Abstract
The TOCA family of F-BAR–containing proteins bind to and remodel lipid bilayers via their conserved F-BAR domains, and regulate actin dynamics via their N-Wasp binding SH3 domains. Thus, these proteins are predicted to play a pivotal role in coordinating membrane traffic with actin dynamics during cell migration and tissue morphogenesis. By combining genetic analysis in Caenorhabditis elegans with cellular biochemical experiments in mammalian cells, we showed that: i) loss of CeTOCA proteins reduced the efficiency of Clathrin-mediated endocytosis (CME) in oocytes. Genetic interference with CeTOCAs interacting proteins WSP-1 and WVE-1, and other components of the WVE-1 complex, produced a similar effect. Oocyte endocytosis defects correlated well with reduced egg production in these mutants. ii) CeTOCA proteins localize to cell–cell junctions and are required for proper embryonic morphogenesis, to position hypodermal cells and to organize junctional actin and the junction-associated protein AJM-1. iii) Double mutant analysis indicated that the toca genes act in the same pathway as the nematode homologue of N-WASP/WASP, wsp-1. Furthermore, mammalian TOCA-1 and C. elegans CeTOCAs physically associated with N-WASP and WSP-1 directly, or WAVE2 indirectly via ABI-1. Thus, we propose that TOCA proteins control tissues morphogenesis by coordinating Clathrin-dependent membrane trafficking with WAVE and N-WASP–dependent actin-dynamics.
Highlights
The coordination and functional cooperation between endocytic trafficking of membranes and membrane proteins with actinbased motility is required for the correct execution of many cellular phenotypes
We tested whether the SH3 domain of CeTOCA-1 is functional and able to associate with one of the known mammalian ligands, N-WASP or C. elegans WSP-1
Immobilized GST-SH3 domains of CeTOCA-1 and CeTOCA-2 bound to mammalian N-WASP and CeWSP-1 (Figure 1C and Figure S1F), CeTOCA-2 SH3 was less efficient than the SH3 domains of human TOCA-1 or CeTOCA-1
Summary
The coordination and functional cooperation between endocytic trafficking of membranes and membrane proteins with actinbased motility is required for the correct execution of many cellular phenotypes. Results obtained in several species have established that endocytosis and trafficking events rely on propelling forces generated by actin treadmilling [1]. Consistent with these results, an increasing number of actin binding and regulatory proteins have been shown to participate in a variety of internalization and trafficking processes, controlling the signaling response of cells to extracellular stimuli. Genetic and cellular biochemical evidence has revealed how cycles of endocytosis and recycling of plasma membranes and plasma membrane proteins are essential to promote the spatial restriction of signaling [2,3,4,5]
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