Physical Chemistry Seminar: Confocal microscopy approaches in studying biomolecular condensates in cells

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Abstract:

Biomolecular condensates (biocondensates, in short) facilitate self-organization of biomolecules (e.g., proteins, nucleic acids, lipids, metabolites) through non-specific multivalent weak intermolecular interactions to form dynamic membraneless sub-compartments that carry multiple biological functions. Systematic in vitro reconstructions have been suggesting that a few predominant biomolecules are sufficient to form biocondensates through different phase separation mechanisms. However, biocondensates in cellulo are more complex and harder to be studied systematically.

In this talk I will strive to focus on two naturally-occurring biocondensates we have been studied recently: (1) heterochromatin condensates in chromocenters of pluripotent embryonic stem cells (ESCs) before and during cell differentiation (published), and (2) α-synuclein (α-syn) bodies in the SH-SY5Y model neuronal cells (unpublished). I will cover several exciting and (sometimes) novel experimental approaches using laser-scanning confocal microscopy, including local density sensing using fluorescence lifetimes of fluorescent proteins, homoFRET through fluorescence anisotropy of fluorescent proteins, probing rare diffusing biomolecular clusters, one cluster at a time, using BLISS, 1- and 2-color immunofluorescence stimulated emission depletion, STED, microscopy) backed by mass spectrometric proteomic analyses. Using these approaches, I will show (1) the process heterochromatin undergoes during the first 10 days differentiating away from pluripotent state and into mature cells facilitating its transition between two distinctly stabilized states, and (2) that α-syn supports previously-unknown physiological functions within biocondensates that can explain what cells use excess α-syn for, aside from getting rid of it.

Seminar Organizer: Prof. Yuval Ebenstein