Abstract
Abstract
Aptamers are artificial oligonucleotides binding to specific molecular targets. They have a promising role in therapeutics and diagnostics but are often difficult to design. Here, we exploited the catRAPID algorithm to generate aptamers targeting TAR DNA-binding protein 43 (TDP-43), whose aggregation is associated with Amyotrophic Lateral Sclerosis. On the pathway to forming insoluble inclusions, TDP-43 adopts a heterogeneous population of assemblies, many smaller than the diffraction-limit of light. We demonstrated that our aptamers bind TDP-43 and used the tightest interactor, Apt-1, as a probe to visualize TDP-43 condensates with super-resolution microscopy. At a resolution of 10 nanometers, we tracked TDP-43 oligomers undetectable by standard approaches. In cells, Apt-1 interacts with both diffuse and condensed forms of TDP-43, indicating that Apt-1 can be exploited to follow TDP-43 phase transition. The de novo generation of aptamers and their use for microscopy opens a new page to study protein condensation.
Aptamers are artificial oligonucleotides binding to specific molecular targets. They have a promising role in therapeutics and diagnostics but are often difficult to design. Here, we exploited the catRAPID algorithm to generate aptamers targeting TAR DNA-binding protein 43 (TDP-43), whose aggregation is associated with Amyotrophic Lateral Sclerosis. On the pathway to forming insoluble inclusions, TDP-43 adopts a heterogeneous population of assemblies, many smaller than the diffraction-limit of light. We demonstrated that our aptamers bind TDP-43 and used the tightest interactor, Apt-1, as a probe to visualize TDP-43 condensates with super-resolution microscopy. At a resolution of 10 nanometers, we tracked TDP-43 oligomers undetectable by standard approaches. In cells, Apt-1 interacts with both diffuse and condensed forms of TDP-43, indicating that Apt-1 can be exploited to follow TDP-43 phase transition. The de novo generation of aptamers and their use for microscopy opens a new page to study protein condensation.
| Original language | English |
|---|---|
| Article number | 3306 |
| Number of pages | 13 |
| Journal | Nature Communications |
| Volume | 13 |
| DOIs | |
| Publication status | Published - 23 Jun 2022 |
Bibliographical note
We thank the “RNA Initiative” at IIT, all members of the M.H.H., A.P., and G.G.T. groups, and especially Fernando Cid Samper. M.H.H. wishes to thank UCB Biopharma and Dr Jim Love for providing funding for the instrument used to generate the super-resolution data in this manuscript. A.P. acknowledges funding from UK DRI (grant REI 3556) and AlzheimerUK (grant ARUK-PG2019B-020). O.K. was supported by a Scottish PhD Research & Innovation Network Traineeships in MND/MS. E.Z. received funding from the Newton fellowship scheme and the MINDED fellowship of the European Union’s Horizon 2020 research and innovation program under the Marie Skłodowska-Curie grant agreement No. 754490. K.J. was funded via the BBSRCEastBIO doctoral training program (BB/M010996/1). The research leading to these results was supported by European Research Council [RIBOMYLOME n. 309545 to G.G.T. and ASTRA n. 855923 to G.G.T.], H2020 [IASIS n. 727658 to G.G.T. and INFORE n. 825080 to G.G.T.] and MND [840-791 to G.G.T. and A.P.] projects. The authors would also like to acknowledge the help and support received during confocal images acquisition by the group of Giuseppe Vicidomini at the Molecular Microscopy and Spectroscopy Department of IIT.Data Availability Statement
Data availabilityThe super-resolution and SAVE image data generated in this study have been deposited in the Zenodo database under accession code 6533779. Not all the calculations for aptamers are publicly available due to the filing of a patent (Italian priority application N. 102022000009500) by E.Z. (IIT), A.A. (IIT and CRG), G.G.T. (IIT, CRG, and ICREA), O.K. (University of Edinburgh), M.H.H. (University of Edinburgh), and A.P. (and King’s College London) but they can be granted upon reasonable request under non-disclosure agreement. Source data are provided with this paper.
Code availability
Predictions for the aptamer sequences were carried out by means of the software “catRAPID omics”, version 2.0, available at http://s.tartaglialab.com/page/catrapid_omics2_group (the RNA Fitness of Apt-1 is at https://tinyurl.com/y3578hr3 and the Protein Fitness of Apt-1 is at https://tinyurl.com/evmybn73). Calculations for sequences <50 nucleic acids are restricted in the webserver due to the filing of a patent (see also Data availability) but they can be provided upon reasonable request under non-disclosure agreement.
Keywords
- computational models
- RNA
- RNA-Binding Proteins
- Super-resolution microscopy