Genetic interaction map of essential genes in M. tuberculosis

Researchers at Rockefeller University, in Jeremy Rock's Laboratory of Host-Pathogen Biology, have published in Nature a map of genetic interactions among essential genes of Mycobacterium tuberculosis (Mtb) and the model species Mycobacterium smegmatis (Msm). The paper is available via dx.doi.org, and the university announced it on 4 November 2026 dx.doi.org. The maps cover nearly all essential genes of both species and, according to the authors, reveal thousands of previously unrecognized interactions dx.doi.org.

Mtb causes tuberculosis, which the authors call the world's leading infectious cause of death. Many of its genes remain uncharacterized, which limits understanding of the disease and constrains drug development dx.doi.org. Treatment typically requires several antibiotics taken over months, and drug-resistant infections take longer to clear.

Dual-sgRNA CRISPRi platform

A genetic interaction is a combined phenotype that deviates from the expectation based on single-gene perturbations. Negative interactions, including synthetic lethality, give a more severe phenotype than expected, and positive ones a milder one dx.doi.org. Earlier transposon-based approaches in Mtb were typically restricted to non-essential genes, which excludes most antibiotic targets dx.doi.org.

The team adapted the CombiGEM framework to the Mtb CRISPR-interference platform. Each sgRNA received a unique barcode, and the cloning workflow keeps the sgRNA–barcode link and places paired barcodes close together. This allows the abundance of each dual-sgRNA strain to be quantified by barcode sequencing dx.doi.org. The authors also implemented a statistical pipeline for detecting interactions with high confidence dx.doi.org.

Results and validation

The maps recapitulate known biology and identify functional connections between essential genes and drug targets dx.doi.org. The authors also ran structure-based protein–protein interaction predictions for about 600,000 gene pairs represented in the maps. They report that a subset of the interactions likely reflects physical associations between proteins dx.doi.org.

Selected interactions were tested experimentally. The validation covered drug–drug interactions and roles in the electron transport chain for several genes of unknown function dx.doi.org. The authors state that the work provides a generalizable framework for dual-gene perturbation and defines the architecture of essential-gene networks in mycobacteria dx.doi.org.

All experiments were done in bacterial cultures. The paper does not report testing of new drugs or treatment regimens. Rock said this is the first comprehensive map of interactions among essential genes in a pathogen.

Rockefeller University maps genetic interactions of essential genes in M. tuberculosis

Dual-sgRNA CRISPRi platform

Results and validation

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