RNA-guided systems refer to proteins that target specific nucleic acid sequences by binding to guide RNA (gRNA). In nature, these systems allow a single enzyme to target different sequences based on the gRNA it carries. For example, CRISPR-Cas9 can precisely target and edit specific DNA sequences using designed gRNAs. Over the past decade, CRISPR-Cas systems from bacteria and archaea have revolutionized basic biological sciences, molecular medicine, and biotechnology through applications in genome and epigenome editing, RNA editing, gene regulation, and pathogen detection.
In addition to various CRISPR-Cas systems, recent discoveries by Feng Zhang's team and others have identified RNA-guided systems with gene-editing potential, such as the OMEGA system, Fanzor system, and CRISPR-associated transposases. However, these systems were discovered based on their evolutionary relationship with CRISPR-Cas systems. To directly expand the diversity of RNA-guided systems, more general database mining methods are needed. On February 27, 2025, Feng Zhang's team published a study in Science titled "TIGR-Tas: A family of modular RNA-guided DNA-targeting systems in prokaryotes and their viruses."
The study began with the gRNA interaction domain of the Cas9 protein and, through iterative structural and sequence homology mining, identified an ancient family of RNA-guided DNA-targeting proteins in phages and parasitic bacteria. This system, named TIGR-Tas, uses tigRNA to guide Tas proteins to specific DNA sites. It can be reprogrammed to target any DNA sequence in the genome (without relying on PAM sequences) and directly modify the target DNA. Importantly, compared to other RNA-guided systems like CRISPR, the TIGR-Tas system is highly compact (Tas proteins are on average only a quarter the size of Cas9), making it more advantageous for gene-editing therapies.
Feng Zhang stated that the TIGR-Tas system is highly versatile, with many different functions. The study found that TIGR-associated proteins (Tas) have an RNA-binding component that interacts with gRNA to guide it to specific genomic locations. Some Tas proteins also contain adjacent DNA-cutting segments that cleave DNA at specific sites. This modularity facilitates the development of gene-editing tools, allowing researchers to introduce useful new functions into natural Tas proteins.
As a pioneer of CRISPR gene-editing technology, Feng Zhang first adapted the bacterial CRISPR system into a gene-editing tool—CRISPR-Cas9—in January 2013, achieving CRISPR gene editing in mammalian cells for the first time. This breakthrough transformed modern biology and ushered in a new era of gene editing. Since then, Zhang and his team have discovered and engineered multiple gene-editing systems.
Zhang emphasized that nature is incredibly diverse, and his team continues to explore this diversity to uncover new biological mechanisms and apply them to manipulate biological processes. In this latest study, to discover new programmable gene-editing systems, the team focused on the structural features of the Cas9 protein that interact with gRNA. This RNA-guided property makes Cas9 a powerful gene-editing tool because RNA binds to DNA or other RNA through precise base-pairing mechanisms, making it easier to reprogram.
The team systematically screened hundreds of millions of known or predicted protein structures to identify candidates with RNA-binding domains similar to Cas9. To uncover distantly related proteins, they employed an iterative research strategy: starting with Cas9, they identified an RNA-binding protein called IS110; focusing on IS110's RNA-binding domain, they resolved its three-dimensional structural features; and based on these new features, they refined their screening criteria and expanded their search.
Using this strategy, the team identified a large number of Cas9 distant relatives, making it difficult to analyze these highly diverse proteins using standard phylogenetic methods reliant on conserved sequences. Instead, they turned to artificial intelligence (AI) to understand these proteins. Using a protein large language model (Protein Large Language Model), the team clustered the proteins based on their potential evolutionary relationships. Notably, one group stood out—its encoded genes had regularly spaced repeat sequences reminiscent of CRISPR systems, where CRISPR stands for "Clustered Regularly Interspaced Short Palindromic Repeats."
The team named this newly discovered system TIGR-Tas, which consists of TIGR arrays and Tas proteins. They identified over 20,000 different Tas proteins, most of which are found in phages, archaeal viruses, and parasitic bacteria. The TIGR arrays are processed into 36-nucleotide gRNAs—tigRNAs—each containing two short spacer regions (spacer A and B) that target both strands of DNA. This dual targeting flexibility avoids self-targeting and eliminates the need for PAM sequences.
Next, the team experimentally tested dozens of Tas proteins, demonstrating that some could be reprogrammed to precisely target and cut DNA in human cells. For example, TaTasR and ParTasR induced gene editing at multiple loci in human cells, with editing efficiencies of up to 3.6%.
Unlike CRISPR-Cas systems, which can only target DNA sequences flanked by short PAM motifs, the TIGR-Tas system has no such requirement, meaning it can theoretically target any site in the genome. Additionally, the compact size of Tas proteins (averaging only a quarter the size of Cas9) makes them easier to deliver, potentially overcoming delivery challenges in clinical gene-editing therapies.
In summary, the TIGR-Tas system fills a gap in the evolution of RNA-guided mechanisms. Its PAM independence, dual-strand targeting capability, and compact size offer new directions for gene editing. Although editing efficiency needs improvement, this new system could become an important complement to the CRISPR-Cas gene-editing toolbox.
Feng Zhang's team is currently exploring the natural functions of the TIGR-Tas system in viruses and how it can be applied to research and disease treatment. They have determined the molecular structure of a Tas protein capable of gene editing in human cells and are working to enhance its functionality. Additionally, they have noted connections between the TIGR-Tas system and certain RNA-processing proteins in human cells.
Resource: https://www.science.org/doi/10.1126/science.adv9789