Screen of human genome reveals set of genes essential for cellular viability

Diagram of the crystal structure of Cas9 in complex with guide RNA and its target DNA.

Diagram of the crystal structure of Cas9 in complex with guide RNA and its target DNA.

Image: Courtesy of H. Nishimasu, et al. (from Wikipedia).

October 15, 2015

Tags: Sabatini LabCancerGenetics + Genomics

CAMBRIDGE, Mass. – Using two complementary analytical approaches, scientists at Whitehead Institute and Broad Institute of MIT and Harvard have for the first time identified the universe of genes in the human genome essential for the survival and proliferation of human cell lines or cultured human cells.

Their findings and the materials they developed in conducting the research will not only serve as invaluable resources for the global research community but should also have application in the discovery of drug-targetable genetic vulnerabilities in a variety of human cancers.

Scientists have long known the essential genes in microorganisms, such as yeast, whose genomes are smaller and more easily manipulated. Most common yeast strains, for example, are haploid, meaning that genes exist in single copies, making it fairly simple for researchers to eliminate or “knock out” individual genes and assess the impact on the organism. However, owing to their greater complexity, diploid mammalian genomes, including the human genome, have been resistant to such knockout techniques—including RNA interference, which is hampered by off-target effects and incomplete gene silencing.

Now, however, through use of the breakthrough CRISPR (for clustered regularly interspersed short palindromic repeats) genome editing system, researchers in the labs of Whitehead Member David Sabatini and Broad Institute Director Eric Lander have been able to generate a genome-wide library of single-guide RNAs (sgRNAs) to screen for and identify the genes required for cellular viability. The sgRNA library targeted slightly more than 18,000 genes, of which approximately 10% proved to be essential. These findings are reported online this week in the journal Science.

“This is the first report of human cell-essential genes,” says Tim Wang, a graduate student in the Sabatini and Lander labs and first author of the Science paper. “This answers a question people have been asking for quite a long time.”

As might have been expected, Wang says that many of the essential genes are involved in fundamental biological processes, including DNA replication, RNA transcription, and translation of messenger RNA. But, as Wang also notes, approximately 300 of these essential genes are of a class not previously characterized, are largely located in the cellular compartment known as the nucleolus, and are associated with RNA processing. Wang says the precise function of these genes is the subject of future investigation.

To validate the results of the CRISPR screens, the group took the added step of screening for essential genes in a unique line of haploid human cells. Using an approach known as gene-trap mutagenesis (a method pioneered in part by former Whitehead Fellow Thijn Brummelkamp) in the haploid cells and comparing it to the CRISPR results, the researchers found significant, consistent overlap in the gene sets found to be essential. In a final step, the group tested their approaches in cell lines derived from two cancers, chronic myelogenous leukemia (CML) and Burkitt’s lymphoma, both of which have been extensively studied. The novel method not only identified the essentiality of the known genes—in the case of CML, it hit on the BCR and ABL1 genes, whose translocation is the target of the successful drug Gleevec—but also highlighted additional genes that may be therapeutic targets in these cancers.

“The ability to zero in on the essential genes in the highly complex human system will give us new insight into how diseases, such as cancer, continue to resist efforts to defeat them,” Lander says.

Wang, Lander, and Sabatini are enthusiastic about the potential applications of their work, as it should accelerate the identification of cancer drug targets while enhancing our understanding of the evolution of drug resistance, a major contributor to therapeutic failure. The researchers attribute this vast potential to the rigor that CRISPR brings to human genetics.

“This is really the first time we can reliably, accurately, and systematically study genetics in mammalian cells,” Sabatini says. “It’s remarkable how well it’s working.”

 

This work was supported by the National Institutes of Health (grant CA103866), the National Human Genome Research Institute (grant 2U54HG003067-10), the National Science Foundation, the MIT Whitaker Health Sciences Fund, and the Howard Hughes Medical Institute.

 

Written by Matt Fearer

 

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David Sabatini's primary affiliation is with Whitehead Institute for Biomedical Research, where his laboratory is located and all his research is conducted. He is also a Howard Hughes Medical Institute investigator and a professor of biology at Massachusetts Institute of Technology.

 

About Whitehead Institute

Whitehead Institute is a world-renowned non-profit research institution dedicated to improving human health through basic biomedical research. Wholly independent in its governance, finances, and research programs, Whitehead shares a close affiliation with Massachusetts Institute of Technology through its faculty, who hold joint MIT appointments.

 

About the Broad Institute of MIT and Harvard

The Eli and Edythe L. Broad Institute of MIT and Harvard was launched in 2004 to empower this generation of creative scientists to transform medicine. The Broad Institute seeks to describe all the molecular components of life and their connections; discover the molecular basis of major human diseases; develop effective new approaches to diagnostics and therapeutics; and disseminate discoveries, tools, methods and data openly to the entire scientific community.

Founded by MIT, Harvard and its affiliated hospitals, and the visionary Los Angeles philanthropists Eli and Edythe L. Broad, the Broad Institute includes faculty, professional staff and students from throughout the MIT and Harvard biomedical research communities and beyond, with collaborations spanning over a hundred private and public institutions in more than 40 countries worldwide.

 

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Full Citation:

“Identification and characterization of essential genes in the human genome”

Science, October 15, 2015 (online)

Tim Wang (1,2,3,4), Kıvanç Birsoy (1,2,3,4,5), Nicholas W. Hughes (3), Kevin M. Krupczak (2,3,4), Yorick Post (2,3,4), Jenny J. Wei (1,2), Eric S. Lander (1,3,6, 8), and David M. Sabatini (1,2,3,4,7,8)

1. Department of Biology, Massachusetts Institute of Technology, Cambridge MA 02139, USA

2. Whitehead Institute for Biomedical Research, Nine Cambridge Center, Cambridge, MA 02142, USA

3. Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA 02142, USA

4. David H. Koch Institute for Integrative Cancer Research at MIT, Cambridge, MA 02139, USA

5. Present address: Laboratory of Metabolic Regulation and Genetics, The Rockefeller University, New York, NY 10065, USA

6. Department of Systems Biology, Harvard Medical School, Boston, MA 02115, USA

7. Howard Hughes Medical Institute, Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA

8. These authors contributed equally to this work

Whitehead Institute is a world-renowned non-profit research institution dedicated to improving human health through basic biomedical research.
Wholly independent in its governance, finances, and research programs, Whitehead shares a close affiliation with Massachusetts Institute of Technology
through its faculty, who hold joint MIT appointments.

© Whitehead Institute for Biomedical Research         Nine Cambridge Center    Cambridge, MA 02142