Integrating protein copy numbers with interaction networks to quantify stoichiometry in clathrin-mediated endocytosis

Abstract

Proteins that drive processes like clathrin-mediated endocytosis (CME) are expressed at copy numbers within a cell and across cell types varying from hundreds (e.g. auxilin) to millions (e.g. clathrin). These variations contain important information about function, but without integration with the interaction network, they cannot capture how supply and demand for each protein depends on binding to shared and distinct partners. Here we construct the interface-resolved network of 82 proteins involved in CME and establish a metric, a stoichiometric balance ratio (SBR), that quantifies whether each protein in the network has an abundance that is sub- or super-stoichiometric dependent on the global competition for binding. We find that highly abundant proteins (like clathrin) are super-stoichiometric, but that not all super-stoichiometric proteins are highly abundant, across three cell populations (HeLa, fibroblast, and neuronal synaptosomes). Most strikingly, within all cells there is significant competition to bind shared sites on clathrin and the central AP-2 adaptor by other adaptor proteins, resulting in most being in excess supply. Our network and systematic analysis, including response to perturbations of network components, show how competition for shared binding sites results in functionally similar proteins having widely varying stoichiometries, due to variations in both abundance and their unique network of binding partners.

Reaction rates estimation for the endocytic reception in extracellular vesicles-mediated communications

“…Please note that this model is just to apply to some of endocytosis mechanisms, which are already studied in the literature. [1,7,10,21].…”

The Regulation of Epidermal Growth Factor Receptor Signaling

“…Another possibility is that other interactions, such as the interaction with EGFR or ubiquitin ligases contributes to Ack1/2 recruitment to CCPs. This is consistent with many endocytosis proteins recruited to CCPs via multiple interactions, instead of single protein-protein interaction interfaces (Smith et al, 2017;Duan et al, 2022;Praefcke et al, 2004). To test this, I could examine the CCP localization of additional mutants of Ack1/2 using similar approaches as I have done for the 4A and ΔSH3 mutants of Ack1/2.…”

“…This domain of Ack1 bears similarity to the clathrinbinding domain of b-arrestin, a clathrin-adaptor protein that is shown to directly mediate clathrin-dependent β2-adrenergic receptor (β2AR) endocytosis (Goodman et al, 1996;Teo et al, 2001b). In the 4A mutant (clathrin binding defective mutant), Ack1 may still be recruited to CCPs based on competitive displacement of other protein-protein interactions in the clathrin interaction network (Duan et al, 2022). Hence, despite Ack1 localizing to CCPs without direct interaction with clathrin heavy chain, this disrupted protein interaction motif may nonetheless impair EGF-stimulated activation of PI3K-Akt signaling.…”

Dynamin1 long- and short-tail isoforms exploit distinct recruitment and spatial patterns to form endocytic nanoclusters

“…We defined a minimal model with three components to limit the number of free parameters as much as possible while still capturing the observed experimental dynamics: (1) Dyn1, (2) a uniformly distributed membrane recruiter and (3) a cluster-associated recruiter or “activator” that is spatially restricted to cluster sites. The membrane recruiter represents a population of proteins (such as a variety of SH3-domain containing proteins 46 ) and lipids (such as PI(4,5)P 2 ) already on the plasma membrane that recruit Dyn1. The cluster-associated “activator” mimics Dyn1-binding proteins assembled at sites of clathrin-coated pits (such as amphiphysin and intersectin) 3 .…”