CRISPR-Fine Screen of Human Genome Identifies Essential Genes

For the first time, a comprehensive set of cell-essential genes in the human genome has been compiled. This feat, which not only points to deeper studies of human genetics but also suggests a strategy for finding vulnerabilities in cancer cells, was achieved thanks to the CRISPR-Cas9 gene-editing system. This system proved sufficiently rigorous to succeed where other gene-knockout approaches, such as RNA interference, had failed, mainly due to off-target effects and incomplete gene silencing.

Using CRISPR-Cas9, scientists at the Whitehead Institute and the Broad Institute constructed a genome-wide single-guide RNA (sgRNA) library to screen for genes required for proliferation and survival in a human cancer cell line. The sgRNA library targeted slightly more than 18,000 genes, of which approximately 10% proved to be essential.

These results appeared October 15 in the journal Science, in an article entitled, “Identification and characterization of essential genes in the human genome.” The article describes how the results obtained with the CRISPR-Cas9 approach were validated by another approach, gene-trap mutagenesis, which involved screening for essential genes in a unique line of haploid human cells.

The set of essential genes identified in the current study largely consists of genes that encode components of fundamental pathways, are expressed at high levels, and contain few inactivating polymorphisms in the human population. For example, the genes associated with fundamental pathways include many that participate in DNA replication, RNA transcription, and translation of messenger RNA.

Of the essential genes found to be involved in RNA processing, about 300 were previously uncharacterized. Moreover, the products of these genes were determined to be largely localized to the nucleolus.

Additional findings pertain specifically to cancer. “[Our screens],” the authors indicated in the Science article, “revealed differences specific to each cell line and cancer type that reflect the developmental origin, oncogenic drivers, paralogous gene expression pattern, and chromosomal structure of each line.”

This work, which was led by Whitehead Member David Sabatini and Broad Institute Director Eric Lander, used the CRISPR-Cas9 approach to investigate cell lines derived from two cancers, chronic myelogenous leukemia (CML) and Burkitt’s lymphoma. 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,” said Dr. Lander.

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