Cep57 regulates human centrosomes through multivalent interactions.

Human Cep57 is a coiled-coil scaffold at the pericentriolar matrix (PCM), controlling centriole duplication and centrosome maturation for faithful cell division. Genetic truncation mutations of Cep57 are associated with the mosaic-variegated aneuploidy (MVA) syndrome. During interphase, Cep57 forms a complex with Cep63 and Cep152, serving as regulators for centrosome maturation. However, the molecular interplay of Cep57 with these essential scaffolding proteins remains unclear. Here, we demonstrate that Cep57 undergoes liquid-liquid phase separation (LLPS) driven by three critical domains (NTD, CTD, and polybasic LMN). In vitro Cep57 condensates catalyze microtubule nucleation via the LMN motif-mediated tubulin concentration. In cells, the LMN motif is required for centrosomal microtubule aster formation. Moreover, Cep63 restricts Cep57 assembly, expansion, and microtubule polymerization activity. Overexpression of competitive constructs for multivalent interactions, including an MVA mutation, leads to excessive centrosome duplication. In Cep57-depleted cells, self-assembly mutants failed to rescue centriole disengagement and PCM disorganization. Thus, Cep57's multivalent interactions are pivotal for maintaining the accurate structural and functional integrity of human centrosomes.

Combating breast cancer progression through combination therapy with hypomethylating agent and glucocorticoid.

Breast cancer is the leading cause of cancer-related death in women. Among breast cancer types, triple-negative breast cancer (TNBC) accounts for 15% of all breast cancers with aggressive tumor behavior. By using bioinformatic approaches, we observed that the microRNA-708 promoter is highly methylated in breast carcinomas, and this methylation is linked to a poor prognosis. Moreover, microRNA-708 expression correlates with better clinical outcomes in TNBC patients. Combination treatment with the hypomethylating agent decitabine and synthetic glucocorticoid significantly increased the expression of microRNA-708, reactivated DNMT-suppressed pathways, and decreased the expression of multiple metastasis-promoting genes such as matrix metalloproteinases (MMPs) and IL-1ß, leading to the suppression of breast cancer cell proliferation, migration, and invasion, as well as reduced tumor growth and distant metastasis in the TNBC xenograft mouse model. Overall, our study reveals a therapeutic opportunity in which a combined regimen of decitabine with glucocorticoid may have therapeutic potential in treating TNBC patients.

MicroRNA-708 emerges as a potential candidate to target undruggable NRAS.

RAS, the most frequently mutated oncogene that drives tumorigenesis by promoting cell proliferation, survival, and motility, has been perceived as undruggable for the past three decades. However, intense research in the past has mainly focused on KRAS mutations, and targeted therapy for NRAS mutations remains an unmet medical need. NRAS mutation is frequently observed in several cancer types, including melanoma (15-20%), leukemia (10%), and occasionally other cancer types. Here, we report using miRNA-708, which targets the distinct 3' untranslated region (3'UTR) of NRAS, to develop miRNA-based precision medicine to treat NRAS mutation-driven cancers. We first confirmed that NRAS is a direct target of miRNA-708. Overexpression of miRNA-708 successfully reduced NRAS protein levels in melanoma, leukemia, and lung cancer cell lines with NRAS mutations, resulting in suppressed cell proliferation, anchorage-independent growth, and promotion of reactive oxygen species-induced apoptosis. Consistent with the functional data, the activities of NRAS-downstream effectors, the PI3K-AKT-mTOR or RAF-MEK-ERK signaling pathway, were impaired in miR-708 overexpressing cells. On the other hand, cell proliferation was not disturbed by miRNA-708 in cell lines carrying wild-type NRAS. Collectively, our data unveil the therapeutic potential of using miRNA-708 in NRAS mutation-driven cancers through direct depletion of constitutively active NRAS and thus inhibition of its downstream effectors to decelerate cancer progression. Harnessing the beneficial effects of miR-708 may therefore offer a potential avenue for small RNA-mediated precision medicine in cancer treatment.

Re-evaluation of the taxonomic confusion of the metallic-blue Pimpla Fabricius, 1804 (Hymenoptera: Ichneumonidae: Pimplinae) species in the Oriental region.

The metallic-blue body coloration is a distinct characteristic found in seven species of the cosmopolitan genus Pimpla Fabricius, 1804. In this study, we focus on two externally similar species, namely P. cyanator Morley, 1914 in India and P. alishanensis (Kusigemati, 1984) in Taiwan, along with the Indian P. indra Cameron, 1899, which are the only three species in the Oriental region exhibiting this trait. By examining the type series of these two taxa and acquiring additional materials from Taiwan, we discovered female morphological variations that resemble the form of the holotype of P. cyanator. Consequently, we propose that P. alishanensis should be synonymized with P. cyanator (syn. nov.). Furthermore, we provide a redescription of P. cyanator based on morphological characters and molecular COI sequences. We also discuss its revised distribution, the diagnostic features distinguishing P. cyanator from other metallic-blue Pimpla species worldwide, the factors influencing the color change in specimens, and some potential misidentifications that have occurred in the past.

Online citizen sciences reveal natural enemies and new occurrence data of Meteorusstellatus Fujie, Shimizu & Maeto, 2021 (Hymenoptera, Braconidae, Euphorinae).

Background: Citizen science is a research approach that involves collaboration between professional scientists and non-professional volunteers. The utilisation of recent online citizen-science platforms (e.g. social networking services) has greatly revolutionised the accessibility of biodiversity data by providing opportunities for connecting professional and citizen scientists worldwide. Meteorusstellatus Fujie, Shimizu & Maeto, 2021 (Hymenoptera, Braconidae, Euphorinae) has been recorded from the Oriental Islands of Japan and known to be a gregarious endoparasitoid of two macro-sized sphingid moths of Macroglossum, Ma.passalus (Drury) and Ma.pyrrhosticta Butler. It constructs characteristic star-shaped communal cocoons, suspended by a long cable. Although M.stellatus has been reported only from the Oriental Islands of Japan, the authors recognise its occurrence and ecological data from Taiwan and the Palaearctic Island of Japan through posts on online citizen-science groups about Taiwanese Insects on Facebook and an article on a Japanese citizen-scientist's website. New information: Through collaboration between professional and citizen scientists via social media (Facebook groups) and websites, the following new biodiversity and ecological data associated with M.stellatus are provided: Meteorusstellatus is recorded for the first time from Taiwan and the Palaearctic Region (Yakushima Is., Japan).Cechetraminor (Butler, 1875), Hippotioncelerio (Linnaeus, 1758) and Macroglossumsitiene (Walker, 1856) (Lepidoptera, Sphingidae) are recorded for the first time as hosts of M.stellatus and two of which (C.minor and H.celerio) represent the first genus-level host records for M.stellatus.Mesochorus sp. (Hymenoptera, Ichneumonidae), indeterminate species of Pteromalidae and Trichogrammatidae (Hymenoptera), are recognised as hyperparasitoid wasps of M.stellatus.Parapolybiavaria (Fabricius, 1787) (Hymenoptera, Vespidae) is reported as a predator of pendulous communal cocoons of M.stellatus. The nature of suspended large-sized communal cocoons of M.stellatus and the importance and limitations of digital occurrence data and online citizen science are briefly discussed.

DNA Sequencing from Subcritical Concentration of Cell-Free DNA Extracted from Electrowetting-on-Dielectric Platform.

Electro-Wetting-On-Dielectric (EWOD) based digital operations have demonstrated outstanding potential in actuating and manipulating liquid droplets. Here, we adapted the EWOD for extracting femtogram quantities of cell-free DNA (cf-DNA) from 1 µL of KSOM mouse embryo culture medium. Our group extracted the femtogram quantity of cf-DNA from 1 µL of mouse embryo culture medium in our previous work. Here, we initially explain a modification from our previous extraction protocol, which improves the extraction percentage to 36.74%. Though the modified extraction protocol improves the extraction percentage from our previously reported work, the quantity is still in the femtogram range. The cf-DNA in femtogram quantity is in subcritical/subthreshold concentration for any further analysis, such as sequencing. To the best of our knowledge, we need a minimum of picogram/nanogram DNA quantities for further analysis. We demonstrated a ground-breaking mechanism of this subcritical concentration of cf-DNA amplification to the nanogram range and performed DNA sequencing. Basic Local Alignment Search Tool (BLAST) is used as a sequence similarity search program to confirm the identity percentage between query and subject. More than 97% of nucleotide identities between query and subject sequences have been obtained from the sequencing result. Hence, we can use the methodology to amplify the subcritical concentration of extracted DNA for further analytics. Moreover, as we extract the cf-DNA from the embryo culture medium, the natural growth of the embryo has not been disrupted. This entire mechanism will pave a new path towards the lab-on-a-chip (LOC) concept.

Lack of social touch alters anxiety-like and social behaviors in male mice.

The importance of social interactions has been reported in a variety of animal species. In human and rodent models, social isolation is known to alter social behaviors and change anxiety or depression levels. During the coronavirus pandemic, although people could communicate with each other through other sensory cues, social touch was mostly prohibited under different levels of physical distancing policies. These social restrictions inspired us to explore the necessity of physical contact, which has rarely been investigated in previous studies on mouse social interactions. We first conducted a long-term observation to show that pair-housed mice in a standard laboratory cage spent nearly half the day in direct physical contact with each other. Furthermore, we designed a split-housing condition to demonstrate that even with free access to visual, auditory, and olfactory social signals, the lack of social touch significantly increased anxiety-like behaviors and changed social behaviors. There were correspondingly higher levels of the pro-inflammatory cytokine interleukin-6 in the hippocampus in mice with no access to physical contact. Our study demonstrated the necessity of social touch for the maintenance of mental health in mice and could have important implications for human social interactions.

BARD1 is an ATPase activating protein for OLA1.

OLA1 is a P-loop ATPase, implicated in centrosome duplication through the interactions with tumor suppressors BRCA1 and BARD1. Disruption of the interaction of OLA1 with BARD1 results in centrosome amplification. However, the molecular interplay and mechanism of the OLA1-BARD1 complex remain elusive. Here, we use a battery of biophysical, biochemical, and structural analyses to elucidate the molecular basis of the OLA1-BARD1 interaction. Our structural and enzyme kinetics analyses show this nucleotide-dependent interaction enhances the ATPase activity of OLA1 by increasing the turnover number (kcat). Unlike canonical GTPase activating proteins that act directly on the catalytic G domain, the BARD1 BRCT domain binds to the OLA1 TGS domain via a highly conserved BUDR motif. A cancer related mutation V695L on BARD1 is known to associate with centrosome abnormality. The V695L mutation reduces the BARD1 BRCT-mediated activation of OLA1. Crystallographic snapshot of the BRCT V695L mutant at 1.88 Å reveals this mutation perturbs the OLA1 binding site, resulting in reduced interaction. Altogether, our findings suggest the BARD1 BRCT domain serves as an ATPase activating protein to control OLA1 allosterically.

Monocytes secrete CXCL7 to promote breast cancer progression.

Certain immune cells and inflammatory cytokines are essential components in the tumor microenvironment to promote breast cancer progression. To identify key immune players in the tumor microenvironment, we applied highly invasive MDA-MB-231 breast cancer cell lines to co-culture with human monocyte THP-1 cells and identified CXCL7 by cytokine array as one of the increasingly secreted cytokines by THP-1 cells. Further investigations indicated that upon co-culturing, breast cancer cells secreted CSF1 to induce expression and release of CXCL7 from monocytes, which in turn acted on cancer cells to promote FAK activation, MMP13 expression, migration, and invasion. In a xenograft mouse model, administration of CXCL7 antibodies significantly reduced abundance of M2 macrophages in tumor microenvironment, as well as decreased tumor growth and distant metastasis. Clinical investigation further suggested that high CXCL7 expression is correlated with breast cancer progression and poor overall survival of patients. Overall, our study unveils an important immune cytokine, CXCL7, which is secreted by tumor infiltrating monocytes, to stimulate cancer cell migration, invasion, and metastasis, contributing to the promotion of breast cancer progression.

The Hidden Role of Hydrogen Sulfide Metabolism in Cancer.

Hydrogen Sulfide (H2S), an endogenously produced gasotransmitter, is involved in various important physiological and disease conditions, including vasodilation, stimulation of cellular bioenergetics, anti-inflammation, and pro-angiogenesis. In cancer, aberrant up-regulation of H2S-producing enzymes is frequently observed in different cancer types. The recognition that tumor-derived H2S plays various roles during cancer development reveals opportunities to target H2S-mediated signaling pathways in cancer therapy. In this review, we will focus on the mechanism of H2S-mediated protein persulfidation and the detailed information about the dysregulation of H2S-producing enzymes and metabolism in different cancer types. We will also provide an update on mechanisms of H2S-mediated cancer progression and summarize current options to modulate H2S production for cancer therapy.

Effects of synthetic glucocorticoids on breast cancer progression.

Glucocorticoids (GCs) are widely prescribed as adjuvant therapy for breast cancer patients. Unlike other steroid hormone receptors, the GC receptor is not considered an oncogene. Research in the past few years has revealed the complexity of GC-mediated signaling, but it remains puzzling whether GCs promote or inhibit tumor progression in different cancer types. Here we evaluated the potential of using a synthetic GC, dexamethasone (DEX), in the treatment of breast cancer. We found that the administration of low-dose DEX suppressed tumor growth and distant metastasis in the MCF-7 and MDA-MB-231 xenograft mouse model, whereas treatment with high-dose DEX enhanced tumor growth and metastasis, respectively. Treatment of breast cancer cells with DEX inhibited cell adhesion, migration, and invasion in a dose-dependent manner. The DEX-mediated inhibition of cell adhesion, migration, and invasion is partly through induction of microRNA-708 and subsequent Rap1B-mediated signaling in MDA-MB-231 cells. On the other hand, in MCF-7 cells, DEX-suppressed cell migration is independent from microRNA-708 mediated signaling. Overall, our data reveal that DEX acts as a double-edged sword during breast-cancer progression and metastasis: Lower concentrations inhibit breast cancer tumor growth and metastasis, whereas higher concentrations may play an undesired role to promote breast cancer progression.

Extraction of Cell-free Dna from An Embryo-culture Medium Using Micro-scale Bio-reagents on Ewod.

As scientific and technical knowledge advances, research on biomedical micro-electromechanical systems (bio-MEMS) is also developing towards lab-on-a-chip (LOC) devices. A digital microfluidic (DMF) system specialized for an electrowetting- on-dielectric (EWOD) mechanism is a promising technique for such point-of-care systems. EWOD microfluidic biochemical analytical systems provide applications over a broad range in the lab-on-a-chip field. In this report, we treated extraction of cell-free DNA (cf-DNA) at a small concentration from a mouse embryo culture medium (2.5 days & 3.5 days) with electro-wetting on a dielectric (EWOD) platform using bio-reagents of micro-scale quantity. For such extraction, we modified a conventional method of genomic-DNA (g-DNA) extraction using magnetic beads (MB). To prove that extraction of cf-DNA with EWOD was accomplished, as trials we extracted designed-DNA (obtained from Chang Gung Memorial Hospital (CGMH), Taiwan which shows properties similar to that of cf-DNA). Using that designed DNA, extraction with both conventional and EWOD methods has been performed; the mean percentage of extraction with both methods was calculated for a comparison. From the cycle threshold (Ct) results with a quantitative polymerase chain reaction (q-PCR), the mean extraction percentages were obtained as 14.8 percent according to the conventional method and 23 percent with EWOD. These results show that DNA extraction with EWOD appears promising. The EWOD extraction involved voltage 100 V and frequency 2 kHz. From this analysis, we generated a protocol for an improved extraction percentage on a EWOD chip and performed cf-DNA extraction from an embryo-culture medium (KSOM medium) at 3.5 and 2.5 days. The mean weight obtained for EWOD-extracted cf-DNA is 0.33 fg from the 3.5-day sample and 31.95 fg from the 2.5-day sample. All these results will pave a new path towards a renowned lab-on-a-chip concept.

Dysregulation of cystathionine γ-lyase promotes prostate cancer progression and metastasis.

Hydrogen sulfide (H2 S), an endogenous signaling gaseous molecule, is involved in various physiological activities, including vessel relaxation, regulation of cellular bioenergetics, inflammation, and angiogenesis. By using xenograft orthotopic implantation of prostate cancer PC3 cells and subsequently comparing bone metastatic with primary tumor-derived cancer cells, we find that H2 S-producing enzyme cystathionine γ-lyase (CTH) is upregulated in bone-metastatic PC3 cells. Clinical data further reveal that the expression of CTH is elevated in late-stage prostate cancer patients, and higher CTH expression correlates with poor survival from The Cancer Genome Atlas (TCGA) prostate cancer RNA-seq datasets. CTH promotes NF-κB nuclear translocation through H2 S-mediated sulfhydration on cysteine-38 of the NF-κB p65 subunit, resulting in increased IL-1ß expression and H2 S-induced cell invasion. Knockdown of CTH in PC3 cells results in the suppression of tumor growth and distant metastasis, while overexpression of CTH in DU145 cells promotes primary tumor growth and lymph node metastasis in the orthotopic implanted xenograft mouse model. Together, our findings provide evidence that CTH generated H2 S promotes prostate cancer progression and metastasis through IL-1ß/NF-κB signaling pathways.

Wild-type p53 upregulates an early onset breast cancer-associated gene GAS7 to suppress metastasis via GAS7-CYFIP1-mediated signaling pathway.

The early onset breast cancer patients (age ≤ 40) often display higher incidence of axillary lymph node metastasis, and poorer five-year survival than the late-onset patients. To identify the genes and molecules associated with poor prognosis of early onset breast cancer, we examined gene expression profiles from paired breast normal/tumor tissues, and coupled with Gene Ontology and public data base analysis. Our data showed that the expression of GAS7b gene was lower in the early onset breast cancer patients as compared to the elder patients. We found that GAS7 was associated with CYFIP1 and WAVE2 complex to suppress breast cancer metastasis via blocking CYFIP1 and Rac1 protein interaction, actin polymerization, and ß1-integrin/FAK/Src signaling. We further demonstrated that p53 directly regulated GAS7 gene expression, which was inversely correlated with p53 mutations in breast cancer specimens. Our study uncover a novel regulatory mechanism of p53 in early onset breast cancer progression through GAS7-CYFIP1-mediated signaling pathways.

Low-dose glucocorticoids suppresses ovarian tumor growth and metastasis in an immunocompetent syngeneic mouse model.

Ovarian cancer has the highest mortality rate among gynecologic malignancies. Despite chemotherapy and surgical debulking options, ovarian cancer recurs and disseminates frequently with a poor prognosis. We previously reported a novel role of glucocorticoids (GCs) in metastatic ovarian cancer by upregulating microRNA-708. In this study, we used an immunocompetent syngeneic mouse model and further evaluated the effect and optimal dosages of GCs in treating metastatic ovarian cancer. The treatment of C57BL/6-derived ovarian cancer ID-8 cells with a synthetic GC, dexamethasone (DEX), induced the expression of microRNA-708, leading to decreased cell migration and invasion through targeting Rap1B. Administration of DEX at a low dose, as low as 5 µg/kg body weight, inhibited the primary tumor size and abdominal metastasis in mice bearing ID-8 cell-derived ovarian tumors. In the treated primary tumors, microRNA-708 was upregulated, whereas some proinflammatory cytokines, namely interleukin (IL)-1ß and IL-18, were downregulated. The number of tumor-associated macrophages (TAMs) and myeloid-derived suppressor cells (MDSCs) in the tumor microenvironment were reduced. Overall, our study shows that low-dose GCs can suppress ovarian cancer progression and metastasis likely through not only the upregulation of the metastasis suppressor microRNA-708, but also the modulation of TAMs and MDSCs in the tumor microenvironment.

New dimension of glucocorticoids in cancer treatment.

Glucocorticoids have been used in clinical oncology for over half a century. The clinical applications of glucocorticoids in oncology are mainly dependent on their pro-apoptotic action to treat lymphoproliferative disorders, and also on alleviating side effects induced by chemotherapy or radiotherapy in non-hematologic cancer types. Researches in the past few years have begun to unveil the profound complexity of glucocorticoids signaling and have contributed remarkably on therapeutic strategies. However, it remains striking and puzzling how glucocorticoids use different mechanisms in different cancer types and different targets to promote or inhibit tumor progression. In this review, we provide an update on glucocorticoids and its receptor, GR-mediated signaling and highlight some of the latest findings on the actions of glucocorticoids signaling during tumor progression and metastasis.

Glucocorticoids mediate induction of microRNA-708 to suppress ovarian cancer metastasis through targeting Rap1B.

Glucocorticoids are widely used in conjunction with chemotherapy for ovarian cancer to prevent hypersensitivity reactions. Here we reveal a novel role for glucocorticoids in the inhibition of ovarian cancer metastasis. Glucocorticoid treatments induce the expression of miR-708, leading to the suppression of Rap1B, which result in the reduction of integrin-mediated focal adhesion formation, inhibition of ovarian cancer cell migration/invasion and impaired abdominal metastasis in an orthotopic xenograft mouse model. Restoring Rap1B expression reverts glucocorticoid-miR-708 cascade-mediated suppression of ovarian cancer cell invasion and metastasis. Clinically, low miR-708 and high Rap1B are found in late-state ovarian tumours, as compared with normal, and patients with high miR-708 show significantly better survival. Overall, our findings reveal an opportunity for glucocorticoids and their downstream mediators, miR-708 or Rap1B, as therapeutic modalities against metastatic ovarian epithelial cancer.

Vav3-rac1 signaling regulates prostate cancer metastasis with elevated Vav3 expression correlating with prostate cancer progression and posttreatment recurrence.

Prostate cancer remains the second leading cause of cancer death in men in the Western world. Yet current therapies do not significantly improve the long-term survival of patients with distant metastasis. In this study, we investigated the role of the guanine nucleotide exchange factor Vav3 in prostate cancer progression and metastasis and found that Vav3 expression correlated positively with prostate cancer cell migration and invasion. Stimulation of the receptor tyrosine kinase EphA2 by ephrinA1 resulted in recruitment and tyrosine phosphorylation of Vav3, leading to Rac1 activation as well as increased migration and invasion in vitro. Reduction of Vav3 resulted in fewer para-aortic lymph nodes and bone metastasis in vivo. Clinically, expression of Vav3 and EphA2 was elevated in late-stage and metastatic prostate cancers. Among patients with stage IIB or earlier prostate cancer, higher Vav3 expression correlated with lower cumulative biochemical failure-free survival, suggesting that Vav3 may represent a prognostic marker for posttreatment recurrence of prostate cancer. Together, our findings provide evidence that the Vav3-mediated signaling pathway may serve as a therapeutic target for prostate cancer metastasis.

Rapamycin by itself and additively in combination with carboplatin inhibits the growth of ovarian cancer cells.

OBJECTIVE: The current standard treatment for ovarian carcinoma, consisting of surgery followed by chemotherapy with carboplatin and paclitaxel, is fraught with a high rate of recurrences. We hypothesized that targeted inhibition of specific signaling pathways in combination with conventional drugs may increase chemotherapeutic efficacy. METHODS: We analyzed the expression and activation profiles of various signaling pathways in nine established ovarian cancer cell lines (CAOV-3, ES2, PA-1, SKOV-3, NIHOVCAR3, OV90, TOV112D, A1847, A2780) and 24 freshly procured human ovarian tumors. The PI3 kinase pathway component Akt was frequently overexpressed and/or activated in tumor cells. The effect of several PI3K pathway inhibitors (rapamycin, LY294002, SH-6) and rapamycin in combination with carboplatin on various tumor cell growth characteristics was tested in cell lines and fresh tumor-derived transient monolayer and organ cultures. RESULTS: Rapamycin by itself and additively with carboplatin inhibited the growth and invasion, and increased the sensitivity to anoikis of most of the ovarian cancer cell lines and fresh tumors. The additive inhibitory effect may be due to enhanced apoptosis as demonstrated by Poly-ADP-Ribose Polymerase (PARP) cleavage and Annexin V staining in cells treated with both rapamycin and carboplatin. CONCLUSIONS: Rapamycin in combination with standard chemotherapeutic agents may improve the efficiency of ovarian cancer treatment.

Ephrin-B2-induced cleavage of EphB2 receptor is mediated by matrix metalloproteinases to trigger cell repulsion.

EphB receptors provide crucial adhesive and repulsive signals during cell migration and axon guidance, but it is unclear how they switch between these opposing responses. Here we provide evidence of an important role for matrix metalloproteinases (MMPs) in repulsive EphB2 signaling. We found that EphB2 is cleaved by MMPs both in vitro and in vivo, and that this cleavage is induced by interaction with its ligand ephrin-B2. Our findings demonstrate that MMP-2/MMP-9-specific inhibition or cleavage-resistant mutations in the ectodomain of EphB2 can prevent EphB2-mediated cell-cell repulsion in HEK293 cells, and block ephrin-B1-induced growth cone withdrawal in cultured hippocampal neurons. Transient expression of wtEphB2, but not noncleavable EphB2-4/5 mutant, restored ephrin-B1-induced growth cone collapse and withdrawal in EphB-deficient neurons. The inhibition of EphB2 cleavage also had potent regulatory effects on EphB2 activity. This study provides the first evidence that MMP-mediated cleavage of EphB2 is induced by receptor-ligand interactions at the cell surface and that this event triggers cell-repulsive responses.