The ERC1 scaffold protein implicated in cell motility drives the assembly of a liquid phase

Abstract

Several cellular processes depend on networks of proteins assembled at specific sites near the plasma membrane. Scaffold proteins assemble these networks by recruiting relevant molecules. The scaffold protein ERC1/ELKS and its partners promote cell migration and invasion, and assemble into dynamic networks at the protruding edge of cells. Here by electron microscopy and single molecule analysis we identify ERC1 as an extended flexible dimer. We found that ERC1 scaffolds form cytoplasmic condensates with a behavior that is consistent with liquid phases that are modulated by a predicted disordered region of ERC1. These condensates specifically host partners of a network relevant to cell motility, including liprin-α1, which was unnecessary for the formation of condensates, but influenced their dynamic behavior. Phase separation at specific sites of the cell periphery may represent an elegant mechanism to control the assembly and turnover of dynamic scaffolds needed for the spatial localization and processing of molecules.

ELKS1 Captures Rab6-Marked Vesicular Cargo in Presynaptic Nerve Terminals

“…They contain coiled-coil stretches that form extended, homodimeric antenna-like tethers and contain Rab-binding sites to capture vesicles. ELKS proteins indeed share key properties of Golgins in addition to the sequence homology: their coiled-coils form elongated homo- and heterodimers ( Deguchi-Tawarada et al, 2004 ; Sala et al, 2019 ), and they bind to Rab6 ( Figure 3 ; Monier et al, 2002 ). The key difference between ELKS and Golgins is their localization.…”

“…How could ELKS1 be anchored? ELKS proteins bind to several active zone proteins, including RIM, Liprin-α, and Bassoon, and they homodimerize with one another and form heterooligomers ( Deguchi-Tawarada et al, 2004 ; Held and Kaeser, 2018 ; Ko et al, 2003 ; Ohtsuka et al, 2002 ; Sala et al, 2019 ; Takao-Rikitsu et al, 2004 ; Wang et al, 2002 ). Through interactions with active zone proteins and oligomerization, it is possible that ELKS1 forms a filamentous network throughout the nerve terminal that is anchored at the active zone.…”

“…An alternative model is that ELKS participates in liquid-phase condensates that tether vesicles. ELKS1 was recently shown to undergo phase transitions and to form liquid droplets within cells, and the N-terminal half alone is sufficient to form phase condensates ( Sala et al, 2019 ). In this model, ELKS is dynamically tethered by its property to from a liquid phase or by participating in a Synapsin-based liquid phase that tethers synaptic vesicles ( Milovanovic et al, 2018 ).…”

Phosphorylation triggers presynaptic phase separation of Liprin-α3 to control active zone structure

“…During development, it is likely that phase separation and recruitment of presynaptic material occur independent of phosphorylation and involve additional proteins. For instance, other Liprin-α isoforms or ELKS, which captures synaptic material and phase separates 32, 61 , may play active roles. In conclusion, an overarching model arises in which phase transition of presynaptic proteins is essential to recruit and assemble presynaptic material into functional molecular machines.…”

“…This may include interactions with vesicles, cytoskeletal elements and trafficking machinery, potentially explaining why some synaptic vesicles are lost in Liprin-α deficient synapses[9][10][11]59,60 .During development, it is likely that phase separation and recruitment of presynaptic material occur independent of phosphorylation and involve additional proteins. For instance, other Liprinα isoforms or ELKS, which captures synaptic material and phase separates32,61 , may play active roles. In conclusion, an overarching model arises in which phase transition of presynaptic proteins is essential to recruit and assemble presynaptic material into functional molecular machines.Materials and methodsAssessments of droplets in transfected HEK293T cellsHEK293T cells were plated on 0.1 mm thick coverslips and transfected with plasmids expressing proteins of interest under the CMV promoter.…”