons. Additionally, they found that all of the heterochromatin — a form of chromatin — is connected during the process of differentiation, contrary to the former belief that heterochromatin form islands surrounded by euchromatin, which carry active genes. According to Larabell, this erroneous theory had resulted from observations made through two-dimensional cell imaging.
One possible reason for the connectedness of heterochromatin is so that the nucleus’s entire mass of euchromatin, instead of being buried within heterochromatin, is accessible to proteins necessary for the construction of proteins from DNA material, Larabell said.
“The heterochromatin is about 30 percent more crowded than euchromatin — that wasn’t known before either,” Larabell said. “So it’s harder for molecules to move through heterochromatin.”
According to Larabell, now that the team has constructed the structural map of the nucleus during differentiation, it will move on to studying the formation of nuclei and the process in which some genes are silenced during cell differentiation.
“The behaviors of the cell is determined by what genes are on and off in that cell, and the next question is: What turns the gene on or off? … That’s the essence of epigenetics,” said UC Berkeley bioengineering professor David Schaffer, who was not involved in the research. “This regulation of genes being turned on and off has been shown to be associated to many diseases.”
For instance, the lethal Huntington’s disease is caused by the expression, or switching on, of a broken gene within the brain neurons of patients, according to Schaffer. Through a broader understanding of epigenetics, scientists may be more likely gain further insights into these health issues.