A Novel Reporter for Quantitative Imaging of RNA Aberrant Splicing

RNA splicing is an important step in the expression of most eukaryote genes. It takes place in the nucleus during or immediately after transcription and involves removing the introns and joining exons together. Typically, precursor messengers (pre-mRNA) splicing follows the GU-AG rule. RNA splicing mechanism generally requires trans-acting factors and cis-acting elements to facilitate the splicing of the spliceosome- a large ribonucleic acid-protein complex comprising of proteins and small nuclear RNAs. In some cases, genes may undergo aberrant splicing due to the abnormal variation in the splicing process. Aberrant splicing affects different biological processes and can cause diseases. Unfortunately, most of the techniques currently used for imaging and detection of aberrant splicing in living subjects are unable to provide the desired dynamic information of splicing events due to several disadvantages such as low specificity of localization.

The use of reporter genes for the quantification of mRNA molecules has exhibited promising results despite several drawbacks reported in recent studies. In an effort to address these challenges, a team of researchers from Xidian University: Jinrong Xie, Haifeng Zheng, Si Chen, Xiaorui Shi, Wenjie Mao, and led by Professor Fu Wang developed a new activatable bioluminescent reporter for real-time imaging of pre-mRNA aberrant splicing process. Their work is currently published in the research journal, Molecular Therapy – Methods and Clinical Development.

In their approach, the cell-based reporter, also known as the Rluc-SMN system, was designed for continuous and noninvasive imaging of pre-mRNA splicing patterns based on the principle that exogenous splicing inhibitors in living organisms can easily detectable positive signals for detecting aberrant splicing. Specifically, the design involved inserting a human survival motor neuron upstream of the renilla luciferase (Rluc) such that only the active luciferase protein is translated by the unspliced mRNA. The feasibility of the Rluc-SMN reporter was validated against its ability to provide real-time and quantitative dynamic information about the splicing events.

The research team reported that the genetically encoded bioluminescence reporter exhibited the ability to visualize and measure real-time pre-mRNA aberrant splicing patterns in a dose- and time-dependent manner. Moreover, it could provide both quantitative and longitudinal information about the splicing process in response to the exogenous splicing inhibitors in living animals- as hypothesized by the researchers. Based on the data, the research team also noted that the activatable reporter could be potentially used for both noninvasive visualization of aberrant splicing process as well as high-throughput screening of splicing modulators in cells.

In summary, the study demonstrated the design of a new genetically encoded bioluminescence reporter for real-time and quantitative imaging of RNA aberrant splicing. Based on the results, the proposed method addressed most of the drawbacks reported in the previous studies and is thus a promising tool for noninvasive visualization of the splicing process and high-throughput screening of splicing modulators in cells. In a statement to Medicine Innovates, Professor Fu Wang said their findings would be useful in developing drugs for the treatment of diseases related to aberrant splicing of RNA.