Remember CRISPR (clustered regularly interspaced short palindromic repeats) – that new gene-editing system which is faster and cheaper than anything that came before it? CRISPR is derived from bacterial systems which uses guide RNA to target a specific sequence on a DNA strand. It is coupled with a Cas (CRISPR Associated) protein which can do things like cleave the DNA at the targeted location. We are really just at the beginning of exploring the potential of this new system, in both research and therapeutics.

Well – we may already have something better than CSRISP: TIGR-Tas. This is also an RNA-based system for targeting specific sequences of DNA and delivering a TIGR associated protein to perform a specific function. TIGR (Tandem Interspaced Guide RNA) may have some useful advantages of CRISPR.

As presented in a new paper, TIGR is actually a family of gene editing systems. It was discovered not by happy accident, but by specifically looking for it. As the paper details: “through iterative structural and sequence homology-based mining starting with a guide RNA-interaction domain of Cas9”. This means they started with Cas9 and then trolled through the vast database of phage and parasitic bacteria for similar sequences. They found what they were looking for – another family of RNA-guided gene editing systems.

Like CRISPR, TIGR is an RNA guided system, and has a modular structure. Different Tas proteins can be coupled with the TIGR to perform different actions at the targeted site. But there are several potential advantages for TIGR over CRISPR. Like CRISPR it is RNA guided, but TIGR uses both strands of the DNA to find its target sequence. This “dual guided” approach may lead to fewer off-target errors. While CRISPR works very well, there is a trade-off in CRISPR systems between speed and precision. The faster it works, the greater the number of off-target actions – like cleaving the DNA in the wrong place. The hope is that TIGR will make fewer off-target mistakes because of better targeting.

TIGR also has “PAM-Independent targeting”. What does that mean? PAM stands for protospacer adjacent motifs – these are short DNA sequences, about 6 base pairs, that exist next to the sequence that his being targeted by CRISPR. The Cas9 protease will not function without the PAM. It appears to have evolved so that the bacteria using CRISPR as an adaptive immune system can tell self from non-self, as invading bacteria or viruses will have the PAM sequences, but the native DNA will not. The end result is that CRISPR needs PAM sequences in order to function, but the TIGR system does not. This makes the TIGR system much more versatile.

I saved what is potentially the best advantage for last – Tas proteins are much smaller than Cas proteins, about a quarter of the size. At first this might not seem like a huge advantage, but for some applications it is. One of the main limiting factors for using CRISPR therapeutically is getting the CRISPR-Cas complex into human cells. There are several available approaches – physical methods like direct injection, chemical methods, and viral vectors. Specific methods, however, generally have a size limit on the package they can deliver into a cell. Adeno-associated vectors (AAVs) for example have lots of advantaged but only can deliver relatively small payloads. Having a much more compact gene-editing system, therefore, is a huge potential advantage.

When it comes to therapeutics, the delivery system is perhaps the greater limiting factor than the gene targeting and editing system itself. There are currently two FDA indications for CRISPR-based therapies, both for blood disorders (sickle cell and thalassemia). For these disorders bone marrow can be removed from the patient, CRISPR is then applied to make the desired genetic changes, and then the bone marrow is transplanted back into the patient. In essence, we bring the cells to the CRISPR rather than the CRISPR to the cells. But how do we deliver CRISPR to a cell population within a living adult human?

We use the methods I listed above, such as the AAVs, but these all have limitations. Having a smaller package to deliver, however, will greatly expand our options.

The world of genetic engineering is moving incredibly fast. We are taking advantage of the fact that nature has already tinkered with these systems for hundreds of millions of years. There are likely more systems and variations out there for us to find. But already we have powerful tools to make precise edits of DNA at targeted locations, and TIGR just adds to our toolkit.