Scientists have taken an important step toward understanding how CRISPR systems work, particularly those known as type IV-A systems, which work differently than most other systems. These systems use unique ways to manage genetic material without cutting it. A research team led by Professor Patrick Pausch, Dr. Lina Malinauskaite, Dr. Rafael Pinilla-Redondo and Professor Lennart Randau, including researchers from Vilnius University, Philipps University of Marburg and the University of Copenhagen, used advanced imaging methods to reveal new for detailed information about these systems. Their findings were published in the journal Nature Communications.
Unlike other CRISPR systems, which cut DNA to render it useless, the Type IV-A system works by stopping the process of converting genetic material into RNA molecules, a step needed to produce proteins in cells. This type of interference is particularly useful in controlling genetic competition and regulating genes. The scientists focused on understanding how these systems recognize DNA targets and introduce a special protein called DinG helicase, which unwinds the DNA strand so that it can be processed further to perform the task.
Speaking about the work, Dr. Malinauskaitė said: “Our results reveal the detailed processes behind type IV-A CRISPR mechanisms and show how they function in unique ways. This understanding can help us develop tools to edit genetic material and regulate genes in new ways.
The researchers used cryo-electron microscopy, a method of freezing a sample to ultra-low temperatures to capture its structure at high resolution, to map the structure of two different versions of the Type IV-A system. One was from a bacterium called Pseudomonas oilovorans, and the other was from Klebsiella pneumoniae. The researchers found that these systems have a shrimp-like shape, in which protein components form a backbone that supports the guide RNA, guiding the system to a specific DNA target and binding to the target DNA. Specific proteins such as Cas8 and Cas5 play a key role in ensuring that the system locks onto the correct DNA sequence. The differences in these proteins indicate that each version works slightly differently, allowing them to adapt to different needs.
Another important discovery was how the systems recruit DinG helicase, a protein that helps them interfere with genetic processes. One system uses narrow interaction zones (small areas where proteins are connected) to attach the protein, while the other system has more extensive connections involving multiple proteins. These differences indicate that systems have evolved to meet the different challenges of managing DNA.
The researchers also highlighted similarities and differences between these systems and other systems that combine RNA and DNA. Although some processes may look familiar, the way these systems use DinG helicases sets them apart. This change reflects the flexibility and adaptability of CRISPR systems over time, demonstrating their evolutionary success in processing genetic material.
Experts believe the research has practical applications beyond understanding genetics. Professor Pausch noted: “The compact design of the Type IV-A system makes it ideal for creating new tools for editing genomes, especially where space is limited, such as in virus-based delivery systems.”
By the end of the study, the scientists had a clearer understanding of how these systems operate, offering potential for future applications. The unique design and mechanisms of the Type IV-A system could be used to develop advanced tools for medical and agricultural purposes. These findings are expected to shape the future of genome editing technology and provide new directions for genetic engineering researchers.
Journal reference
Čepaitė R., Klein N., Mikšys A. et al. “Structural variation in type IV-A1- and type IV-A3-mediated CRISPR interference.” Nature Communications (2024). DOI: https://doi.org/10.1038/s41467-024-53778-1
About the author
Professor Patrick Bausch He is an outstanding researcher in the field of genome editing technology and leads breakthrough research on the CRISPR system. Working at Vilnius University, he specializes in decoding the molecular mechanisms of gene regulation with the aim of developing advanced genetic engineering tools.
Dr. Lena Malinasket is a molecular biologist whose work focuses on understanding DNA-protein interactions. Her research has made significant contributions to innovation in CRISPR systems, with a focus on structural biology, to unlock their potential in medical and agricultural applications.
Dr. Rafael Pinilla-Redondo is a distinguished microbiologist who specializes in the study of bacterial immune systems and their applications in biotechnology. He is affiliated with the University of Copenhagen and is dedicated to exploring the diversity and evolution of CRISPR systems to address pressing scientific challenges.
Professor Lennart Landau is a molecular scientist known for his work on RNA biology and microbial defense systems. His work at Philipps University of Marburg has greatly improved our understanding of CRISPR adaptive mechanisms in microorganisms and is of great significance for future biotechnology innovation.
