Abstract
Assemblies of actin and its regulators underlie the dynamic morphology of all eukaryotic cells. To understand how actin regulatory proteins work together to generate actin-rich structures such as filopodia, we analyzed the localization of diverse actin regulators within filopodia in Drosophila embryos and in a complementary in vitro system of filopodia-like structures (FLSs). We found that the composition of the regulatory protein complex where actin is incorporated (the filopodial tip complex) is remarkably heterogeneous both in vivo and in vitro. Our data reveal that different pairs of proteins correlate with each other and with actin bundle length, suggesting the presence of functional subcomplexes. This is consistent with a theoretical framework where three or more redundant subcomplexes join the tip complex stochastically, with any two being sufficient to drive filopodia formation. We provide an explanation for the observed heterogeneity and suggest that a mechanism based on multiple components allows stereotypical filopodial dynamics to arise from diverse upstream signaling pathways.
Highlights
The regulation of actin polymerization is crucial for numerous preexisting actin network generated by the Arp2/3 complex cell functions, including cell migration, adhesion, and epithelial becoming bundled by Fascin (Svitkina et al, 2003; Vignjevic closure (Jacinto et al, 2002; Jacquemet et al, 2015) and is often et al, 2003; Yang and Svitkina, 2011); and (3) membrane-bound disrupted in disease, such as cancer metastasis and intracellular adaptor proteins recruiting Ena/vasodilator-stimulated phosphoprotein (VASP) (Disanza et al, 2013; infection by pathogens (Bendris and Schmid, 2017; Molinie and Mattila et al, 2007), which could coexist with either formin or Gautreau, 2018; Stradal and Schelhaas, 2018; Tan et al, 2013)
We found that a cell-free system of filopodia-like structures (FLSs) is characterized by similar heterogeneities, and it allowed us to make large-scale combinatorial measurements of the correlations of actin regulators with each other and the morphology of the actin bundle
Heterogeneous tip complexes and exponentially distributed filopodial lengths in vivo in Drosophila We first examined the localization of Ena and the Arp2/3 complex nucleation promoting factor Scar/WAVE at the filopodial tip complex in vivo using the Drosophila embryo
Summary
The regulation of actin polymerization is crucial for numerous preexisting actin network generated by the Arp2/3 complex cell functions, including cell migration, adhesion, and epithelial becoming bundled by Fascin (Svitkina et al, 2003; Vignjevic closure (Jacinto et al, 2002; Jacquemet et al, 2015) and is often et al, 2003; Yang and Svitkina, 2011); and (3) membrane-bound disrupted in disease, such as cancer metastasis and intracellular adaptor proteins recruiting Ena/VASP (Disanza et al, 2013; infection by pathogens (Bendris and Schmid, 2017; Molinie and Mattila et al, 2007), which could coexist with either formin or Gautreau, 2018; Stradal and Schelhaas, 2018; Tan et al, 2013). One of the best examples is filopodia, with their whether the subtypes reflect differences between cell types or characteristic membrane-associated “tip complex” where new coexist in the same cell and whether they impart particular actin monomers are incorporated, leading to rapid extension of properties to the growing filopodia. We recently examined this the filopodia from the cell surface (Applewhite et al, 2007; question by measuring whether the amount of Ena and VASP at Mallavarapu and Mitchison, 1999).
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