Visualizing translation and co-translational folding of proteins live
Our current projects focus on tracking protein translation and co-translational folding of disease and aging-related targets with a high spatiotemporal resolution in living cells. We apply the state-of-art single-particle tracking approach to study the dynamics and uncover any heterogeneity in individual mRNAs.
How can we visualize the translation of proteins in living cells?
The translation of a single mRNA in living cells is first visualized in 2016. Our lab mainly applies a technology named Nascent Chain Tracking (NCT) to study protein translation. In our standard reporter, we insert 10 repeated epitope tags, such as the HA tag or FLAG tag, to the N-terminus of the protein of interest and 24 repeated MS2 stem loops in the 3' UTR. For imaging, we co-transfect the report, anti-HA or FLAG intrabodies-GFP (also named Frankenbodies), and MCP (MS2 Coat Protein) -Halo-JF549 into cells. When the reporter DNA is transcribed, the mRNA is lit up upon binding of the MCP-Halo-JF549 to the MS2 stem loops. When the mRNA is translated, the nascent chains are lit up upon binding of the intrabody-GFP to the epitope tags. The colocalized nascent chain spots in green and mRNA spots in magenta are translation spots. The spots can then be detected and tracked, providing insight into translation dynamics in living cells.
Visualizing Co-Translational Folding
Developing live-cell imaging tools that enable tracking protein translation and folding
To expand the toolbox of tracking translation and enable tracking protein folding in a living cell, our lab focuses on developing genetically-encodable intrabodies as live-cell imaging tools. We aim to develop a high throughput in vitro display technology for generating intrabodies.