Aidesigned protein awakens silenced genes one
By putting together CRISPR technology and a protein made with AI, it is possible to wake up dormant genes by turning off the chemical “off switches” that keep them from working. In the journal Cell Reports, researchers from the University of Washington School of Medicine in Seattle talk about this finding.
Shiri Levy, the lead author of the paper and a postdoctoral fellow at the UW Institute for Stem Cell and Regenerative Medicine (ISCRM), said that this method will help researchers figure out how individual genes affect normal cell growth and development, ageing, and diseases like cancer.
Levy said, “The great thing about this approach is that we can safely upregulate certain genes to change how cells work without permanently changing the genome and making mistakes we didn’t mean to.”
Hannele Ruohola-Baker, who is a professor of biochemistry and an associate director of ISCRM, was in charge of the project. The AI-designed protein was made at the UW Medicine Institute for Protein Design (IPD) under the direction of David Baker, who is also a professor of biochemistry and the head of the IPD.
The new method controls gene activity without changing the DNA sequence of the genome. It does this by focusing on chemical changes that help pack genes into our chromosomes and control how they work. The word “epigenetic” comes from the Greek word “epi,” which means “over” or “above” the genes. This is because these changes don’t happen inside the genes but on top of them. Epigenetic markers are the chemical changes that control how a gene works.
Scientists are especially interested in epigenetic changes because they not only change the way genes work in normal cell function, but they also build up over time and contribute to ageing. They can also affect the health of future generations because we can pass them on to our children.
In their work, Levy and her colleagues focused on a group of proteins called PRC2. PRC2 turns off genes by attaching a small molecule called a methyl group to a protein called histones, which is part of the package that holds genes. These methyl groups need to be reactivated so that if PRC2 is stopped, the genes it shut down can be turned back on.
PRC2 is active during all stages of development, but it is especially important in the first few days of life, when embryonic cells change into the different cell types that will make up the tissues and organs of the growing embryo. Chemicals can stop PRC2 from working, but they are not very precise and change how PRC2 works all over the genome. The UW researchers wanted to find a way to block PRC2 so that it would only affect one gene at a time.
David Baker and his team use AI to make a protein that will bind to PRC2 and stop it from using another protein to change the histones. Ruohola-Baker and Levy then joined this designed protein with a version of the Cas9 protein that didn’t work.
CRISPR is a way to change genes. Cas9 is a protein that is used in this process. Cas9 binds to RNA, which it then uses as an address tag. By making a specific “address-tag” RNA, scientists can use the system to bring Cas9 to a specific place in the genome and cut and splice genes there. In this experiment, however, the Cas9 protein’s ability to cut is turned off, so the genomic DNA sequence does not change.
So, it’s called dCas9, which means “dead.” But Cas9 still works as a “vehicle” to take cargo to a certain place. The dCas9-RNA construct was carrying the blocking protein that was made by AI. “Levy says, “dCas9 is like UBER. It will take you anywhere you want to go on the genome.” The guide RNA is like a person in an Uber who tells the driver where to go.”
In the new paper, Levy and her colleagues show that they were able to block PRC2 and turn on only four genes by using this technique. They also showed that by turning on just two genes, they could change induced pluripotent stem cells into placental progenitor cells.
Levy said, “This method lets us avoid bombarding cells with different growth factors, gene activators, and gene repressors to make them differentiate.” “Instead, we can target specific sites on the gene transcription promoters’ region, remove those marks, and let the cell do the rest in a natural, holistic way.”
Lastly, the researchers were able to show how the method can be used to find the location of specific PRC2-controlled regulatory regions where individual genes are turned on. Many of these don’t know where they are. In this case, they found a promoter area for a gene called TBX18. This area is called a TATA box. Even though it is thought that these promoter regions are within 30 DNA base pairs of the gene, they found that this gene’s promoter region was more than 500 base pairs away.
Ruohola-Baker said, “This was a very important find.” “There are TATA boxes all over the genome, but the important ones are close to the gene transcription site, and the others don’t seem to matter. This tool’s strength is that it can find the important TATA boxes that depend on PRC2, in this case.”
In both normal and abnormal cells, epigenetic changes decorate large parts of the genome. But it’s not clear what the smallest unit of epigenetic change is and how it works. Ruohola-Baker says, “With AI-designed proteins and CRISPR technology, we can now find the exact epigenetic marks that are important for gene expression, learn the rules, and use them to control cell function, drive cell differentiation, and create 21st century therapies.”
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