Modeling Asynchronous Nuclear Divisions in Growing Fungal Networks

We aimed to understand how nuclei within the multinucleate fungus Ashbya gossypii maintain asynchronous cell cycles despite sharing a common cytosol, while the mycelium as a whole simultaneously regulates its nuclear-to-cytoplasmic ratio. This is inherently a multiscale problem, with nanometer scale cell cycle regulators diffusing between nuclei spaced micrometers apart, all within a continuous fungal cell spanning millimeters. Asynchronous nuclear divisions in Ashbya suggest that regulatory molecules do not diffuse uniformly, so we first investigated the physical properties of the cytosol. Using genetically encoded multimeric nanoparticles (GEMs) as passive probes, we estimated local diffusivity throughout individual fungal hyphae and related it to intracellular structures. GEM diffusivity varied by orders of magnitude within single cells, with reduced mobility near nuclei, suggesting that cytosolic organization can slow molecular movement around nuclei to potentially insulate them and enable asynchronous cell cycles. We next turned to the scale of the entire mycelium, combining mathematical modeling with experimental biology to determine how the nuclear-to-cytoplasmic ratio is controlled while nuclei simultaneously remain asynchronous from one another. Modeling dividing nuclei as a growing system of phase oscillators in a branching fungal cell revealed that asynchronous cell cycles are required to prevent fluctuations in nuclear density that would arise from simultaneous divisions, and that a cell cycle checkpoint alone leads to synchrony rather than stabilizing density. Instead, if branching and nuclear density are coupled, the cell can both stabilize the nuclear-to-cytoplasmic ratio and avoid synchronous cell cycles. Finally, we addressed how nuclei maintain asynchrony while sharing a common cytosol by testing a model in which nuclei are asymmetrically coupled through production of the G1 cyclin Cln3. In a fixed population of oscillators, this Cln3-mediated coupling strongly generates asynchrony, but introducing nuclear division continually increases local synchrony, requiring stronger coupling for asynchrony to begin to emerge. Incorporating nuclear mobility disperses daughters and brings local asynchrony close to the experimental value, suggesting that Cln3-mediated asymmetric coupling, combined with nuclear mobility, can generate cell cycle asynchrony between nuclei coupled through a common cytosol.