transcription factors
The Amazing Brain: Tracking Molecular Events with Calling Cards
Posted on by Dr. Francis Collins
In days mostly gone by, it was fashionable in some circles for people to hand out calling cards to mark their arrival at special social events. This genteel human tradition is now being adapted to the lab to allow certain benign viruses to issue their own high-tech calling cards and mark their arrival at precise locations in the genome. These special locations show where there’s activity involving transcription factors, specialized proteins that switch genes on and off and help determine cell fate.
The idea is that myriad, well-placed calling cards can track brain development over time in mice and detect changes in transcription factor activity associated with certain neuropsychiatric disorders. This colorful image, which won first place in this year’s Show Us Your BRAINs! Photo and Video contest, provides a striking display of these calling cards in action in living brain tissue.
The image comes from Allen Yen, a PhD candidate in the lab of Joseph Dougherty, collaborating with the nearby lab of Rob Mitra. Both labs are located in the Washington University School of Medicine, St. Louis.
Yen and colleagues zoomed in on this section of mouse brain tissue under a microscope to capture dozens of detailed images that they then stitched together to create this high-resolution overview. The image shows neural cells (red) and cell nuclei (blue). But focus in on the neural cells (green) concentrated in the brain’s outer cortex (top) and hippocampus (two lobes in the upper center). They’ve been labelled with calling cards that were dropped off by adeno-associated virus [1].
Once dropped off, a calling card doesn’t bear a pretentious name or title. Rather, the calling card, is a small mobile snippet of DNA called a transposon. It gets dropped off with the other essential component of the technology: a specialized enzyme called a transposase, which the researchers fuse to one of many specific transcription factors of interest.
Each time one of these transcription factors of interest binds DNA to help turn a gene on or off, the attached transposase “grabs” a transposon calling card and inserts it into the genome. As a result, it leaves behind a permanent record of the interaction.
What’s also nice is the calling cards are programmed to give away their general locations. That’s because they encode a fluorescent marker (in this image, it’s a green fluorescent protein). In fact, Yen and colleagues could look under a microscope and tell from all the green that their calling card technology was in place and working as intended.
The final step, though, was to find out precisely where in the genome those calling cards had been left. For this, the researchers used next-generation sequencing to produce a cumulative history and map of each and every calling card dropped off in the genome.
These comprehensive maps allow them to identify important DNA-protein binding events well after the fact. This innovative technology also enables scientists to attribute past molecular interactions with observable developmental outcomes in a way that isn’t otherwise possible.
While the Mitra and Dougherty labs continue to improve upon this technology, it’s already readily adaptable to answer many important questions about the brain and brain disorders. In fact, Yen is now applying the technology to study neurodevelopment in mouse models of neuropsychiatric disorders, specifically autism spectrum disorder (ASD) [2]. This calling card technology also is available for any lab to deploy for studying a transcription factor of interest.
This research is supported by the Brain Research through Advancing Innovative Neurotechnologies® (BRAIN) Initiative. One of the major goals of BRAIN Initiative is to accelerate the development and application of innovative technologies to gain new understanding of the brain. This award-winning image is certainly a prime example of striving to meet this goal. I’ll look forward to what these calling cards will tell us in the future about ASD and other important neurodevelopmental conditions affecting the brain.
References:
[1] A viral toolkit for recording transcription factor-DNA interactions in live mouse tissues. Cammack AJ, Moudgil A, Chen J, Vasek MJ, Shabsovich M, McCullough K, Yen A, Lagunas T, Maloney SE, He J, Chen X, Hooda M, Wilkinson MN, Miller TM, Mitra RD, Dougherty JD. Proc Natl Acad Sci U S A. 2020 May 5;117(18):10003-10014.
[2] A MYT1L Syndrome mouse model recapitulates patient phenotypes and reveals altered brain development due to disrupted neuronal maturation. Jiayang Chen, Mary E. Lambo, Xia Ge, Joshua T. Dearborn, Yating Liu, Katherine B. McCullough, Raylynn G. Swift, Dora R. Tabachnick, Lucy Tian, Kevin Noguchi, Joel R. Garbow, John N. Constantino. bioRxiv. May 27, 2021.
Links:
Brain Research through Advancing Innovative Neurotechnologies® (BRAIN) Initiative (NIH)
Autism Spectrum Disorder (National Institute of Mental Health/NIH)
Dougherty Lab (Washington University School of Medicine, St. Louis)
Mitra Lab (Washington University School of Medicine)
Show Us Your BRAINs! Photo and Video Contest (BRAIN Initiative/NIH)
NIH Support: National Institute of Neurological Disorders and Stroke; National Institute of Mental Health; National Center for Advancing Translational Sciences; National Human Genome Research Institute; National Institute of General Medical Sciences
Protein Links Gut Microbes, Biological Clocks, and Weight Gain
Posted on by Dr. Francis Collins
Caption: Lipids (red) inside mouse intestinal cells with and without NFIL3.
Credit: Lora V. Hooper, University of Texas Southwestern Medical Center, Dallas
The American epidemic of obesity is a major public health concern, and keeping off the extra pounds is a concern for many of us. Yet it can also be a real challenge for people who may eat normally but get their days and nights mixed up, including night-shift workers and those who regularly travel overseas. Why is that?
The most obvious reason is the odd hours throw a person’s 24-hour biological clock—and metabolism—out of sync. But an NIH-funded team of researchers has new evidence in mice to suggest the answer could go deeper to include the trillions of microbes that live in our guts—and, more specifically, the way they “talk” to intestinal cells. Their studies suggest that what gut microbes “say” influences the activity of a key clock-driven protein called NFIL3, which can set intestinal cells up to absorb and store more fat from the diet while operating at hours that might run counter to our fixed biological clocks.
Snapshots of Life: A Van Gogh Moment for Pancreatic Cancer
Posted on by Dr. Francis Collins
Salt Lake City
Last year, Nathan Krah sat down at his microscope to view a thin section of pre-cancerous pancreatic tissue from mice. Krah, an MD/PhD student in the NIH-supported lab of Charles Murtaugh at the University of Utah, Salt Lake City, had stained the tissue with three dyes, each labelling a different target of interest. As Krah leaned forward to look through the viewfinder, he fully expected to see the usual scattershot of color. Instead, he saw enchanting swirls reminiscent of the famous van Gogh painting, The Starry Night.
In this eye-catching image featured in the University of Utah’s 2016 Research as Art exhibition, red indicates a keratin protein found in the cytoskeleton of precancerous cells; green, a cell adhesion protein called E-cadherin; and yellow, areas where both proteins are present. Finally, blue marks the cell nuclei of the abundant immune cells and fibroblasts that have expanded and infiltrated the organ as a tumor is forming. Together, they paint a fascinating new portrait of pancreatic ductal adenocarcinoma (PDAC), the most common form of pancreatic cancer.
Creative Minds: A Transcriptional “Periodic Table” of Human Neurons
Posted on by Dr. Francis Collins
Caption: Mouse fibroblasts converted into induced neuronal cells, showing neuronal appendages (red), nuclei (blue) and the neural protein tau (yellow).
Credit: Kristin Baldwin, Scripps Research Institute, La Jolla, CA
Writers have The Elements of Style, chemists have the periodic table, and biomedical researchers could soon have a comprehensive reference on how to make neurons in a dish. Kristin Baldwin of the Scripps Research Institute, La Jolla, CA, has received a 2016 NIH Director’s Pioneer Award to begin drafting an online resource that will provide other researchers the information they need to reprogram mature human skin cells reproducibly into a variety of neurons that closely resemble those found in the brain and nervous system.
These lab-grown neurons could be used to improve our understanding of basic human biology and to develop better models for studying Alzheimer’s disease, autism, and a wide range of other neurological conditions. Such questions have been extremely difficult to explore in mice and other animal models because they have shorter lifespans and different brain structures than humans.
Regenerative Medicine: Making Blood Stem Cells in the Lab
Posted on by Dr. Francis Collins
Caption: Arrow in first panel points to an endothelial cell induced to become hematopoietic stem cell (HSC). Second and third panels show the expansion of HSCs over time.
Credit: Raphael Lis, Weill Cornell Medicine, New York, NY
Bone marrow transplants offer a way to cure leukemia, sickle cell disease, and a variety of other life-threatening blood disorders.There are two major problems, however: One is many patients don’t have a well-matched donor to provide the marrow needed to reconstitute their blood with healthy cells. Another is even with a well-matched donor, rejection or graft versus host disease can occur, and lifelong immunosuppression may be needed.
A much more powerful option would be to develop a means for every patient to serve as their own bone marrow donor. To address this challenge, researchers have been trying to develop reliable, lab-based methods for making the vital, blood-producing component of bone marrow: hematopoietic stem cells (HSCs).
Two new studies by NIH-funded research teams bring us closer to achieving this feat. In the first study, researchers developed a biochemical “recipe” to produce HSC-like cells from human induced pluripotent stem cells (iPSCs), which were derived from mature skin cells. In the second, researchers employed another approach to convert mature mouse endothelial cells, which line the inside of blood vessels, directly into self-renewing HSCs. When these HSCs were transplanted into mice, they fully reconstituted the animals’ blood systems with healthy red and white blood cells.
