Tying Human Genes to Their Function Using CRISPR Technology

Researchers have expanded on data from the Human Genome Project, using their single-cell sequencing tool – Perturb-seq – to link each human gene to its function and create a free resource for other researchers.

Comprehensive roadmap links genes to their function

An international collaboration of researchers headed by Whitehead Institute member Professor Jonathan Weissman has expanded on data collected from the 2003 Human Genome Project, creating a comprehensive roadmap that links human genes to their function. Weissman is also a professor of biology at Massachusetts Institute of Technology (MIT), and an investigator at the Howard Hughes Medical Institute.

The full database was published on June 9 in the journal Cell and made freely available on the Weissman Lab website, providing opportunities for other researchers to make use of this vast resource in their own studies.

“It’s a big resource in the way the human genome is a big resource, in that you can go in and do discovery-based research,” explains Weissman.

Former Weissman Lab postdoc and co-senior author Dr. Tom Norman adds, “I think this dataset is going to enable all sorts of analyses that we haven't even thought up yet by people who come from other parts of biology and suddenly they just have this available to draw on.”

Revolutionizing new techniques

This study improved upon an approach known as Perturb-seq, first published in 2016 by Weissman and colleagues at MIT. Perturb-seq allows researchers to study the implications of turning genes “on” or “off” in great detail.

Initially, it was only possible to study small sets of genes at one time, and this came at great expense. However, researchers went on to publish a proof-of-concept paper in which they combined this technique with CRISPR/Cas9 genome editing to create a new, scaled-up approach.

In the new paper, the researchers delved deeper into this method and expanded it to the entire human genome using blood cancer cell lines and non-cancerous retina cells. Applying this technique to over 2 million cells, they were able to produce a detailed resource mapping genotypes to their corresponding phenotypes.

Utilizing the data

After assembling this ground-breaking dataset that permits such unbiased analysis, the researchers then asked themselves the question, “What do you actually do with it?”.

Firstly, they focused their attention on looking into genes with as-yet-unknown functions. By comparing genes with known functions to those that were unknown, they were able to tease out genes with similar transcriptional outcomes.

For example, after noticing the effects of a mutation in the gene C7orf26, researchers drew comparisons to the removal of other genes that resulted in a similar phenotype. These proteins were discovered to form part of the protein complex Integrator, responsible for creating small nuclear RNAs. This was previously thought to be made of a complex of 14 proteins, but the use of Perturb-seq confirmed C7orf26 as the fifteenth component.

Furthermore, the researchers were able to use this data to examine how mitochondria respond to stress and delve deeper into the question of why they have their own DNA. Joseph Replogle, MD-PhD student in Weissman’s lab and co-first author of the paper said, “People have been interested for a long time in how nuclear and mitochondrial DNA are coordinated and regulated in different cellular conditions, especially when a cell is stressed.”

The findings showed that when different mitochondria-related genes were perturbed, the nuclear genome responded in a similar way to various genetic changes. However, the mitochondrial genome responses were much more variable.

“A big-picture takeaway from our work is that one benefit of having a separate mitochondrial genome might be having localized or very specific genetic regulation in response to different stressors,” Replogle added.

Future prospects

The researchers that developed Perturb-seq hope to build upon their technique further, branching out to study other cell types beyond their blood cancer cell lines. They aim to examine many other genes in detail within their roadmap and hope other groups of researchers do the same.

“This really is the culmination of many years of work by the authors and other collaborators, and I’m really pleased to see it continue to succeed and expand,” Dr. Norman summarized.

Reference: Replogle JM, Saunders RA, Pogson AN, et al. Mapping information-rich genotype-phenotype landscapes with genome-scale Perturb-seq. Cell. 2022;0(0). doi: 10.1016/j.cell.2022.05.013