WIP-1 and DBN-1 promote scission of endocytic vesicles by bridging actin and Dynamin-1 in theC. elegansintestine

Abstract

There has been a consensus that actin plays an important role in scission of the clathrin-coated pits (CCPs) together with large GTPases of the dynamin family in metazoan cells. However, the recruitment, regulation and functional interdependence of actin and dynamin during this process remain inadequately understood. Here, based on small-scale screening and in vivo live-imaging techniques, we identified a novel set of molecules underlying CCP scission in the multicellular organism Caenorhabditis elegans. We found that loss of Wiskott−Aldrich syndrome protein (WASP)-interacting protein (WIP-1) impaired CCP scission in a manner that is independent of the C. elegans homolog of WASP/N-WASP (WSP-1) and is mediated by direct binding to G-actin. Moreover, the cortactin-binding domain of WIP-1 serves as the binding interface for DBN-1 (also known in other organisms as Abp1), another actin-binding protein. We demonstrate that the interaction between DBN-1 and F-actin is essential for Dynamin-1 (DYN-1) recruitment at endocytic sites. In addition, the recycling regulator RME-1, a homolog of mammalian Eps15 homology (EH) domain-containing proteins, is increasingly recruited at the arrested endocytic intermediates induced by F-actin loss or DYN-1 inactivation, which further stabilizes the tubular endocytic intermediates. Our study provides new insights into the molecular network underlying F-actin participation in the scission of CCPs. This article has an associated First Person interview with the first author of the paper.

SNX-3 mediates retromer-independent tubular endosomal recycling by opposing EEA-1-facilitated trafficking

“…elegans . It specifically localizes to basolateral recycling endosomes and promotes the recycling of proteins from endosomes back to the cell surface [ 21 , 28 , 29 ]. In WT intestines, most of the subplasmalemmal hTAC tubules colocalized with RME-1-labeled structures, indicating efficient recycling.…”

“…The lmp-1 cDNA fused by TagRFP-T with a linker sequence Gly-Ala at 3 0 -terminus was amplified and cloned into Pvha-6::TagRFP-T by using Xmal and KpnI sites to generate Pvha-6::LMP-1::TagRFP-T. Pvha-6::GFP::EEA-1 and Pvha-6:: GFP::SNX-3 were constructed by ligating eea-1 genomic DNA or snx-3 cDNA into Pvha-6:: GFP vector at NheI and KpnI sites, respectively. The Pvha-6::TagRFP-T::RAB-5 were generated previously [28]. To generate Pvha-6::ARF-6::mCherry, the arf-6 cDNA was amplified and inserted into Pvha-6::mCherry vector at the Smal and Agel sites.…”

Drebrin-like protein regulates body bending ofC. elegansvia suppression of NCA cation leak channels

“…C. elegans DBN-1 is a multi-domain protein that contains an N-terminal ADF-H domain, three coiled-coils (3xCC), a proline-rich sequence and a C-terminal SH3 domain, of which the 3xCC domain of DBN-1 was shown to bind F-actin in vitro 29 . The ok925 mutation that truncates DBN-1 after two coiled-coils leads to a mild disorganization of actin filaments during body-wall muscle contraction 29 and affects vesicle scission during endocytosis in the intestine 27 . However, the remaining N-terminal part of DBN-1 in the ok925 mutant is sufficient to support its wild-type-like locomotion (Fig.…”

“…Drebrin-like protein is, for example, an important component of the endocytic machinery. Being selectively recruited at the late stage of the formation of clathrin-coated pits, it directly interacts with dynamin, mediating vesicle scission 25,26,27 . Elimination of the mAbp1 in mice results in moderate reduction of both receptor-mediated and synaptic endocytosis and in a severe impairment of synaptic vesicle recycling.…”

Dimerization of human drebrin-like protein governs its biological activity

“…). However, the ok925 has a mild disorganization of actin filaments during body-wall muscle contraction 3 and defective vesicle scission during endocytosis in the intestines37 . Taken together, our data demonstrate that the amino-acid sequence of DBNL defines its existence as a dimer and that DBNL can efficiently interact with actin filaments inside a cell only in its native conformation.…”