Microtubule Plus-end Tracking Proteins Research Articles

CLIP170 inhibits the metastasis and EMT of papillary thyroid cancer through the TGF-β pathway.

Metastasis poses a significant challenge in combating tumors. Even in papillary thyroid cancer (PTC), which typically exhibits a favorable prognosis, high recurrence rates are attributed to metastasis. Cytoplasmic linker protein 170 (CLIP170) functions as a classical microtubule plus-end tracking protein (+TIP) and has shown close association with cell migration. Nevertheless, the specific impact of CLIP170 on PTC cells remains to be elucidated. Our analysis of the GEO and TCGA databases unveiled an association between CLIP170 and the progression of PTC. To explore the impact of CLIP170 on PTC cells, we conducted various assays. We evaluated its effects through CCK-8, wound healing assay, and transwell assay after knocking down CLIP170. Additionally, the influence of CLIP170 on the cellular actin structure was examined via immunofluorescence; we further investigated the molecular expressions of epithelial-mesenchymal transition (EMT) and the transforming growth factor-β (TGF-β) signaling pathways through Western blotting and RT-qPCR. These findings were substantiated through an in vivo nude mouse model of lung metastasis. We observed a decreased expression of CLIP170 in PTC in contrast to normal thyroid tissue. Functionally, the knockdown of CLIP170 (CLIP170KD) notably enhanced the metastatic potential and EMT of PTC cells, both in vitro and in vivo. Mechanistically, CLIP170KD triggered the activation of the TGF-β pathway, subsequently promoting tumor cell migration, invasion, and EMT. Remarkably, the TGF-β inhibitor LY2157299 effectively countered TGF-β activity and significantly reversed tumor metastasis and EMT induced by CLIP170 knockdown. In summary, these findings collectively propose CLIP170 as a promising therapeutic target to mitigate metastatic tendencies in PTC.

Studying Microtubule Dynamics in Human Neurons: Two-Dimensional Microtubule Tracing and Kymographs in iPSC- and SH-SY5Y-Derived Neurons for Tau Research.

The study of microtubule (MT) dynamics is essential for the understanding of cellular transport, cell polarity, axon formation, and other neurodevelopmental mechanisms. All these processes rely on the constant transition between assembly and disassembly of tubulin polymers to/from MTs, known as dynamic instability. This process is well-regulated, among others, by phosphorylation of microtubule-associated proteins (MAP), including the Tau protein. Protein kinases, in particular the microtubule affinity regulating kinase (MARK), regulate the MT-Tau interaction, inducingTau dissociation by phosphorylation. Phosphorylated Tau dissociates from microtubules forming insoluble aggregates known as neurofibrillary tangles. These accumulations of hyperphosphorylated Tau in the neurons disrupt the physiological MT-based transport machinery within the cell and can potentially lead to the development of neurodegenerative disorders, such as Alzheimer's disease (AD) and related tauopathies. Further investigations on the MT cytoskeleton dynamics are essential as they may elucidate pathomechanisms of neurodegenerative diseases - particularly tauopathies - as well as fundamental neurodevelopmental processes.The study of thedynamic assembly and disassembly of the MT network requires live-cell imaging rather than conventional immunocytochemistry based on fixed samples. To investigate MT dynamics, we perform live-cell imaging of neurons transfected with a fluorescently tagged version of the microtubule plus-end tracking protein (+TIP) EB3. This protein associates with the growing ends of MTs and thus visualizes MT growth in real time. Our imaging analysis protocol allows the determination of quantity, orientation, and velocity of MT growth in the soma and neurites of transfected neurons,using ImageJ-based tracking software and kymographs. Furthermore, functional effects of Tau and MARK kinases on the MT cytoskeleton can be assessed by overexpression or downregulation experiments of the respective protein prior to the live imaging assay. We use two different human neuronal cell models, naive and differentiated SH-SY5Y neuroblastoma cells, and neurons derived from induced pluripotent stem cells (iPSCs), both of which have shown success as models to study Tau-related pathologies.This protocol describes an optimized method for analysis of microtubule dynamics using fluorescent tagged EB3 protein as microtubule plus end marker. In this chapter, we outline the process of neuronal transfection, live-cell imaging, and necessary time-lapse image analysis based on ImageJ in two human-derived neuronal systems, which are suitable for the analysis of Tau trafficking and sorting studies.

Activation of the DNA damage checkpoint perturbs asymmetric localization of Kar9 to spindle pole bodies in Saccharomyces cerevisiae

During cell cycle progression in Saccharomyces cerevisiae, spindle pole bodies (SPBs) are duplicated during the G1/S-phase transition. SPBs are crucial for the organization of both the spindle and astral microtubules, and their orientation defines the direction of nuclear division. In this process, an old SPB, which serves as the template SPB during the duplication process, is oriented toward the bud side. The patterning microtubule plus-end tracking protein, Kar9, plays an important role in the orientation of SPBs by asymmetrically localizing to the old SPB. Here, methylglyoxal (MG), a metabolite derived from glycolysis, was found to perturb asymmetric Kar9 localization and influence proper positioning of the old SPB. MG activated the DNA damage checkpoint pathway, and MG-induced perturbation of asymmetric Kar9 localization was abolished by the deletion of MEC1, a sensor for the DNA damage checkpoint pathway. Methyl methanesulfonate, a DNA-alkylating agent, also perturbed asymmetric Kar9 localization. Our results suggest that activation of the DNA damage checkpoint pathway perturbs the asymmetric Kar9 localization required for proper positioning of SPBs.

Interaction between Teneurin-2 and microtubules via EB proteins provides a platform for GABAA receptor exocytosis.

Neurons form dense neural circuits by connecting to each other via synapses and exchange information through synaptic receptors to sustain brain activities. Excitatory postsynapses form and mature on spines composed predominantly of actin, while inhibitory synapses are formed directly on the shafts of dendrites where both actin and microtubules (MTs) are present. Thus, it is the accumulation of specific proteins that characterizes inhibitory synapses. In this study, we explored the mechanisms that enable efficient protein accumulation at inhibitory postsynapse. We found that some inhibitory synapses function to recruit the plus end of MTs. One of the synaptic organizers, Teneurin-2 (TEN2), tends to localize to such MT-rich synapses and recruits MTs to inhibitory postsynapses via interaction with MT plus-end tracking proteins EBs. This recruitment mechanism provides a platform for the exocytosis of GABAA receptors. These regulatory mechanisms could lead to a better understanding of the pathogenesis of disorders such as schizophrenia and autism, which are caused by excitatory/inhibitory (E/I) imbalances during synaptogenesis.

Microtubule plus-end tracking proteins: novel modulators of cardiac sodium channels and arrhythmogenesis.

The cardiac sodium channel NaV1.5 is an essential modulator of cardiac excitability, with decreased NaV1.5 levels at the plasma membrane and consequent reduction in sodium current (INa) leading to potentially lethal cardiac arrhythmias. NaV1.5 is distributed in a specific pattern at the plasma membrane of cardiomyocytes, with localization at the crests, grooves, and T-tubules of the lateral membrane and particularly high levels at the intercalated disc region. NaV1.5 forms a large macromolecular complex with and is regulated by interacting proteins, some of which are specifically localized at either the lateral membrane or intercalated disc. One of the NaV1.5 trafficking routes is via microtubules (MTs), which are regulated by MT plus-end tracking proteins (+TIPs). In our search for mechanisms involved in targeted delivery of NaV1.5, we here provide an overview of previously demonstrated interactions between NaV1.5 interacting proteins and +TIPs, which potentially (in)directly impact on NaV1.5 trafficking. Strikingly, +TIPs interact extensively with several intercalated disc- and lateral membrane-specific NaV1.5 interacting proteins. Recent work indicates that this interplay of +TIPs and NaV1.5 interacting proteins mediates the targeted delivery of NaV1.5 at specific cardiomyocyte subcellular domains, while also being potentially relevant for the trafficking of other ion channels. These observations are especially relevant for diseases associated with loss of NaV1.5 specifically at the lateral membrane (such as Duchenne muscular dystrophy), or at the intercalated disc (for example, arrhythmogenic cardiomyopathy), and open up potential avenues for development of new anti-arrhythmic therapies.

Multivalency ensures persistence of a +TIP body at specialized microtubule ends

Microtubule plus-end tracking proteins (+TIPs) control microtubule specialization and are as such essential for cell division and morphogenesis. Here we investigated interactions and functions of the budding yeast Kar9 network consisting of the core +TIP proteins Kar9 (functional homologue of APC, MACF and SLAIN), Bim1 (orthologous to EB1) and Bik1 (orthologous to CLIP-170). A multivalent web of redundant interactions links the three +TIPs together to form a ‘+TIP body’ at the end of chosen microtubules. This body behaves as a liquid condensate that allows it to persist on both growing and shrinking microtubule ends, and to function as a mechanical coupling device between microtubules and actin cables. Our study identifies nanometre-scale condensates as effective cellular structures and underlines the power of dissecting the web of low-affinity interactions driving liquid–liquid phase separation in order to establish how condensation processes support cell function.

Ubiquitination of CLIP-170 family protein restrains polarized growth upon DNA replication stress

Microtubules play a crucial role during the establishment and maintenance of cell polarity. In fission yeast cells, the microtubule plus-end tracking proteins (+TIPs) (including the CLIP-170 homologue Tip1) regulate microtubule dynamics and also transport polarity factors to the cell cortex. Here, we show that the E3 ubiquitin ligase Dma1 plays an unexpected role in controlling polarized growth through ubiquitinating Tip1. Dma1 colocalizes with Tip1 to cortical sites at cell ends, and is required for ubiquitination of Tip1. Although the absence of dma1+ does not cause apparent polar growth defects in vegetatively growing cells, Dma1-mediated Tip1 ubiquitination is required to restrain polar growth upon DNA replication stress. This mechanism is distinct from the previously recognized calcineurin-dependent inhibition of polarized growth. In this work, we establish a link between Dma1-mediated Tip1 ubiquitination and DNA replication or DNA damage checkpoint-dependent inhibition of polarized growth in fission yeast.

MAPRE2 regulates the first meiotic progression in mouse oocytes

Microtubule plus-end tracking proteins (+TIPs) associate with growing microtubule plus ends and control microtubule dynamics and interactions with different cellular structures during cell division, cell migration and morphogenesis. Microtubule-associated RP/EB family member 2 (MAPRE2/EB2) is a highly conserved core component of +TIPs networks, but whether this molecule is required for mammalian meiotic progression is unknown. In this study, we investigated the expression and function of MAPRE2 during oocyte maturation. Our results showed that MAPRE2 was consistently expressed from germinal vesicle (GV) to metaphase II (MII) stages and that MAPRE2 was distributed in the cytoplasm of oocytes at GV stage and along the spindle at metaphase I (MI) and MII stages. Small interfering RNA-mediated knockdown of Mapre2 severely impaired microtubule stability, kinetochore-microtubule attachment, and chromosome alignment and subsequently caused spindle assembly checkpoint (SAC) activation and cyclin B1 nondegradation, leading to failure of chromosome segregation and first polar body extrusion. This study demonstrates for the first time that MAPRE2 plays an important role during mouse oocyte meiosis.

The CLIP-170 N-terminal domain binds directly to both F-actin and microtubules in a mutually exclusive manner

The cooperation between the actin and microtubule (MT) cytoskeletons is important for cellular processes such as cell migration and muscle cell development. However, a full understanding of how this cooperation occurs has yet to be sufficiently developed. The MT plus-end tracking protein CLIP-170 has been implicated in this actin–MT coordination by associating with the actin-binding signaling protein IQGAP1 and by promoting actin polymerization through binding with formins. Thus far, the interactions of CLIP-170 with actin were assumed to be indirect. Here, we demonstrate using high-speed cosedimentation assays that CLIP-170 can bind to filamentous actin (F-actin) directly. We found that the affinity of this binding is relatively weak but strong enough to be significant in the actin-rich cortex, where actin concentrations can be extremely high. Using CLIP-170 fragments and mutants, we show that the direct CLIP-170–F-actin interaction is independent of the FEED domain, the region that mediates formin-dependent actin polymerization, and that the CLIP-170 F-actin-binding region overlaps with the MT-binding region. Consistent with these observations, in vitro competition assays indicate that CLIP-170–F-actin and CLIP-170–MT interactions are mutually exclusive. Taken together, these observations lead us to speculate that direct CLIP-170–F-actin interactions may function to reduce the stability of MTs in actin-rich regions of the cell, as previously proposed for MT end-binding protein 1.

The microtubule plus-end tracking protein Bik1 is required for chromosome congression.

During mitosis, sister chromatids congress on both sides of the spindle equator to facilitate the correct partitioning of the genomic material. Chromosome congression requires a finely tuned control of microtubule dynamics by the kinesin motor proteins. In Saccharomyces cerevisiae, the kinesin proteins Cin8, Kip1, and Kip3 have a pivotal role in chromosome congression. It has been hypothesized that additional proteins that modulate microtubule dynamics are involved. Here, we show that the microtubule plus-end tracking protein Bik1—the budding yeast ortholog of CLIP-170—is essential for chromosome congression. We find that nuclear Bik1 localizes to the kinetochores in a cell cycle–dependent manner. Disrupting the nuclear pool of Bik1 with a nuclear export signal (Bik1-NES) leads to slower cell-cycle progression characterized by a delayed metaphase–anaphase transition. Bik1-NES cells have mispositioned kinetochores along the spindle in metaphase. Furthermore, using proximity-dependent methods, we identify Cin8 as an interaction partner of Bik1. Deleting CIN8 reduces the amount of Bik1 at the spindle. In contrast, Cin8 retains its typical bilobed distribution in the Bik1-NES mutant and does not localize to the unclustered kinetochores. We propose that Bik1 functions with Cin8 to regulate kinetochore–microtubule dynamics for correct kinetochore positioning and chromosome congression.

Dynactin 1 negatively regulates HIV-1 infection by sequestering the host cofactor CLIP170

Many viruses directly engage and require the dynein-dynactin motor-adaptor complex in order to transport along microtubules (MTs) to the nucleus and initiate infection. HIV type 1 (HIV-1) exploits dynein, the dynein adaptor BICD2, and core dynactin subunits but unlike several other viruses, does not require dynactin-1 (DCTN1). The underlying reason for HIV-1's variant dynein engagement strategy and independence from DCTN1 remains unknown. Here, we reveal that DCTN1 actually inhibits early HIV-1 infection by interfering with the ability of viral cores to interact with critical host cofactors. Specifically, DCTN1 competes for binding to HIV-1 particles with cytoplasmic linker protein 170 (CLIP170), one of several MT plus-end tracking proteins (+TIPs) that regulate the stability of viral cores after entry into the cell. Outside of its function as a dynactin subunit, DCTN1 also functions as a +TIP that we find sequesters CLIP170 from incoming particles. Deletion of the Zinc knuckle (Zn) domain in CLIP170 that mediates its interactions with several proteins, including DCTN1, increased CLIP170 binding to virus particles but failed to promote infection, further suggesting that DCTN1 blocks a critical proviral function of CLIP170 mediated by its Zn domain. Our findings suggest that the unique manner in which HIV-1 binds and exploits +TIPs to regulate particle stability leaves them vulnerable to the negative effects of DCTN1 on +TIP availability and function, which may in turn have driven HIV-1 to evolve away from DCTN1 in favor of BICD2-based engagement of dynein during early infection.

Structure and regulation of the microtubule plus-end tracking protein Kar9

In many eukaryotes, coordination of chromosome segregation with cell cleavage relies on the patterned interaction of specific microtubules with actin filaments through dedicated microtubule plus-end tracking proteins (+TIPs). However, how these+TIPs are spatially controlled is unclear. The yeast+TIP Kar9 drives one of the spindle aster microtubules along actin cables to align the mitotic spindle with the axis of cell division. Here, we report the crystal structure of Kar9's folded domain, revealing spectrin repeats reminiscent of the+TIPs MACF/ACF7/Shot and PRC1/Ase1. Point mutations abrogating spectrin-repeat-mediated dimerization of Kar9 reduced and randomized Kar9 distribution to microtubule tips, and impaired spindle positioning. Six Cdk1 sites surround the Kar9 dimerization interface. Their phosphomimetic substitution inhibited Kar9 dimerization, displaced Kar9 from microtubules, and affected its interaction with the myosin motor Myo2. Our results provide molecular-level understanding on how diverse cell types may regulate and pattern microtubule-actin interactions to orchestrate their divisions.

Human Cytomegalovirus Exploits TACC3 To Control Microtubule Dynamics and Late Stages of Infection.

While it is well established that microtubules (MTs) facilitate various stages of virus replication, how viruses actively control MT dynamics and functions remains less well understood. Recent work has begun to reveal how several viruses exploit End-Binding (EB) proteins and their associated microtubule plus-end tracking proteins (+TIPs), in particular to enable loading of viral particles onto MTs for retrograde transport during early stages of infection. Distinct from other viruses studied to date, at mid- to late stages of its unusually protracted replication cycle, human cytomegalovirus (HCMV) increases the expression of all three EB family members. This occurs coincident with the formation of a unique structure, termed the assembly compartment (AC), which serves as a Golgi-derived MT organizing center. Together, the AC and distinct EB proteins enable HCMV to increase the formation of dynamic and acetylated microtubule subsets to regulate distinct aspects of the viral replication cycle. Here, we reveal that HCMV also exploits EB-independent +TIP pathways by specifically increasing the expression of transforming acidic coiled coil protein 3 (TACC3) to recruit the MT polymerase, chTOG, from initial sites of MT nucleation in the AC out into the cytosol, thereby increasing dynamic MT growth. Preventing TACC3 increases or depleting chTOG impaired MT polymerization, resulting in defects in early versus late endosome organization in and around the AC as well as defects in viral trafficking and spread. Our findings provide the first example of a virus that actively exploits EB-independent +TIP pathways to regulate MT dynamics and control late stages of virus replication. IMPORTANCE Diverse viruses rely on host cell microtubule networks to transport viral particles within the dense cytoplasmic environment and to control the broader architecture of the cell to facilitate their replication. However, precisely how viruses regulate the dynamic behavior and function of microtubule filaments remains poorly defined. We recently showed that the assembly compartment (AC) formed by human cytomegalovirus (HCMV) acts as a Golgi-derived microtubule organizing center. Here, we show that at mid- to late stages of infection, HCMV increases the expression of transforming acidic coiled coil protein 3 (TACC3) to control the localization of the microtubule polymerase, chTOG. This, in turn, enables HCMV to generate dynamic microtubule subsets that organize endocytic vesicles in and around the AC and facilitate the transport of new viral particles released into the cytosol. Our findings reveal the first instance of viral targeting of TACC3 to control microtubule dynamics and virus spread.

Targeting the Microtubule EB1-CLASP2 Complex Modulates NaV1.5 at Intercalated Discs.

[Figure: see text].

Kinetochore-independent mechanisms of sister chromosome separation.

Although kinetochores normally play a key role in sister chromatid separation and segregation, chromosome fragments lacking kinetochores (acentrics) can in some cases separate and segregate successfully. In Drosophila neuroblasts, acentric chromosomes undergo delayed, but otherwise normal sister separation, revealing the existence of kinetochore- independent mechanisms driving sister chromosome separation. Bulk cohesin removal from the acentric is not delayed, suggesting factors other than cohesin are responsible for the delay in acentric sister separation. In contrast to intact kinetochore-bearing chromosomes, we discovered that acentrics align parallel as well as perpendicular to the mitotic spindle. In addition, sister acentrics undergo unconventional patterns of separation. For example, rather than the simultaneous separation of sisters, acentrics oriented parallel to the spindle often slide past one another toward opposing poles. To identify the mechanisms driving acentric separation, we screened 117 RNAi gene knockdowns for synthetic lethality with acentric chromosome fragments. In addition to well-established DNA repair and checkpoint mutants, this candidate screen identified synthetic lethality with X-chromosome-derived acentric fragments in knockdowns of Greatwall (cell cycle kinase), EB1 (microtubule plus-end tracking protein), and Map205 (microtubule-stabilizing protein). Additional image-based screening revealed that reductions in Topoisomerase II levels disrupted sister acentric separation. Intriguingly, live imaging revealed that knockdowns of EB1, Map205, and Greatwall preferentially disrupted the sliding mode of sister acentric separation. Based on our analysis of EB1 localization and knockdown phenotypes, we propose that in the absence of a kinetochore, microtubule plus-end dynamics provide the force to resolve DNA catenations required for sister separation.

Destabilization of Long Astral Microtubules via Cdk1-Dependent Removal of GTSE1 from Their Plus Ends Facilitates Prometaphase Spindle Orientation

The precise regulation of microtubule dynamics over time and space in dividing cells is critical for several mitotic mechanisms that ultimately enable cell proliferation, tissue organization, and development. Astral microtubules, which extend from the centrosome toward the cell cortex, must be present for the mitotic spindle to properly orient, as well as for the faithful execution of anaphase and cytokinesis. However, little is understood about how the dynamic properties of astral microtubules are regulated spatiotemporally, or the contribution of astral microtubule dynamics to spindle positioning. The mitotic regulator Cdk1-CyclinB promotes destabilization of centrosomal microtubules and increased microtubule dynamics as cells enter mitosis, but how Cdk1 activity modulates astral microtubule stability, and whether it impacts spindle positioning, is unknown. Here, we uncover a mechanism revealing that Cdk1 destabilizes astral microtubules in prometaphase and thereby influences spindle reorientation. Phosphorylation of the EB1-dependent microtubule plus-end tracking protein GTSE1 by Cdk1 in early mitosis abolishes its interaction with EB1 and recruitment to microtubule plus ends. Loss of Cdk1 activity, or mutation of phosphorylation sites in GTSE1, induces recruitment of GTSE1 to growing microtubule plus ends in mitosis. This decreases the catastrophe frequency of astral microtubules and causes an increase in the number of long astral microtubules reaching the cell cortex, which restrains the ability of cells to reorient spindles along the long cellular axis in early mitosis. Astral microtubules thus must not only be present but also dynamic to allow the spindle to reorient, a state assisted by selective destabilization of long astral microtubules via Cdk1.

A cilium-independent role for intraflagellar transport 88 in regulating angiogenesis

Endothelial cilia are microtubule-based hair-like protrusions in the lumen ofblood vessels that function as fluid mechanosensors to regulate vascular hemodynamics.However, the functions of endothelial cilia in vascular development remain controversial. In this study, depletion of several key proteins responsible for ciliogenesis allows us to identify a cilium-independent role for intraflagellartransport88 (IFT88) in mammalian angiogenesis. Disruption of primary cilia by heat shock does not affect the angiogenic process. However, depletion of IFT88 significantly inhibits angiogenesis both in vitro and in vivo. IFT88 mediates angiogenesis by regulating the migration, polarization, proliferation, and oriented division of vascular endothelial cells. Further mechanistic studies demonstrate that IFT88 interacts with γ-tubulin and microtubule plus-end tracking proteins and promotes microtubule stability. Our findings indicate that IFT88 regulates angiogenesis through its actions in microtubule-based cellular processes, independent of its role in ciliogenesis.

Redox-dependent regulation of end-binding protein 1 activity by glutathionylation.

Cytoskeletal proteins are susceptible to glutathionylation under oxidizing conditions, and oxidative damage has been implicated in several neurodegenerative diseases. End-binding protein 1 (EB1) is a master regulator of microtubule plus-end tracking proteins (+TIPs) and is critically involved in the control of microtubule dynamics and cellular processes. However, the impact of glutathionylation on EB1 functions remains unknown. Here we reveal that glutathionylation is important for controlling EB1 activity and protecting EB1 from irreversible oxidation. In vitro biochemical and cellular assays reveal that EB1 is glutathionylated. Diamide, a mild oxidizing reagent, reduces EB1 comet number and length in cells, indicating the impairment of microtubule dynamics. Three cysteine residues of EB1 are glutathionylated, with mutations of these three cysteines to serines attenuating microtubule dynamics but buffering diamide-induced decrease in microtubule dynamics. In addition, glutaredoxin 1 (Grx1) deglutathionylates EB1, and Grx1 depletion suppresses microtubule dynamics and leads to defects in cell division orientation and cell migration, suggesting a critical role of Grx1-mediated deglutathionylation in maintaining EB1 activity. Collectively, these data reveal that EB1 glutathionylation is an important protective mechanism for the regulation of microtubule dynamics and microtubule-based cellular activities.

Articles of Significant Interest in This Issue

Human immunodeficiency virus type 1 (HIV-1) exploits specialized microtubule (MT) plus-end tracking proteins (commonly known as +TIPs) to induce the formation of stable MTs soon after virus entry and to promote early stages of infection. Mitra et al. (e00404-20) found that distinct domains in cytoplasmic linker-associated protein 2 (CLASP2), a +TIP that captures cortical MT plus ends to enable filament stabilization, mediate interactions with incoming HIV-1 particles, and facilitate MT stabilization and trafficking of incoming cores. These findings identify a new +TIP that acts as a host cofactor to promote early stages of viral infection.

The +TIP Navigator-1 is an actin-microtubule crosslinker that regulates axonal growth cone motility.

Microtubule (MT) plus-end tracking proteins (+TIPs) are central players in the coordination between the MT and actin cytoskeletons in growth cones (GCs) during axon guidance. The +TIP Navigator-1 (NAV1) is expressed in the developing nervous system, yet its neuronal functions remain poorly elucidated. Here, we report that NAV1 controls the dynamics and motility of the axonal GCs of cortical neurons in an EB1-dependent manner and is required for axon turning toward a gradient of netrin-1. NAV1 accumulates in F-actin-rich domains of GCs and binds actin filaments in vitro. NAV1 can also bind MTs independently of EB1 in vitro and crosslinks nonpolymerizing MT plus ends to actin filaments in axonal GCs, preventing MT depolymerization in F-actin-rich areas. Together, our findings pinpoint NAV1 as a key player in the actin-MT crosstalk that promotes MT persistence at the GC periphery and regulates GC steering. Additionally, we present data assigning to NAV1 an important role in the radial migration of cortical projection neurons in vivo.