Phospho-regulated tuning of viscoelastic properties balances centrosome growth and strength

Abstract

ABSTRACTCentrosomes nucleate microtubule asters and comprise centrioles surrounded by pericentriolar material (PCM). In preparation for mitosis, the PCM scaffold grows dramatically while resisting microtubule pulling forces. How PCM achieves dynamic growth while maintaining mechanical strength is unclear. Here we probed the dynamic and material properties of theC. elegansPCM scaffold. Single-embryo extrusion experiments revealed that the protein SPD-5 forms distinct but co-existing dynamic and stable scaffolds within PCM. The stable scaffold grew in preparation for mitosis, then disappeared at anaphase onset. SPD-5 mutants that lacked PLK-1 phosphorylation at four key sites (4A) could not build the stable scaffold, whereas phospho-mimetic SPD-5(4E) could. Expression of SPD-5(4A) impaired PCM assembly, but this phenotype was partially rescued by eliminating microtubule pulling forces, indicating material weakness. SPD-5(4A) expression also resulted in chromosome segregation defects, revealing the importance of PCM strength for development. Unexpectedly, expression of SPD-5(4E) prevented full-scale PCM growth and caused embryonic lethality.In vitro, PLK-1-induced phosphorylation increased the viscoelastic moduli of minimal SPD-5 scaffolds, which increased their solidity. This caused faster initial growth that then plateaued, in effect setting an upper limit to SPD-5 scaffold size. Thus, PCM scaffold assembly and strength are optimized through phospho-regulated equilibrium of dynamic and stable scaffold components. Our results further reveal kinase-driven kinetic arrest as a potential mechanism of centrosome size scaling.SIGNIFICANCE STATEMENTTo build and position the mitotic spindle, centrosomes must grow large enough to nucleate many hundreds of microtubules and resist their pulling forces. How centrosomes simultaneously achieve dynamic growth and tensile strength is unknown. Here, we show that centrosomes behave as phosphorylation-tunable hydrogels with both liquid-like and solid-like properties, which contribute to dynamics and material strength, respectively. Phospho-null or phospho-mimicking mutations in the scaffold protein SPD-5 dysregulate the balance of liquid- and sold-like behaviors within the centrosome, impairing function and embryo viability in both cases. Our results showcase how meso-scale material properties contribute to the function of a membrane-less organelle and how they are finely tuned by chemical modifications.