Oligomerization, F-actin Interaction, and Membrane Association of the Ubiquitous Mammalian Coronin 3 Are Mediated by Its Carboxyl Terminus

Abstract

Coronin 3 is a ubiquitously expressed member of the coronin protein family in mammals. In fibroblasts and HEK 293 cells, it is localized both in the cytosol and in the submembranous cytoskeleton, especially at lamellipodia and membrane ruffles. The carboxyl terminus of all coronins contains a coiled coil suggested to mediate dimerization. We show here that in contrast to other coronin homologues, the recombinant human coronin 3 carboxyl terminus forms oligomers rather than dimers, and that this part is sufficient to bind to and cross-link F-actin in vitro. The carboxyl terminus alone also conferred membrane association in vivo, and removal of the coiled coil abolished membrane localization but not in vitro F-actin binding. Coronin 3 is exclusively extracted as an oligomer from both the cytosol and the membrane fraction. Because oligomerization was not reported for other coronins, it might be a key feature governing coronin 3-specific functions. Cytosolic coronin 3 showed a high degree of phosphorylation, which is likely to regulate the subcellular localization of the protein.

The identification of a new actin-binding region in p57

“…In this study, we found that the C-terminal region of p57, p57 , had the similar actin-binding property as full-length p57, as demonstrated by co-sedimentation of expressed protein with F-actin in vitro and co-localization of expressed protein with actin cytoskeleton in vivo. These results revealed that the C-terminal region of p57 might also be responsible for binding F-actin which was consistent with actin-binding sites of other coronin-like proteins such as Xcoronin and Hcoronin3 [13,14].…”

“…However, it has been reported that the deletion of the C-terminal 65 amino acids from Xcoronin, a Xenopus homologue of coronin, significantly reduces its actin-binding activity [13]. In another report, the F-actin interaction of human coronin 3 (Hcoronin3) is mediated by its C-terminus [14]. These studies suggest that the C-terminus of p57 may contribute to its actin binding.…”

The GDP-dependent Rab27a effector coronin 3 controls endocytosis of secretory membrane in insulin-secreting cell lines

“…By contrast, FM4-64 uptake was inhibited in cells expressing GFP-fused dominant-negative coronin-3-ΔC, which bound GDP-Rab27a (Fig. 2C) without actinbundling activity (Spoerl et al, 2002) (Fig. 3C).…”

“…Therefore, residues 1-314 of coronin 3, which form the β-propeller structure for protein-protein interactions (Appleton et al, 2006), are essential for the binding to GDP-Rab27a. Rab27a-T23N preferentially interacted with coronin-3-N, coronin-3-WD, and coronin-3ΔC compared with coronin-3-WT, probably because Rab27a binding might be hindered by the C-terminus of coronin 3, which intramolecularly binds the N-terminus (Spoerl et al, 2002).…”

“…2C). Rab27a-T23N bound coronin-3-WT (amino acids 1-474), coronin-3-N (1-71), coronin-3-WD (72-314, containing the tryptophan and aspartate repeat), and coronin-3-ΔC (1-314) but not GFP and coronin-3-C (315-474), which is responsible for the actin binding (Spoerl et al, 2002) (Fig. 2C).…”