posted on 2023-11-02, 13:00authored byZoe Gidden, Curran Oi, Emily J. Johnston, Zuzanna Konieczna, Haresh Bhaskar, Lorena Mendive-Tapia, Fabio de Moliner, Susan J. Rosser, Simon G. J. Mochrie, Marc Vendrell, Mathew H. Horrocks, Lynne Regan
Fluorescence microscopy
enables specific visualization of proteins
in living cells and has played an important role in our understanding
of the protein subcellular location and function. Some proteins, however,
show altered localization or function when labeled using direct fusions
to fluorescent proteins, making them difficult to study in live cells.
Additionally, the resolution of fluorescence microscopy is limited
to ∼200 nm, which is 2 orders of magnitude larger than the
size of most proteins. To circumvent these challenges, we previously
developed LIVE-PAINT, a live-cell super-resolution approach that takes
advantage of short interacting peptides to transiently bind a fluorescent
protein to the protein-of-interest. Here, we successfully use LIVE-PAINT
to image yeast membrane proteins that do not tolerate the direct fusion
of a fluorescent protein by using peptide tags as short as 5-residues.
We also demonstrate that it is possible to resolve multiple proteins
at the nanoscale concurrently using orthogonal peptide interaction
pairs.