Experimental Imaging Center
Imaging of gene regulation
Despite being first introduced in biomedical research over a century ago, fluorescence microscopy is ageing extremely well. Thanks to genetically encoded fluorescent reporters such as GFP (Nobel Prize in Chemistry, 2008), it is possible to localize and follow biological events in real time in living cells. Further, the interaction between light and fluorescent molecules allows to circumvent physical barriers once thought unsurpassable and collect images of biological samples with details on the nanometer spatial scale, as demonstrated by the recent advances in super-resolution microscopy (Nobel Prize in Chemistry 2014). Fluorescence microscopy is also sensitive enough to detect and track an individual ("single") molecule as it moves and interacts with its partners, leading to quantitative analysis on a "molecule by molecule" base of protein dynamics, protein-protein interactions, gene expression and more.
Research activity
Our team focuses on developing and applying single molecule approaches to quantifying dynamic events in individual living cells. In particular, we have pioneered a single- molecule approach to quantify the interaction kinetics between nuclear proteins (transcription factors) and its binding sites on DNA, and we are currently applying it to dissect the causes that underlie activation and inactivation of those transcription factors that act as tumor suppressors or oncogenes in cancer settings.
Together with these activities, we support members of our scientific community interested in developing/applying assays in advanced fluorescence microscopy, ranging from protocol development, to data acquisition and data analysis for F-techniques ultrasensitive, single molecule and single-particle imaging and super-resolution microscopy.
Mazzocca, M., Loffreda, A., Colombo, E., Fillot, T.,Gnani, D., Falletta, P., Monteleone, E., Capozi, S., Bertrand, E., Legube, G., Lavagnino, Z., Tacchetti, C. & Mazza, D. (2023). Chromatin organization drives the search mechanism of nuclear factors. Nature Communications, 14(1), 6433.
Louphrasitthiphol, P., Loffreda, A. , Pogenberg V., Picaud, S., Schepsky, A., Friedrichsen, H., Zeng, Z. Lashgari, A., Thomas, B., Patton, E.E.. Wilmanns, M., Filippakopoulos, P., Lambert, J., Steingrímsson, E., Mazza, D., & Goding, C.R. (2023) Acetylation reprograms MITF target selectivity. Nature Communications, 14(1), 6051.
Biasin, M., Strizzi, S., Bianco, A., Macchi, A., Utyro, O., Pareschi, G., Loffreda, A., Cavalleri, A., Lualdi, M., Trabattoni, D., Tacchetti, C., Mazza, D., & Clerici, M. (2022). UV and violet light can Neutralize SARS-CoV-2 Infectivity. Journal of Photochemistry and Photobiology, 10, 100107.
Mazzocca, M., Colombo, E., Callegari, A., & Mazza, D. (2021). Transcription factor binding kinetics and transcriptional bursting: What do we really know? Current Opinion in Structural Biology, 71, 239–248.
Mazzocca, M., Fillot, T., Loffreda, A., Gnani, D., & Mazza, D. (2021). The needle and the haystack: single molecule tracking to probe the transcription factor search in eukaryotes. Biochemical Society Transactions, 49(3), 1121–1132.
Louphrasitthiphol, P., Siddaway, R., Loffreda, A., Pogenberg, V., Friedrichsen, H., Schepsky, A., Zeng, Z., Lu, M., Strub, T., Freter, R., Lisle, R., Suer, E., Thomas, B., Schuster-Böckler, B., Filippakopoulos, P., Middleton, M., Lu, X., Patton, E. E., Davidson, I., …, Mazza, D.*, Goding, C. R.* (2020). Tuning Transcription Factor Availability through Acetylation-Mediated Genomic Redistribution. Molecular Cell, 79(3), 472-487.e10.
Zambrano, S., Loffreda, A., Carelli, E., Stefanelli, G., Colombo, F., Bertrand, E., Tacchetti, C., Agresti, A., Bianchi, M. E., Molina, N., & Mazza, D. (2020). First Responders Shape a Prompt and Sharp NF-κB-Mediated Transcriptional Response to TNF-α. IScience, 23(9), 101529.
Callegari, A., Sieben, C., Benke, A., Suter, D. M., Fierz, B., Mazza, D., & Manley, S. (2019). Single-molecule dynamics and genome-wide transcriptomics reveal that NF-kB (p65)-DNA binding times can be decoupled from transcriptional activation. PLoS Genetics, 15(1), e1007891.
Cammarota, E., & Mazza, D. (2019). Monitoring Transcription Factor Oligomerization in Single Living Cells by Number and Brightness Analysis. Methods in Molecular Biology (Clifton, N.J.), 2038, 223–237.
Loffreda, A., Jacchetti, E., Antunes, S., Rainone, P., Daniele, T., Morisaki, T., Bianchi, M. E., Tacchetti, C., & Mazza, D. (2017). Live-cell p53 single-molecule binding is modulated by C-terminal acetylation and correlates with transcriptional activity. Nature Communications, 8(1), 313.
Swinstead, E. E., Miranda, T. B., Paakinaho, V., Baek, S., Goldstein, I., Hawkins, M., Karpova, T. S., Ball, D., Mazza, D., Lavis, L. D., Grimm, J. B., Morisaki, T., Grøntved, L., Presman, D. M., & Hager, G. L. (2016). Steroid Receptors Reprogram FoxA1 Occupancy through Dynamic Chromatin Transitions. Cell, 165(3), 593–605.
Caldieri, G., Barbieri, E., Nappo, G., Raimondi, A., Bonora, M., Conte, A., Verhoef, L. G. G. C., Confalonieri, S., Malabarba, M. G., Bianchi, F., Cuomo, A., Bonaldi, T., Martini, E., Mazza, D., Pinton, P., Tacchetti, C., Polo, S., Di Fiore, P. P., & Sigismund, S. (2017). Reticulon 3-dependent ER-PM contact sites control EGFR nonclathrin endocytosis. Science (New York, N.Y.), 356(6338), 617–624.
Morisaki, T., Müller, W. G., Golob, N., Mazza, D.*, & McNally, J. G.* (2014). Single-molecule analysis of transcription factor binding at transcription sites in live cells. Nature Communications, 5, 4456.
Mazza, D., Mueller, F., Stasevich, T. J., & McNally, J. G. (2013). Convergence of chromatin binding estimates in live cells. Nature Methods, 10(8), 691–692.
Mazza, D., Abernathy, A., Golob, N., Morisaki, T., & McNally, J. G. (2012). A benchmark for chromatin binding measurements in live cells. Nucleic Acids Research, 40(15), e119.
