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- Title
From Cell Populations to Molecular Complexes: Multiplexed Multimodal Microscopy to Explore p53-53BP1 Molecular Interaction.
- Authors
Pelicci, Simone; Furia, Laura; Pelicci, Pier Giuseppe; Faretta, Mario
- Abstract
Surpassing the diffraction barrier revolutionized modern fluorescence microscopy. However, intrinsic limitations in statistical sampling, the number of simultaneously analyzable channels, hardware requirements, and sample preparation procedures still represent an obstacle to its widespread diffusion in applicative biomedical research. Here, we present a novel pipeline based on automated multimodal microscopy and super-resolution techniques employing easily available materials and instruments and completed with open-source image-analysis software developed in our laboratory. The results show the potential impact of single-molecule localization microscopy (SMLM) on the study of biomolecules' interactions and the localization of macromolecular complexes. As a demonstrative application, we explored the basis of p53-53BP1 interactions, showing the formation of a putative macromolecular complex between the two proteins and the basal transcription machinery in situ, thus providing visual proof of the direct role of 53BP1 in sustaining p53 transactivation function. Moreover, high-content SMLM provided evidence of the presence of a 53BP1 complex on the cell cytoskeleton and in the mitochondrial space, thus suggesting the existence of novel alternative 53BP1 functions to support p53 activity.
- Subjects
CELL populations; MOLECULAR interactions; P53 antioncogene; MICROSCOPY; FLUORESCENCE microscopy; DNA repair
- Publication
International Journal of Molecular Sciences, 2024, Vol 25, Issue 9, p4672
- ISSN
1661-6596
- Publication type
Article
- DOI
10.3390/ijms25094672