CRISPR's Comrades in Arms Discovered in Microbial Genomes

CRISPR doesn’t stand against bacteriophages – viruses that infect bacteria – all by itself. No, this bacterial immune system has fellows deep in the microbial genome’s ranks. When these ranks were subjected to systematic inspection by researchers at the Weizmann School of Science, they were found to include 10 previously unknown immune defense systems. Some of these systems, like the CRISPR system, may be of use in genome editing.

Besides CRISPR, bacteria have been known to rely on the protections afforded by restriction-modification systems, which target specific sequences on invading phages, and abortive infection systems, which lead to cell death or metabolic arrest upon infection. Yet other bacterial immune defense systems have been thought to exist.

The existence of additional defense systems could help explain how bacteria manage to cope with phages that have “anti-CRISPR” proteins that cancel CRISPR activity. Also, additional defense systems may lurk in “defense islands,” genomic regions where defense-related genes tend to cluster. Such defense islands may harbor genes of unknown function that participate in antiphage defense.

A systematic exploration of defense islands was in order, decided a team of Weizmann Institute scientists led by Rotem Sorek, Ph.D., an associate professor of molecular genetics. The team began its work by creating a computer program that would scan all the bacterial genomes that have ever been sequenced – around 50,000 genomes in all.

Rather than look for sequences with predefined characteristics, the team searched for the “statistical signatures” of genes involved in defense. For example, the team considered whether a gene was located a defense island.

Aware that immune system genes rarely work alone – even in bacteria – the researchers developed complex computer analytic methods so as to understand which genes join forces and work together to form a defense system.

Details of this work were presented in the journal Science, in an article entitled, “Systematic Discovery of Antiphage Defense Systems in the Microbial Pangenome.” The article also described how the Weizmann scientists narrowed down defense gene candidates from millions to several hundred.

“Candidate defense systems were systematically engineered and validated in model bacteria for their antiphage activities,” the article’s authors wrote. “We report nine previously unknown antiphage systems and one antiplasmid system that are widespread in microbes and strongly protect against foreign invaders.”

Rather than attempting to isolate the genetic sequences from hundreds of different bacteria, the scientists turned to synthetic biology: getting the genes made to order. They sent the strings of gene code – totaling something like 400,000 bases, or “letters” of genetic code – to a commercial lab where dozens of different multigene systems were synthesized for testing. These synthetic systems were inserted into lab bacteria that had had their natural immune systems inactivated. The bacteria were then exposed to phages and other infective elements to see if the transplanted defense system was a viable one.

“The systems we discovered are unlike anything we had seen before,” said Dr. Sorek. “But among them, we think, are one or two that might have the potential to increase the gene-editing toolbox, and others that point to the origins of the human immune system.”

Some of the defense genes identified in the study appear to have been borrowed from nondefensive bacterial systems. “These include systems that adopted components of the bacterial flagella and condensin complexes,” the article’s authors indicated. “Our data also suggest a common, ancient ancestry of innate immunity components shared between animals, plants, and bacteria.”

The researchers still don’t know how the new bacterial immune systems function, and some according to Dr. Sorek, “seem to have surprising functions that we are now starting to investigate.” One of these systems contains Toll-interleukin receptor (or TIR) domains. TIR domains were already known to be involved in immune systems, but until now not those of microorganisms.

“The fact that we managed to find 10 new bacterial defense systems implies there are even more out there,” noted Dr. Sorek. “My lab is continuing to search for new ones. In addition, we are starting to focus on several of the more promising ones to understand how they function.”

Dr. Sorek suggested that the new discoveries are exciting because of the new windows they provide on the evolution of immune systems and the eternal battle between viruses and the organisms they infect. But he also believes that some may turn out to be powerful tools for biological research.

“Every immune system, by definition, needs to target invading elements in a very specific yet flexible way, and we can use this targeting for biotechnological purposes—as we’ve done with CRISPR and with restriction enzymes before it,” he stated. “Any one of the new systems we found might be the next gene-editing tool—or perhaps even the foundation of even more exciting molecular tools.”