Pseudouridine (Ψ) is a common modification found in RNA, the molecule that plays a vital role in coding, decoding, regulation, and expression of genes. Think of RNA as a messenger that carries instructions from DNA to make proteins. The addition of pseudouridine to RNA molecules can change how they function, but until recently, scientists struggled to fully understand its role due to challenges in detecting it accurately.
What is Pseudouridine?
Pseudouridine is formed when a uridine nucleotide (one of the building blocks of RNA) undergoes a chemical change. This modification can help stabilize RNA structures, influence how RNA is read by other molecules, and potentially affect gene expression. Even though it’s one of the most abundant modifications found in RNA, the specific functions of pseudouridine have been difficult to pinpoint because traditional methods for detecting it have limitations.
A New Method for Detection: BACS
Researchers at the University of Oxford have developed a new technique called 2-bromoacrylamide-assisted cyclization sequencing (BACS) to improve the detection and study of pseudouridine. This innovative method allows scientists to observe where pseudouridine is located in RNA sequences with high precision, even in areas where it is densely packed.
Here’s how BACS works:
It helps convert pseudouridine back into regular cytidine (C) in RNA sequences, making it easier to identify its presence during sequencing.
By analyzing the modified RNA, scientists can create a detailed map showing exactly where pseudouridine is located.
BACS detected known Ψ sites in human rRNA and spliceosomal snRNAs
a, Flowchart of BACS library construction. b, Numbers of Ψ sites identified in HeLa cy-rRNAs and mt-rRNAs. c–e, Conversion rates of BACS (pink) and control (gray) samples in HeLa 28S rRNA (c), 18S rRNA (d) and 5.8S rRNA (e), respectively. Data are presented as means of two independent experiments. f, Venn diagram illustrating the overlap of Ψ sites detected in human cy-rRNAs between BACS and SILNAS MS. g, Numbers of Ψ sites identified in HeLa spliceosomal snRNAs. h, Venn diagram illustrating the overlap of Ψ sites detected in human spliceosomal snRNAs between BACS and SILNAS MS. i, Conversion rates of BACS (pink) and control (gray) samples in HeLa U2 snRNA. Data are presented as means of two independent experiments.
Key Findings from the Study
Using BACS, the researchers were able to detect all known pseudouridine sites in human ribosomal RNA (rRNA), which is crucial for protein synthesis, as well as in small nuclear RNAs involved in splicing (the process of editing RNA before it is translated into proteins). They also created a quantitative map of pseudouridine in other types of RNA, including small nucleolar RNA (which helps modify rRNA) and transfer RNA (tRNA, which brings amino acids to the ribosome during protein synthesis).
In addition to detecting pseudouridine, BACS can also identify other important RNA modifications, such as adenosine-to-inosine editing and N1-methyladenosine, further expanding its utility in RNA research.
Insights into Viruses and Pseudouridine
Interestingly, the researchers discovered a specific pseudouridine site (Ψ114) in a small RNA produced by the Epstein–Barr virus (a virus that can cause various diseases). However, when they tested a variety of RNA viruses, they found that these viruses typically do not contain pseudouridine in their RNA genomes. This finding highlights important differences in how various virus families modify their RNA.
Conclusion
The introduction of the BACS method represents a significant advancement in RNA research. By providing a more precise way to detect pseudouridine and other modifications, this technique could lead to a better understanding of how these changes affect RNA function in health and disease. As researchers continue to explore the roles of RNA modifications like pseudouridine, we may uncover new insights that could impact fields ranging from genetics to virology. This work is paving the way for further discoveries in RNA biology and could eventually contribute to the development of novel therapeutic strategies.