Phosphorylation of EPYC1 by KEY1 regulates pyrenoid condensate dynamics

Abstract

Biomolecular condensates are membrane-less compartments that undergo dynamic changes in size, number, and composition. Despite the importance of these changes for proper cell function, how these dynamics are regulated remains largely unexplored. One tractable model system to study condensate dynamics is the pyrenoid, a singular condensate in the alga Chlamydomonas reinhardtii that enhances global carbon-fixation. Composed of the CO2-fixing enzyme Rubisco and linker protein EPYC1, the pyrenoid dynamically reorganizes during cell division. However, the mechanisms underlying these dynamic phase behaviors remain elusive. Preliminary findings suggest that a kinase KEY1 regulates pyrenoid dynamics. KEY1 directly phosphorylates EPYC1, and loss of KEY1 function results in the formation of multiple pyrenoid condensates. In this thesis, key1 mutants and EPYC1 phospho-mutants were imaged to elucidate the mechanistic role of KEY1 in regulating pyrenoid dynamics. Confocal images of EPYC1 phospho-mutants reveal that KEY1 phosphorylation of EPYC1 is essential for controlling pyrenoid count. Time lapse imaging of synchronized key1 mutant cells indicate that KEY1 enables symmetric pyrenoid inheritance during cell division and may facilitate pyrenoid growth in the G1 phase. We propose a mechanistic model for how KEY1 phosphorylation of EPYC1 regulates pyrenoid inheritance and growth during the cell cycle. Given that aberrant condensate dynamics are often associated with disease, these results provide an important framework to understand how kinases may regulate the dynamics of other condensates. Furthermore, this work provides additional insights to pyrenoid biogenesis, advancing efforts to engineer synthetic pyrenoids into crops to increase yields and mitigate climate change.