Biophysical principles governing spatiotemporal regulation of compartmentalization in cells [HBNI Th292] 

Abstract

Compartmentalization is a fundamental process underlying the spatiotemporal organization of biological systems. Compartments provide several advantages, such as creating distinct microenvironments for biochemical reactions, enabling the localization of biomolecules, and allowing multiple processes to be carried out simultaneously within the cell. Cells achieve compartmentalization in two ways: (i) via membraneless organelles, or biomolecular condensates, and (ii) via membrane-bound organelles. In this thesis, we aim to understand the physical principles governing compartmentalization in cells for both membraneless and membrane-bound organelles.

Proteins can form membraneless compartments called biomolecular condensates, which facilitate various cellular processes, such as stress responses and transcriptional regulation. These proteins also interact with intracellular surfaces, such as membranes and cytoskeletal filaments; however, the physical principles governing these interactions remain poorly characterized. In this thesis, we study how tau, a microtubule-associated protein, interacts with the microtubule surface. Tau, which is implicated in neurodegenerative disorders, forms condensates in solution at micromolar concentrations. However, recent experiments show that, at much lower concentrations in the nanomolar range, tau forms dense, cohesive patches on microtubules. These patches, previously referred to as envelopes, are distributed non-uniformly along microtubules and form in a concentration-dependent manner. To shed light on these observations, we employ a lattice-gas framework incorporating tau–tau and tau–microtubule interactions. We show that tau envelopes are multilayered condensates formed via a prewetting-like transition. The spatial patterns of tau condensates are dictated by inhomogeneities on the microtubule surface, likely arising from post-translational modifications. We demonstrate that the multilayered nature of the condensates can give rise to kinetically distinct populations of tau with different functional capabilities. In particular, the innermost layer can restrict the access of other proteins, such as severing enzymes, to the microtubule, while the outer layers can recruit tau-interacting molecules, such as soluble tubulin. Our results thus show that tau forms surface-induced condensates on microtubules at physiological concentrations, thereby providing additional regulatory control over microtubule-based functional pathways. Our theoretical framework can be generalized to study phase-separating proteins interacting with other surfaces, such as DNA and membranes.

Compartmentalization in cells is also achieved via various membrane-bound organelles, such as nuclei, mitochondria, and lysosomes, which carry out specialized functions. The abundance of these organelles in cells is tightly regulated in response to external stimuli, but the underlying mechanisms remain elusive. Time-lapse imaging of fluorescently labeled organelles enables single-cell measurements of organelle copy numbers, revealing the time evolution of their distribution across a cell population. In this thesis, we aim to provide a theoretical framework that can leverage these measurements to discern the underlying regulatory mechanisms. Building on a recently proposed kinetic model of organelle biogenesis, which incorporates de novo synthesis, fission, fusion, and degradation, we explore the time-dependent dynamics of organelle abundance. While previous studies have focused on steady-state properties, here we calculate the first two moments of: (i) organelle copy numbers over time, and (ii) first-passage times to reach a specified organelle count. We show that these two moments provide a powerful means of discriminating between different mechanisms of organelle biogenesis. Notably, the time-dependent behavior of organelle biogenesis reveals richer dynamics compared to the steady-state scenario. Our findings shed light on how cells attain steady-state organelle abundance after cell division and environmental perturbation, which can be used to elucidate the underlying mechanisms regulating organelle abundance.

To summarize, in this thesis, we explored two distinct physical routes through which intracellular organization via compartmentalization can emerge. First, we showed that surface-induced condensation of tau can give rise to a functionally diverse microenvironment on microtubules, even below the bulk saturation concentration. Second, we investigated the regulation of membrane-bound organelles, providing a theoretical framework to elucidate mechanisms of organelle abundance control using time-lapse measurements. Together, these studies provide a physical basis for understanding the formation and regulation of diverse intracellular compartments.