Physical principles and functional consequences of nuclear compartmentalization in budding yeast

Abstract

One striking feature of eukaryotic nuclei is the existence of discrete regions in which specific factors concentrate while others are excluded, thus forming microenvironments with different molecular compositions and biological functions. These domains are often referred to as sub-compartments even though they are not membrane enclosed. Despite their functional importance the physical nature of these structures remains largely unknown. Here we describe how the Saccharomyces cerevisiae nucleus is compartmentalized and discuss possible physical models underlying the formation and maintenance of chromatin associated sub-compartments. Focusing on three particular examples: the nucleolus, silencing foci and repair foci, we discuss the biological implications of these different models as well as possible approaches to challenge them in living cells.

Single molecule microscopy reveals key physical features of repair foci in living cells

“…Thus, our results indicate that upon 2 DSBs, Rad52 foci do not merely cluster, but form a focus of size consistent with the fusion of 2 foci inside which molecules explore the entire larger sub-compartment. Finally, in LLPS, increasing protein amounts should increase the focus size while concentrations remain the same inside foci and in the nucleoplasm 2,22 . In the case a LLPS is formed but the molecule observed is not the main species driving the LLPS, the concentration inside foci and in the background increases linearly with over-expression of the observed molecule.…”

“…These interactions can be driven by interactions between chromatin binding proteins or chromatin components. In this model, referred to as "Bridging" Model, or Polymer Polymer Phase Separation (PPPS) model 2,22 , the existence of sub-compartments relies on both the binding and bridging properties of these proteins to chromatin. A third scenario is the Liquid-Liquid Phase Separation (LLPS) Model, also referred to as "Droplet Model" 23 .…”

“…Despite a large literature in the field, many in vivo tests commonly used to probe the nature of sub-compartments are extremely phenomenological and insufficient to rule out other possible mechanisms 2,24 . In addition, most of the optical methods used to discriminate models are at the limit of the diffraction and suffer from severe artefacts 2,24 . Overall, there is a lack of solid experimental criteria to confirm one model versus another in living cells, in particular at the microscopic level.…”

“…The cell nucleus contains membrane-less sub-compartments inside which specific proteins are more concentrated than elsewhere in the nucleus [1][2][3] . Such regions of high local protein concentration, called "foci", are hypothesized to help proteins coordinate and collectively perform their function 2,[4][5][6][7] . The formation of these foci at the right place in the nucleus and within a well-defined time window is essential for the functioning of the cell.…”

“…How repair foci and more generally membrane-less sub-compartments are formed, maintained and disassembled at time scales relevant for their biological function remains unknown. Several models are intensively debated in the literature to understand the nature of membrane-less sub-compartments 2,[21][22][23][24][25][26] .…”