RNA FISH for detecting expanded repeats in human diseases.

Area of Science:

• Molecular Biology
• Genetics
• Neuroscience

Background:

• RNA fluorescence in situ hybridization (FISH) is crucial for detecting transcripts in cells and tissues.
• Variants of RNA FISH enhance signal, resolution, and specificity for subcellular transcript localization.
• RNA FISH is vital for studying nuclear RNA foci in repeat expansion diseases.

Purpose of the Study:

• To summarize findings from FISH studies examining RNA nuclear inclusions.
• To provide a detailed protocol for detecting expanded CAG and CUG repeat RNAs in various cellular models.
• To present the initial single-molecule FISH application in a polyglutamine disease model.

Main Methods:

• Utilizing RNA fluorescence in situ hybridization (FISH) to detect transcripts.
• Applying FISH to various cellular models: fibroblasts, lymphoblasts, iPSCs, and neuronal progenitors.
• Implementing single-molecule FISH for high-resolution analysis.

Main Results:

• FISH successfully visualizes nuclear RNA foci in repeat expansion disease models.
• The protocol facilitates detection of expanded CAG and CUG repeat RNAs.
• Single-molecule FISH provides novel insights into polyglutamine disease mechanisms.

Conclusions:

• RNA FISH is a powerful tool for investigating RNA nuclear inclusions in repeat expansion disorders.
• The detailed protocol enables broader application of FISH in disease research.
• Single-molecule FISH opens new avenues for understanding disease pathogenesis at the molecular level.