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New research has revealed that “random DNA” is actively transcribed in yeast, but remains largely inactive in mammalian cells, despite both organisms sharing a common ancestor and molecular mechanisms. This study involved inserting a synthetic gene in reverse order into yeast and mouse stem cells, which revealed significant differences in transcription activity. The findings suggest that, while yeast cells actively transcribe almost all genes, mammalian cells naturally repress transcription. This research not only challenges our understanding of genetic transcription across species, but also has implications for the future of genetic engineering and the discovery of new genes.
A new study reveals that in single-celled fungal yeast, “randomly DNA” is naturally active, while in mammalian cells, this DNA is deactivated as its natural state in mammalian cells, despite having a common ancestor billion years ago and the same basic molecular machinery.
The new finding revolves around the process by which the genetic instructions in DNA are first converted into a related material called RNA and then into proteins that form the body’s structures and signals. In yeast, mice, and humans, the first step in gene expression, transcription, occurs when the molecular “letters” of DNA (nucleobases) are read in one direction. While 80% of the human genome (the complete set of DNA in our cells) is actively decoded into RNA, less than 2% actually encodes genes that direct the construction of proteins.
So a long-standing mystery in genomics is what all this non-gene transcription accomplishes. Is it just noise, a side effect of evolution, or does it have functions?
A research team at New York University Langone Health attempted to answer the question by creating a large synthetic gene, with its DNA code in the reverse order of its natural parent. They then put synthetic genes into yeast and mouse stem cells and looked at the transcription levels in each. Published in the magazine Nature, The new study reveals that in yeast the genetic system is set up so that almost all genes are continuously transcribed, while the same “default state” in mammalian cells is that transcription is turned off.
Methodology and findings
Interestingly, the study authors say, the reverse order of the code meant that all the mechanisms that evolved in yeast and mammalian cells to turn transcription on or off were absent because the reversed code didn’t make sense. However, like a mirror image, the inverted code reflected some basic patterns observed in the natural code in terms of how often DNA letters were present, how close they were, and how often they were repeated. Since the inverted code was 100,000 molecular letters long, the team found that it randomly included many small stretches of previously unknown code that likely started transcription much more frequently in yeast and stopped it in mammalian cells.
“Understanding the default transcription differences between species “It will help us better understand which parts of the genetic code have functions and which are accidents of evolution,” said corresponding author Jef Boeke, PhD, Sol and Judith Bergstein Director of the Systems Genetics Institute at NYU Langone Health. “This, in turn, promises to guide yeast engineering to make new drugs, create new gene therapies or even help us find new genes buried in the vast code.”
The work lends weight to the theory that the very active transcriptional state of yeast is established so that foreign DNA, rarely injected into the yeast, for example by a virus As it copies itself, it is likely to be transcribed into RNA. If that RNA builds a protein with a useful function, the code will be preserved by evolution as a new gene. Unlike the single-celled yeast organism, which can afford risky new genes that drive more rapid evolution, mammalian cells, as part of bodies with millions of cooperating cells, have less freedom to take in new DNA each time a cell find a virus. Many regulatory mechanisms protect the delicately balanced code as it is.
Great DNA
The new study had to take into account the size of DNA strands, with 3 billion “letters” included in the human genome, and some genes are 2 million letters long. While famous techniques allow for letter-by-letter changes, some engineering tasks are more efficient if researchers build DNA from scratch, with extensive changes made to large swaths of preassembled code exchanged in a cell rather than its natural counterpart. Because human genes are so complex, Boeke’s lab first developed its “genome writing” method in yeast, but then recently adapted it to the genetic code of mammals. The study authors use yeast cells to assemble long sequences of DNA in a single step and then introduce them into mouse embryonic stem cells.
For the current study, the research team addressed the question of how widespread transcription is throughout evolution by introducing a synthetic 101-kilobase stretch of engineered DNA: the human hypoxanthine phosphoribosyl transferase 1 (HPRT1) gene in order reverse coding. They observed widespread activity of the gene in yeast despite the nonsense code’s lack of promoters, fragments of DNA that evolved to signal the start of transcription.
Additionally, the team identified small sequences in the inverted code, repeated stretches of adenosine and thymine building blocks, that are known to be recognized by transcription factors, proteins that bind to DNA to initiate transcription. These sequences, which are only 5 to 15 letters long, could easily occur randomly and may partly explain the default state of highly active yeast, the authors said.
On the contrary, the same code reversed, inserted into the genome of mouse embryonic stem cells, they did not cause widespread transcription. In this scenario, transcription was repressed even though the evolved CpG dinucleotides, known to actively inhibit (silence) genes, were not functional in the reverse code. The team hypothesizes that other building blocks in the mammalian genome may restrict transcription much more than in yeast, perhaps by directly recruiting a group of proteins (the Polycomb complex) that are known to silence genes.
“The closer we get to introducing a nonsense DNA ‘genome’ into living cells, the better they will be able to compare it to the actual evolved genome,” said first author Brendan Camellato, a graduate student in Boeke’s lab. “This could lead to a new frontier of engineered cell therapies, as the ability to insert increasingly longer synthetic DNA allows for a better understanding of what insertions genomes will tolerate and perhaps the inclusion of one or more larger, complete engineered genes. “
Reference: “Synthetic inverted sequences reveal predetermined genomic states” by Brendan R. Camellato, Ran Brosh, Hannah J. Ashe, Matthew T. Maurano and Jef D. Boeke, March 6, 2024, Nature.
DOI: 10.1038/s41586-024-07128-2