Phosphoregulation of hierarchical protein condensate formation during clathrin-mediated endocytosis
- 1University of California Berkeley
Abstract
The sequential assembly of over 50 proteins during clathrin-mediated endocytosis (CME) is well documented in yeast. However, the mechanisms governing this ordered recruitment remain poorly understood. Biomolecular condensation is proposed to orchestrate CME, but defining biophysical parameters in vivo remains challenging. Here, we developed a quantitative in vivo titration strategy to systematically determine CME protein assembly profiles. Beyond the previously reported Ede1 (yeast Eps15), seven additional proteins formed enlarged, concentration-dependent assemblies with distinct biophysical properties. Single-cell profiling revealed two biophysical classes among these eight proteins: homeostatic buffers with defined saturation boundaries and continuous assemblers that scale linearly. Acting as scaffolds and clients, these conserved proteins form an interconnected network of kinases, phosphatase recruiters, and substrates. Within this network, reversible phosphorylation modulates assembly stability to drive recruitment transitions. Collectively, this work establishes a two-stage model of protein condensation that drives CME progression from initiation to maturation and offers a general framework for how protein phosphorylation dynamically modulates biomolecular condensation in vivo.
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BibTeX
@misc{sun2026phosphoregulation,
title = {{Phosphoregulation of hierarchical protein condensate formation during clathrin-mediated endocytosis}},
author = {Y. Sun and A. Wong and G. Hu and A. Yeam and K. Wong and L. V. Dao and A. Tschanz and D. G. Drubin},
howpublished = {bioRxiv},
year = {2026},
month = sep,
doi = {10.64898/2026.09.17.752504},
url = {https://www.biorxiv.org/content/10.64898/2026.09.17.752504v1},
note = {CC BY-NC-ND},
}
RIS
TY - UNPB
TI - Phosphoregulation of hierarchical protein condensate formation during clathrin-mediated endocytosis
AU - Y. Sun
AU - A. Wong
AU - G. Hu
AU - A. Yeam
AU - K. Wong
AU - L. V. Dao
AU - A. Tschanz
AU - D. G. Drubin
PY - 2026
DA - 2026/09/17
PB - bioRxiv
DO - 10.64898/2026.09.17.752504
UR - https://www.biorxiv.org/content/10.64898/2026.09.17.752504v1
AB - The sequential assembly of over 50 proteins during clathrin-mediated endocytosis (CME) is well documented in yeast. However, the mechanisms governing this ordered recruitment remain poorly understood. Biomolecular condensation is proposed to orchestrate CME, but defining biophysical parameters in vivo remains challenging. Here, we developed a quantitative in vivo titration strategy to systematically determine CME protein assembly profiles. Beyond the previously reported Ede1 (yeast Eps15), seven additional proteins formed enlarged, concentration-dependent assemblies with distinct biophysical properties. Single-cell profiling revealed two biophysical classes among these eight proteins: homeostatic buffers with defined saturation boundaries and continuous assemblers that scale linearly. Acting as scaffolds and clients, these conserved proteins form an interconnected network of kinases, phosphatase recruiters, and substrates. Within this network, reversible phosphorylation modulates assembly stability to drive recruitment transitions. Collectively, this work establishes a two-stage model of protein condensation that drives CME progression from initiation to maturation and offers a general framework for how protein phosphorylation dynamically modulates biomolecular condensation in vivo.
ER -