Scientists have discovered a way to switch genes on or off inside yeast and human cells by controlling the point at which DNA is copied into messenger RNA, according to a study published in the journal ACS Synthetic Biology.
Researchers from Massachusetts Institute of Technology (MIT) say that this discovery could enable scientists to better understand the role of the genes, make it easier to engineer cells and lead to better drugs and treatments.
The new method is based on a system called CRISPR, which is made up of two components. The first is a protein that “binds to and slices DNA,” while the second component is a short strand of RNA that guides the protein to the required location within the genome.
“The CRISPR system is quite powerful in that it can be targeted to different DNA binding regions based on simple recoding of these guide RNAs,” says Timothy Lu, assistant professor of electrical engineering, computer science and biological engineering at MIT.
“By simply reprogramming the RNA sequence, you can direct this protein to any location you want on the genome or on a synthetic circuit,” he adds.
For the study, the researchers decided to use CRISPR to control gene transcription – the process by which a sequence of DNA is copied into messenger RNA (mRNA). This carries out instructions from the gene.
Proteins known as “transcription factors” heavily regulate transcription, explain the researchers. These proteins attach to certain DNA sequences in the gene’s “promoter region,” then initiate or block the enzymes needed in order to copy the gene into mRNA.
To use the CRISPR as a transcription factor, the researchers modified the standard CRISPR protein, called Cas9, to ensure it no longer “snipped” DNA after binding to it. A segment was also added to the protein that triggers or inhibits gene expression by changing the transcriptional machinery of the cell.
The researchers also delivered a gene for an “RNA guide” to the target cells in order to push Cas9 to the correct region. The RNA guide works by corresponding to a DNA sequence on the promoter of the gene they want to activate.
The outcome of the study revealed that once the RNA guide and the Cas9 protein linked inside the target cell, the researchers could accurately target the correct gene and switch on transcription.
Additionally, they were surprised to find that this process could also be used to block gene transcription when aimed at a different area of the gene.
“This is nice in that it allows you do to positive and negative regulation with the same protein, but with different guide RNAs targeted to different positions in the promoter,” says Prof. Lu.
The researchers add that this system is much more flexible and easier to use compared with other transcription-control systems based on DNA-binding proteins.
Prof. Lu says:
“There’s a lot of flexibility with CRISPR, and it really comes from the fact that you don’t have to spend any more time doing protein engineering. You can just change the nucleic acid sequence of the RNAs.
I think it is going to make it a lot easier to build synthetic circuits. It should increase the scale and the speed at which we can build a variety of synthetic circuits in yeast cells and mammalian cells.”
The new system has also been created so that it can be induced by certain small molecules that can be added to the cell, such as sugars, the researchers say.
To carry out this process, the genes were engineered for guide RNAs to ensure they are only produced in presence of the small molecule. They add that if there is no small molecule, there is no guide RNA, which means the target gene is undisturbed.
Prof. Lu notes that this type of control may prove useful for further studies looking at the role of naturally occurring genes by switching them on and off throughout different points of disease progression or development.
He adds that the next steps from this research are to build further advanced synthetic circuits that can “make decisions” based on a variety of inputs from a cell’s environment.
“We’d like to be able to scale this up and demonstrate the most complex circuits that anyone’s ever built in yeast and mammalian cells,” Prof. Lu says.