PT  - JOURNAL ARTICLE
AU  - Alexander H. Clowsley
AU  - William T. Kaufhold
AU  - Tobias Lutz
AU  - Anna Meletiou
AU  - Lorenzo Di Michele
AU  - Christian Soeller
TI  - Detecting nanoscale distribution of protein pairs by proximity dependent super-resolution microscopy
AID  - 10.1101/591081
DP  - 2020 Jan 01
TA  - bioRxiv
PG  - 591081
4099  - http://biorxiv.org/content/early/2020/05/30/591081.short
4100  - http://biorxiv.org/content/early/2020/05/30/591081.full
AB  - Interactions between biomolecules such as proteins underlie most cellular processes. It is crucial to visualize these molecular-interaction complexes directly within the cell, to show precisely where these interactions occur and thus improve our understanding of cellular regulation. Currently available proximity-sensitive assays for in-situ imaging of such interactions produce diffraction-limited signals and therefore preclude information on the nanometer-scale distribution of interaction complexes. By contrast, optical super-resolution imaging provides information about molecular distributions with nanometer resolution which has greatly advanced our understanding of cell biology. However, current co-localization analysis of super-resolution fluorescence imaging is prone to false positive signals as the detection of protein proximity is directly dependent on the local optical resolution. Here we present Proximity-Dependent PAINT (PD-PAINT), a method for sub-diffraction imaging of protein pairs, in which proximity detection is decoupled from optical resolution. Proximity is detected via the highly distance-dependent interaction of two DNA labels anchored to the target species. Labeled protein pairs are then imaged with high contrast and nanoscale resolution using the super-resolution approach of DNA-PAINT. The mechanisms underlying the new technique are analyzed by means of coarse-grained molecular simulations and experimentally demonstrated by imaging DNA-origami tiles and epitopes of cardiac proteins in isolated cardiomyocytes. We show that PD-PAINT can be straightforwardly integrated in a multiplexed super-resolution imaging protocol and benefits from advantages of DNA-based super-resolution localization microscopy, such as high specificity, high resolution and the ability to image quantitatively.Competing Interest StatementThe authors have declared no competing interest.