Collapsin Response Mediator Protein-2 (Crmp2) Regulates Trafficking by Linking Endocytic Regulatory Proteins to Dynein Motors

Abstract

Endocytosis is a conserved cellular process in which nutrients, lipids, and receptors are internalized and transported to early endosomes, where they are sorted and either channeled to degradative pathways or recycled to the plasma membrane. MICAL-L1 and EHD1 are important regulatory proteins that control key endocytic transport steps. However, the precise mechanisms by which they mediate transport, and particularly the mode by which they connect to motor proteins, have remained enigmatic. Here we have identified the collapsin response mediator protein-2 (Crmp2) as an interaction partner of MICAL-L1 in non-neuronal cells. Crmp2 interacts with tubulin dimers and kinesin and negatively regulates dynein-based transport in neuronal cells, but its expression and function in non-neuronal cells have remained poorly characterized. Upon Crmp2 depletion, we observed dramatic relocalization of internalized transferrin (Tf) from peripheral vesicles to the endocytic recycling compartment (ERC), similar to the effect of depleting either MICAL-L1 or EHD1. Moreover, Tf relocalization to the ERC could be inhibited by interfering with microtubule polymerization, consistent with a role for uncoupled motor proteinbased transport upon depletion of Crmp2, MICAL-L1, or EHD1. Finally, transfection of dynamitin, a component of the dynactin complex whose overexpression inhibits dynein activity, prevented the relocalization of internalized Tf to the ERC upon depletion of Crmp2, MICAL-L1, or EHD1. These data provide the first trafficking regulatory role for Crmp2 in non-neuronal cells and support a model in which Crmp2 is an important endocytic regulatory protein that links MICAL-L1⅐EHD1-based vesicular transport to dynein motors.The complexity of the endocytic pathways has broadened in recent years upon discovery of the involvement of the four C-terminal Eps 15 homology domain (EHD) 3 proteins (reviewed in Ref. 1). Since the single Caenorhabditis elegans ortholog (known as RME-1) was originally identified as a regulator of yolk receptor recycling (2), its closest mammalian homolog, EHD1, was found to regulate the recycling of receptors that traverse both the clathrin-dependent (3, 4) and the clathrin-independent (5, 6) internalization pathways. Despite similarities to the Ras family of GTP-binding proteins (5, 7), EHD proteins bind and hydrolyze ATP (7-9), a function necessary for their localization to tubular and vesicular membranes (5,7,10).Proteins that contain EH domains interact with the tripeptide asparagine-proline-phenylalanine (NPF) (11). Recent studies have determined that the EH domains of the C-terminal EHD proteins have a highly positively charged surface (12) and selectively interact with NPF motifs followed by clusters of acidic residues (13, 14). Moreover, EHD proteins coordinate endocytic transport with Rab proteins through their interactions with common effectors that contain such NPF motifs (9, 15). More recently, it was demonstrated that EHD1 interacts with the NPF-containing MICAL family protein, MICAL-L1, a Rab8 ef...

A functional coupling between CRMP1 and Nav1.7 for retrograde propagation of Semaphorin3A signaling

“…These findings suggest that CRMP1 mediates Sema3A-induced retrograde axonal transport. It has been reported that CRMP2 is involved in kinesin-1-dependent transport of the Sra1-WAVE1 complex (Arimura et al, 2009) and in regulation of trafficking by linking endocytic regulatory proteins to dynein motors (Rahajeng et al, 2010). CRMP2 has been reported to be able to directly bind to voltage-dependent Na + channels, modulating the channel's slow inactivation (Dustrude et al, 2013;Wang et al, 2010).…”

MICAL-Family Proteins: Complex Regulators of the Actin Cytoskeleton

“…The CH and LIM domains of MICAL-L1 bind to CRMP2 (60). On the other hand, the LIM domain of MICAL-L2 interacts with F-actin and promotes F-actin crosslinking (60).…”

Human T Lymphotropic Virus Type 1 Increases T Lymphocyte Migration by Recruiting the Cytoskeleton Organizer CRMP2

“…Collectively, these data demonstrated that HTLV-1 infection, possibly through Tax activity, profoundly altered posttranslational processing of CRMP2, resulting in more active molecular form in T lymphocyte and elevated migration as a consequence. Tax and CRMP2 colocalized at uropod and cell-cell contact in HTLV-1-infected T cells Given 1) the crucial role of CRMP2 in remodeling neural cell and T lymphocyte cytoskeleton by direct binding to microtubules (25,44) and also in participation, as a cargo receptor, in the transport of specific vesicles (45,46), 2) the ability of Tax protein to modulate CRMP2 activity in infected lymphocyte, in particular cytoskeleton binding (shown in this paper), 3) the capacity of infected cell to transfer Tax though a well organized cell-cell contact (the so-called virological synapse (47), and 4) the role of microtubule in T cell signaling (48), we suspected CRMP2 to bind Tax protein at specific sites in lymphocyte cytoplasm to cooperate. Preliminary analyses using immunoprecipitation and CRMP2-GST pull-down strategy were unable to detect a direct association between these proteins.…”