Complex cell engineering shows genome is resilient to changes, Wellcome Sanger Institute researchers and colleagues find

Scientists have achieved the most complex engineering of human cell lines ever – and it has shown our genomes are more resilient to significant structural changes than previously thought.

Multiple versions of human genomes were created in cell lines, each with different structural changes by researchers from the Wellcome Sanger Institute, Imperial College London, Harvard University in the US and their collaborators, using CRISPR prime editing.

They then deployed genome sequencing to analyse the genetic effects of these structural variations on cell survival. It showed that as long as essential genes remain intact, our genomes can tolerate significant structural changes, including large deletions of the genetic code.

The study, published on 30 January in Science, could help us explore the role of structural variation in developmental diseases and cancer.

Structural variation relates to a change in the structure of an organism’s genome, such as deletions, duplications and inversions of the genetic sequence.

These can be significant changes, affecting hundreds to many thousands of nucleotides – the basic building blocks of DNA and RNA.

Dr Jonas Koeppel, co-first author previously at the Wellcome Sanger Institute, and now at the University of Washington, said: “If the genome was a book, you could think of a single nucleotide variant as a typo, whereas a structural variant is like ripping out a whole page. These structural variants are known to play roles in developmental diseases and cancer, but it has been difficult to study them experimentally. Through creative and collaborative thinking, we’ve been able to do complex engineering in human cells that no-one has done before. By shuffling the genomes of human cell lines at large scale, we’ve shown that our genomes are flexible enough to tolerate significant structural changes. These tools will help focus future studies into structural variations and their roles in disease.”

Using prime editing, the researchers inserted a recognition sequence into the genomes of the human cell lines to target with recombinase – an enzyme that enabled them to ‘shuffle’ the genome.

Inserting these recombinase handles into hundreds and thousands of identical sequences in the genome meant that with a single prime editor they were able to integrate up to almost 1,700 recombinase recognition sites into each cell line. This led to more than 100 random large-scale genetic structural changes per cell–- representing the first time that it’s been possible to ‘shuffle’ a mammalian genome at scale.

Using sequencing, they then took ‘snapshots’ of the human cells and their ‘shuffled’ genomes over the course of a few weeks, watching which cells survived and which died.

When structural variation deleted essential genes, the cells died, as expected. But they found groups of cells with large-scale deletions in the genomes that avoided essential genes were able to survive.

RNA sequencing of the human cell lines to measure gene expression – meaning activity – showed large-scale deletions of the genetic code, especially in non-coding regions, did not seem to impact the gene expression of the rest of the cell.

The study suggests that human genomes are extremely tolerant of structural variation, including variants that change the position of hundreds of genes, providing essential genes are not deleted.