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The multidomain protein Trio regulates among others neuronal outgrowth and axonal guidance in vertebrates and invertebrates. Trio contains two Dbl-homology/pleck- strin homology (DH/PH) tandem domains that activate several RhoGTPases. Here, we present the x-ray structure of the N-terminal DH/PH, hereafter TrioN, refined to 1.7-A resolution. We show that the relative orientations of the DH and PH domains of TrioN and free Dbs are similar. However, this relative orientation is dissimilar to Dbs in the Dbs/Cdc42 structure. In vitro nucleotide exchange ex- periments catalyzed by TrioN show that RhoG is ∼3× more efficiently exchanged than Rac and support the conclusion that RhoG is likely the downstream target of TrioN. Residues 54 and 69, which are not conserved between the two GTPases, are responsible for this specificity. Dot-blot assay reveals that the TrioN-PH domain does not detectably bind phosphatidylinositol 3,4-bisphosphate, PtdIns(3,4)P2, or other phospholipids. This finding is supported by our three-dimensional structure and affinity binding experiments. Interestingly, the presence of RhoG but not Rac or a C-terminal-truncated RhoG mutant allows TrioN to bind PtdIns(3,4)P2 with a micromolar affinity constant. We conclude the variable C-terminal basic tail of RhoG specifically assists the recruitment of the TrioN-PH domain to specific membrane-bound phospholipids. Our data suggest a role for the phosphoinositide 3-kinase, PI 3-kinase, in modulating the Trio/RhoG signaling pathway. [ABSTRACT FROM AUTHOR]

A recent study shows that the protumorigenic guanine nucleotide exchange factor (GEF) Vav1 functions as a tumor suppressor in T cell acute lymphoblastic leukemia (T-ALL) through its ability to complex with the Cbl-b ubiquitin ligase and the intracellular domain of Notch1 (ICN1) and to promote ICN1 degradation. Vav1can act as a double-edged sword in tumorigenesis. [ABSTRACT FROM AUTHOR]

Alzheimer's disease, the most common cause of dementia, is a chronic degenerative disease with typical pathological features of extracellular senile plaques and intracellular neurofibrillary tangles and a significant decrease in the density of neuronal dendritic spines. Cdc42 is a member of the small G protein family that plays an important role in regulating synaptic plasticity and is regulated by Cdc42GAP, which switches Cdc42 from active GTP-bound to inactive GDP-bound states regulating downstream pathways via effector proteins. However, few studies have focused on Cdc42 in the progression of Alzheimer's disease. In a heterozygous Cdc42GAP mouse model that exhibited elevated Cdc42-GTPase activity accompanied by increased Cdc42-PAK1-cofilin signalling, we found impairments in cognitive behaviours, neuron senescence, synaptic loss with depolymerization of F-actin and the pathological phenotypes of Alzheimer's disease, including phosphorylated tau (p-T231, AT8), along with increased soluble and insoluble Aβ1–42 and Aβ1–40, which are consistent with typical Alzheimer's disease mice. Interestingly, these impairments increased significantly with age. Furthermore, the results of quantitative phosphoproteomic analysis of the hippocampus of 11-month-old GAP mice suggested that Cdc42GAP deficiency induces and accelerates Alzheimer's disease-like phenotypes through activation of GSK-3β by dephosphorylation at Ser9, Ser389 and/or phosphorylation at Tyr216. In addition, overexpression of dominant-negative Cdc42 in the primary hippocampal and cortical neurons of heterozygous Cdc42GAP mice reversed synaptic loss and tau hyperphosphorylation. Importantly, the Cdc42 signalling pathway, Aβ1–42, Aβ1–40 and GSK-3β activity were increased in the cortical sections of Alzheimer's disease patients compared with those in healthy controls. Together, these data indicated that Cdc42GAP is involved in regulating Alzheimer's disease-like phenotypes such as cognitive deficits, dendritic spine loss, phosphorylated tau (p-T231, AT8) and increased soluble and insoluble Aβ1–42 and Aβ1–40, possibly through the activation of GSK-3β, and these impairments increased significantly with age. Thus, we provide the first evidence that Cdc42 is involved in the progression of Alzheimer's disease-like phenotypes, which may provide new targets for Alzheimer's disease treatment. [ABSTRACT FROM AUTHOR]

Background In our previous study, Activin B induced actin stress fiber formation and cell migration in Bone marrow-derived mesenchymal stem cells (BMSCs) in vitro. However, the underlying molecular mechanisms are not well studied. RhoA is recognized to play a critical role in the regulation of actomyosin cytoskeletal organization and cell migration. Methods Pull-down assay was performed to investigate the activity of RhoA. The dominant-negative mutants of RhoA (RhoA(N19)) was used to determine whether RhoA has a role in Activin B-induced cytoskeleton organization and cell migration in BMSCs. Cytoskeleton organization was examined by fluorescence Rhodamine-phalloidin staining, and cell migration by transwell and cell scratching assay. Western blot was carried out to investigate downstream signaling cascade of RhoA. Inhibitor and siRNAs were used to detect the role of downstream signaling in stress fiber formation and/or cell migration. Results RhoA was activated by Activin B in BMSCs. RhoA(N19) blocked Activin B-induced stress fiber formation and cell migration. ROCK inhibitor blocked Activin B-induced stress fiber formation but enhanced BMSCs migration. Activin B induced phosphorylation of LIMK2 and Cofilin, which was abolished by ROCK inhibition. Both of siRNA LIMK2 and siRNA Cofilin inhibited Activin B-induced stress fiber formation. Conclusions RhoA regulates Activin B-induced stress fiber formation and migration of BMSCs. A RhoA-ROCK-LIMK2-Cofilin signaling node exists and regulates actin stress fiber formation. RhoA regulates Activin B-induced cell migration independent of ROCK. General significance Better understanding of the molecular mechanisms of BMSCs migration will help optimize therapeutic strategy to target BMSCs at injured tissues. [ABSTRACT FROM AUTHOR]

Substantial spine loss in striatal medium spiny neurons (MSNs) and abnormal behaviors are common features of Parkinson's disease (PD). The caudate putamen (CPu) mainly contains MSNs expressing dopamine D1 receptor (dMSNs) and dopamine D2 receptor (iMSNs) exerting critical effects on motor and cognition behavior. However, the molecular mechanisms contributing to spine loss and abnormal behaviors in dMSNs and iMSNs under parkinsonian state remain unknown. In the present study, we revealed that Cell division control protein 42 (Cdc42) signaling was significantly decreased in the caudate putamen (CPu) in parkinsonian mice. In addition, overexpression of constitutively active Cdc42 in the CPu reversed spine abnormalities and improved the behavior deficits in parkinsonian mice. Utilizing conditional dopamine D1 receptor (D1R) or D2 receptor (D2R) knockout mice, we found that such a decrease under parkinsonian state was further reduced by conditional knockout of the D2R but not D1R. Moreover, the thin spine loss in iMSNs and deficits in motor coordination and cognition induced by conditional knockout of D2R were reversed by overexpression of constitutively active Cdc42 in the CPu. Additionally, conditional knockout of Cdc42 from D2R‐positive neurons in the CPu was sufficient to induce spine and behavior deficits similar to those observed in parkinsonian mice. Overall, our results indicate that impaired Cdc42 signaling regulated by D2R plays an important role in spine loss and behavioral deficits in PD. [ABSTRACT FROM AUTHOR]

Extracellular matrix (ECM) actively participates in normal cell regulation and in the process of tumor progression. The Rho GTPase Cdc42 has been shown to regulate cell-ECM interaction in conventional two-dimensional culture conditions by using dominant mutants of Cdc42 in immortalized cell lines that may introduce nonspecific effects. Here, we employ three-dimensional culture systems for conditional gene targeted primary mouse embryonic fibroblasts that better simulate the reciprocal and adaptive interactions between cells and surrounding matrix to define the role of Cdc42 signaling pathways in ECM organization. Cdc42 deficiency leads to a defect in global cell-matrix interactions reflected by a decrease in collagen gel contraction. The defect is associated with an altered cell-matrix interaction that is evident by morphologic changes and reduced focal adhesion complex formation. The matrix defect is also associated with a reduction in synthesis and activation of matrix metalloproteinase 9 (MMP9) and altered fibronectin deposition patterning. A Cdc42 mutant rescue experiment found that downstream of Cdc42, p21-activated kinase (PAK), but not Par6 or WASP, may be involved in regulating collagen gel contraction and fibronectin organization. Thus, in addition to the previously implicated roles in intracellular regulation of actin organization, proliferation, and vesicle trafficking, Cdc42 is essential in ECM remodeling in three dimensions. [ABSTRACT FROM AUTHOR]

Methamphetamine (METH) is a highly addictive psychostimulant that strongly activates dopamine receptor signaling in the nucleus accumbens (NAc). However, how dopamine D 1 and D 2 receptors (D 1 Rs and D 2 Rs, respectively) as well as downstream signaling pathways, such as those involving Rac1 and Cdc42, modulate METH-induced behavioral and structural plasticity is largely unknown. Using NAc conditional D 1 R and D 2 R deletion mice, Rac1 and Cdc42 mutant viruses, and a series of behavioral and morphological methods, we assessed the effects of D 1 Rs and D 2 Rs on Rac1 and Cdc42 in modulating METH-induced behavioral and structural plasticity in the NAc. D 1 Rs and D 2 Rs in the NAc consistently regulated METH-induced conditioned place preference, locomotor activation, and dendritic and spine remodeling of medium spiny neurons but differentially regulated METH withdrawal–induced spatial learning and memory impairment and anxiety. Interestingly, Rac1 and Cdc42 signaling were oppositely modulated by METH, and suppression of Rac1 signaling and activation of Cdc42 signaling were crucial to METH-induced conditioned place preference and structural plasticity but not to locomotor activation. D 1 Rs activated Rac1 and Cdc42 signaling, while D 2 Rs inhibited Rac1 signaling but activated Cdc42 signaling to mediate METH-induced conditioned place preference and structural plasticity but not locomotor activation. In addition, NAc D 1 R deletion aggravated METH withdrawal–induced spatial learning and memory impairment by suppressing Rac1 signaling but not Cdc42 signaling, while NAc D 2 R deletion aggravated METH withdrawal–induced anxiety without affecting Rac1 or Cdc42 signaling. D 1 Rs and D 2 Rs differentially regulate Rac1 and Cdc42 signaling to modulate METH-induced behavioral plasticity and the structural remodeling of medium spiny neurons in the NAc. [ABSTRACT FROM AUTHOR]

The purpose of this paper is to analytically explore that an electromagnetic ultrasonic transducer (EMAT) can be used to do the activity determination of crack defects. Acoustic emission (AE) testing has been done through the innovative use of EMAT, and the activity of defects can be determined based on the Kaiser effect. In consideration of the Lorentz force and magnetostrictive force, the magnetism, force, and vibration displacement of aluminum sheet and steel sheet are analyzed through the finite-element method and fast Fourier transformation. The db3 wavelet is used to obtain the rule of signal change with excitation current. The experimental system of aluminum sheet and steel sheet was established for verification. This method effectively expands the AE theory based on global loading and also gives full play and expands the defect detection capability of EMAT equipment. [ABSTRACT FROM AUTHOR]

Summary: Background: Asthma is an allergic airway inflammation‐driven disease that affects more than 300 million people world‐wide. Targeted therapies for asthma are largely lacking. Although asthma symptoms can be prevented from worsening, asthma development cannot be prevented. Cdc42 GTPase has been shown to regulate actin cytoskeleton, cell proliferation and survival. Objectives: To investigate the role and targeting of Cdc42 in Th2 cell differentiation and Th2‐mediated allergic airway inflammation. Methods: Post‐thymic Cdc42‐deficient mice were generated by crossing Cdc42flox/flox mice with dLckicre transgenic mice in which Cre expression is driven by distal Lck promoter. Effects of post‐thymic Cdc42 deletion and pharmacological targeting Cdc42 on Th2 cell differentiation were evaluated in vitro under Th2‐polarized culture conditions. Effects of post‐thymic Cdc42 deletion and pharmacological targeting Cdc42 on allergic airway inflammation were evaluated in ovalbumin‐ and/or house dust mite‐induced mouse models of asthma. Results: Post‐thymic deletion of Cdc42 led to reduced peripheral CD8+ T cells and attenuated Th2 cell differentiation, with no effect on closely related Th1, Th17 and induced regulatory T (iTreg) cells. Post‐thymic Cdc42 deficiency ameliorated allergic airway inflammation. The selective inhibition of Th2 cell differentiation by post‐thymic deletion of Cdc42 was recapitulated by pharmacological targeting of Cdc42 with CASIN, a Cdc42 activity‐specific chemical inhibitor. CASIN also alleviated allergic airway inflammation. CASIN‐treated Cdc42‐deficient mice showed comparable allergic airway inflammation to vehicle‐treated Cdc42‐deficient mice, indicative of negligible off‐target effect of CASIN. CASIN had no effect on established allergic airway inflammation. Conclusion and Clinical Relevance: Cdc42 is required for Th2 cell differentiation and allergic airway inflammation, and rational targeting Cdc42 may serve as a preventive but not therapeutic approach for asthma control. [ABSTRACT FROM AUTHOR]

Background: Endothelial cell (EC) regeneration is essential for inflammation resolution and vascular integrity recovery after inflammatory vascular injury. Cdc42 is a central regulator of cell survival and vessel formation in EC development. However, it is unknown that whether Cdc42 could be a regulating role of EC repair following the inflammatory injury in the lung. The study sought to test the hypothesis that Cdc42 is required for endothelial regeneration and vascular integrity recovery after LPS-induced inflammatory injury.Methods and Results: The role of Cdc42 for the regulation of pulmonary vascular endothelial repair was tested in vitro and in vivo. In LPS-induced acute lung injury (ALI) mouse models, knockout of the Cdc42 gene in ECs increased inflammatory cell infiltration and pulmonary vascular leakage and inhibited vascular EC proliferation, which eventually resulted in more severe inflammatory lung injury. In addition, siRNA-mediated knockdown of Cdc42 protein on ECs disrupted cell proliferation and migration and tube formation, which are necessary processes for recovery after inflammatory vascular injury, resulting in inflammatory vascular injury recovery defects.Conclusion: We found that Cdc42 deficiency impairs EC function and regeneration, which are crucial in the post-inflammatory vascular injury repair process. These findings indicate that Cdc42 is a potential target for novel treatments designed to facilitate endothelial regeneration and vascular repair in inflammatory pulmonary vascular diseases, such as ALI/ARDS. [ABSTRACT FROM AUTHOR]

Asthma is a chronic airway inflammation in which Th2 and Th17 cells play critical roles in its pathogenesis. We have reported that atypical protein kinase (PKC) λ/ι is a new regulator for Th2 differentiation and function. However, the role of PKCλ/ι for Th17 cells remains elusive. In this study, we explored the effect of PKCλ/ι on Th17 cells in the context of ex vivo cell culture systems and an in vivo murine model of allergic airway inflammation with the use of activated T cell-specific conditional PKCλ/ι-deficient mice. Our findings indicate that PKCλ/ι regulates Th17 cells. The secretion of Th17 effector cytokines, including IL-17, IL-21 and IL-22, were inhibited from PKCλ/ι-deficient T cells under non-skewing or Th17-skewing culture conditions. Moreover, the impaired Th17 differentiation and function by the PKCλ/ι-deficiency was associated with the downregulation of Stat3 and Rorγt, key Th17 transcription factors. We developed a model of Th17 and neutrophil-involved allergic airway inflammation by intratracheal inoculation of house dust mites. PKCλ/ι-deficiency significantly inhibited airway inflammations. The infiltrating cells in the lungs and bronchoalveolar lavage fluids were significantly reduced in conditional PKCλ/ι-deficient mice. Th17 effector cytokines were reduced in the bronchoalveolar lavage fluids and lungs at protein and mRNA levels. Thus, PKCλ/ι emerges as a critical regulator of Th17 differentiation and allergic airway hyperresponsiveness. [ABSTRACT FROM AUTHOR]

Synaptic plasticity plays a critical role in cocaine addiction. The dopamine D1 and D3 receptors differentially regulate the cocaine-induced gene expression, structural remodeling and behavioral response. However, how these two receptors coordinately mediate the ultra-structural changes of synapses after cocaine exposure and whether these changes are behaviorally relevant are still not clear. Here, using quantitative electron microscopy, we show that D1 and D3 receptors have distinct roles in regulating cocaine-induced ultra-structural changes of synapses in the nucleus accumbens and caudoputamen. Pre-treatment of cocaine-treated mice with D3 receptor antagonist NGB2904 resulted in an increase in the ratio of total and asymmetric synapse to neuron and in the length of postsynaptic densities, compared with cocaine treatment alone. In contrast, pre-treatment of cocaine-treated mice with D1 receptor antagonist SCH23390 caused a reduction in synapse-to-neuron ratio and in postsynaptic densities length. Similarly, NGB2904 and SCH23390 showed opposite/differential effects on cocaine-induced structural plasticity, conditioned place preference and locomotor activity and signaling activation, including the activation of ERK, CREB and NR1 and the expression of c-fos and Cdk5. Therefore, we provide direct electron microscopy evidence that dopamine D1 and D3 receptors reciprocally regulate the ultra-structural changes of synapses following chronic exposure to cocaine. In addition, our data suggest that D1 and D3 receptors may regulate cocaine-induced ultra-structural changes and behavior responses by impact on structural plasticity and signaling transduction. [ABSTRACT FROM AUTHOR]

The mammalian target of rapamycin (mTOR), present in mTOR complex 1 (mTORC1) and mTORC2, is a serine/threonine kinase that integrates nutrients, growth factors, and cellular energy status to control protein synthesis, cell growth, survival and metabolism. However, it remains elusive whether mTOR plays a developmental stage-specific role in tissue development and whether mTOR can function independent of its complexes and kinase activity. In this study, by inducible genetic manipulation approach, we investigated the role of mTOR and its dependence on mTOR complexes and kinase activity in mitochondrial fitness of early, progenitor stage (lineage-negative; Lin-) versus later, lineage-committed stage (lineage-positive; Lin+) of hematopoietic cells. We found that oxidative phosphorylation (OXPHOS), ATP production and mitochondrial DNA synthesis were decreased in mTOR-/- Lin- cells but increased in mTOR-/- Lin+ cells, suggesting that mTOR plays a developmental stage-specific role in OXPHOS, ATP production and mitochondrial DNA synthesis. In contrast to mTOR deletion, simultaneous deletion of Raptor, a key component of mTORC1, and Rictor, a key component of mTORC2, led to increased mitochondrial DNA in Lin- cells and decreased mitochondrial DNA and ATP production in Lin+ cells, suggesting that mTOR regulates mitochondrial DNA synthesis in Lin- and Lin+ cells and ATP production in Lin+ cells independent of mTORC1 and mTORC2. Similar to mTOR deletion, deletion of Raptor alone attenuated glycolysis and increased mitochondrial mass and mitochondrial membrane potential in Lin- cells and increased mitochondrial mass and OXPHOS in Lin+ cells, whereas deletion of Rictor alone had no effect on these mitochondrial parameters in Lin- and Lin+ cells, suggesting that mTOR regulates glycolysis and mitochondrial membrane potential in Lin- cells, OXPHOS in Lin+ cells, and mitochondrial mass in both Lin- and Lin+ cells dependent on mTORC1, but not mTORC2. Either Raptor deficiency or Rictor deficiency recapitulated mTOR deletion in decreasing OXPHOS in Lin- cells and glycolysis in Lin+ cells, suggesting that mTOR regulates OXPHOS in Lin- cells and glycolysis in Lin+ cells dependent on both mTORC1 and mTORC2. Finally, mice harboring a mTOR kinase dead D2338A knock-in mutant showed decreased glycolysis in Lin+ cells, as seen in mTOR-/- Lin+ cells, but no change in glycolysis in Lin- cells, in contrast to the decreased glycolysis in mTOR-/- Lin- cells, suggesting that mTOR regulates glycolysis in Lin+ cells dependent on its kinase activity, whereas mTOR regulates glycolysis in Lin- cells independent of its kinase activity. [ABSTRACT FROM AUTHOR]

The “hygiene hypothesis” is a theory try to explain the dramatic increases in the prevalence of autoimmune and allergic diseases over the past two to three decades in developed countries. According to this theory, reduced exposure to parasites and microorganisms in childhood is the main cause for the increased incidences of both T helper 1 (Th1)-mediated autoimmunity and Th2-mediated allergy. In this study, we investigated the impact of Schistosoma japonicum infection on the allergic airway inflammation induced by repeated intracheal inoculations of house dust mites (HDM), which is a Th17 and neutrophils dominant murine asthma model, mimicking severe asthma. We found that S. japonicum infection downregulated airway hyperresponsiveness. The infiltrating cells, Th17 and Th2 effector cytokines in the bronchoalveolar lavage (BAL) fluids and lungs were significantly reduced in the infected mice. Our findings indicated that S. japonicum infection was able to effectively inhibit host’s allergic airway inflammation, which may be related to the upregulated Treg cells upon infection. To our knowledge, it is the first study to reveal the impact of S. japonicum infection on house dust mite induced severe asthma. More in depth investigation is need to elucidate the underlying mechanisms. [ABSTRACT FROM AUTHOR]

Hippocampus plays critical roles in drug addiction. Cocaine-induced modifications in dopamine receptor function and the downstream signaling are important regulation mechanisms in cocaine addiction. Rac regulates actin filament accumulation while Cdc42 stimulates the formation of filopodia and neurite outgrowth. Based on the region specific roles of small GTPases in brain, we focused on the hippocampal subregions to detect the regulation of Cdc42 signaling in long-term morphological and behavioral adaptations to cocaine. Genetically modified mouse models of Cdc42, dopamine receptor D1 (D1R) and D2 (D2R) and expressed Cdc42 point mutants that are defective in binding to and activation of its downstream effector molecules PAK and N-WASP were generated, respectively, in CA1 or dentate gyrus (DG) subregion. Cocaine induced upregulation of Cdc42 signaling activity. Cdc42 knockout or mutants blocked cocaine-induced increase in spine plasticity in hippocampal CA1 pyramidal neurons, leading to a decreased conditional place preference (CPP)-associated memories and spatial learning and memory in water maze. Cdc42 knockout or mutants promoted cocaine-induced loss of neurogenesis in DG, leading to a decreased CPP-associated memories and spatial learning and memory in water maze. Furthermore, by using D1R knockout, D2R knockout, and D2R/Cdc42 double knockout mice, we found that D2R, but not D1R, regulated Cdc42 signaling in cocaine-induced neural plasticity and behavioral changes. Cdc42 acts downstream of D2R in the hippocampus and plays an important role in cocaine-induced neural plasticity through N-WASP and PAK-LIMK-Cofilin, and Cdc42 signaling pathway correlatively links specific brain regions (CA1, dentate gyrus) to cocaine-induced CPP behavior. • Cocaine induced increasing hippocampal Cdc42 signaling • Cdc42 signaling regulated cocaine-associated hippocampal plasticity. • D2R regulated Cdc42 signaling in cocaine-induced hippocampal plasticity. [ABSTRACT FROM AUTHOR]

Repeated exposure to cocaine was previously found to cause sensitized behavioral responses and structural remodeling on medium spiny neurons of the nucleus accumbens (NAc) and caudate putamen (CPu). Rac1 has emerged as a key integrator of environmental cues that regulates dendritic cytoskeletons. In this study, we investigated the role of Rac1 in cocaine-induced dendritic and behavioral plasticity in the CPu. We found that Rac1 activation was reduced in the NAc but increased in the CPu following repeated cocaine treatment. Inhibition of Rac1 activity by a Rac1-specific inhibitor NSC23766, overexpression of a dominant negative mutant of Rac1 (T17N-Rac1) or local knockout of Rac1 attenuated the cocaine-induced increase in dendrites and spine density in the CPu, whereas overexpression of a constitutively active Rac1 exert the opposite effect. Moreover, NSC23766 reversed the increased number of asymmetric spine synapses in the CPu following chronic cocaine exposure. Downregulation of Rac1 activity likewise attenuates behavioral reward responses to cocaine exposure, with activation of Rac1 producing the opposite effect. Thus, Rac1 signaling is differentially regulated in the NAc and CPu after repeated cocaine treatment, and induction of Rac1 activation in the CPu is important for cocaine exposure-induced dendritic remodeling and behavioral plasticity. [ABSTRACT FROM AUTHOR]

Chemotaxis promotes neutrophil participation in cellular defense by enabling neutrophil migration to infected tissue and is controlled by persistent cell polarization. One long-standing question of neutrophil polarity has been how the pseudopod and the uropod are coordinated. In our previous report, we suggested that Rho GTPase Cdc42 controls neutrophil polarity through CD11 b signaling at the uropod, albeit through an unknown mechanism. Here, we show that Cdc42 controls polarity, unexpectedly, via its effector WASp. Cdc42 controls WASp activation and its distant localization to the uropod. At the uropod, WASp regulates the reorganization of CD11b integrin into detergent resistant membrane domains; in turn, CD11b recruits the microtubule end binding protein EB1 to capture and stabilize microtubules at the uropod. This organization is necessary to maintain neutrophil polarity during migration and is critical for neutrophil emigration into inflamed lungs. These results suggest unrecognized mechanism of neutrophil polarity in which WASp mediates long-distance control of the uropod by Cdc42 to maintain a proper balance between the pseudopod and the uropod. Our study reveals a new function for WASp in the control of neutrophil polarity via crosstalk between CD11 b and microtubules. [ABSTRACT FROM AUTHOR]

Abstract: Repeated exposure to cocaine can induce persistent alterations in the brain''s reward system, including increases in the number of dendrites and spine density on medium-sized spiny neurons (MSNs) in the nucleus accumbens (NAc). The structural remodeling of dendrites and spines in the NAc is thought to play a critical role in cocaine addiction. MSNs in the NAc can be classified by expression of either D1 or D2 dopamine receptors, which are localized to the direct and indirect pathway, respectively. It is unknown whether the dendritic changes induced by repeated cocaine treatment occur in MSNs of the direct or indirect pathway. Because the traditional Golgi-Cox impregnation of neurons precludes identifying particular subpopulations of MSNs, we performed dendritic morphology analysis after biocytin-labeling and Golgi-Cox impregnation. We found that the biocytin staining MSNs showed higher dendritic spine density and higher number of dendrites than that in Golgi impregnation group. In addition, we found that the increasing spine density induced by repeated cocaine treatment in female mice was higher than that in male mice. Next we used biocytin staining and dynorphin/D2 receptor colocalization to determine which cell type(s) displayed dendritic changes after repeated cocaine treatment. We found that cocaine-induced changes in dendritic parameters occurred in MSNs of both the direct (D1-expressing) and indirect (D2-expressing) pathways. [Copyright &y& Elsevier]

Abstract: Activation of RhoA prevents NGF-induced outgrowth and causes retraction of neurites in neuronal cells, including PC12 cells. Despite its inhibitory effect on neurite outgrowth, NGF activates GTP loading of and effector binding to RhoA, setting up an apparent contradiction. According to the molecular switch hypothesis of GTPase function GTP-loading of RhoA should be sufficient to activate its effectors uniformly. However, when monitoring NGF-induced binding of GTP-RhoA to multiple targets, we noted differential interactions with its effectors. We found that NGF elicits a protein kinase A-mediated phosphorylation of RhoA on serine188, which renders it unable to bind to Rho-associated kinase (ROK), whereas it retains the ability to interact with other RhoA targets including rhotekin, mDia-1 and PKN. We show in vitro and in vivo that phosphorylation of serine188 represents an additional switch, capable of directing signals among effector pathways. In the context of PC12 cell differentiation, NGF-induced phosphorylation of RhoA on serine188 prevents it from interacting with ROK, which would otherwise block neurite outgrowth. Transfection of RhoAS188A mutant into PC12 cells prevents NGF-induced neurite outgrowth, just like constitutively activated RhoA14V does, indicating the requirement of this phosphorylation site. Replacement of serine188 with the phosphomimetic glutamate residue in RhoAV14/S188E selectively impairs interaction with ROK and when transfected into PC12 cells restores NGF-induced neurite outgrowth. Therefore, phosphorylation of serine188 may serve as a novel secondary switch of RhoA capable of overriding GTP-binding-elicited effector activation to a subset of targets such as ROK, which interact with the C-terminus of RhoA. [Copyright &y& Elsevier]

Background: Schistosomiasis is an immunopathogenic disease in which Th17 cells play vital roles. Hepatic granuloma formation and subsequent fibrosis are its main pathologic manifestations and the leading causes of hepatic cirrhosis, and effective therapeutic interventions are lacking. In this study, we explored the effects of fasudil, a selective RhoA–Rho-associated kinase (ROCK) inhibitor, on Th17 cells and the pathogenesis of schistosomiasis. Methods: Mice were infected with Schistosoma japonicum and treated with fasudil. The worm burden, hepatic granuloma formation, and fibrosis were evaluated. The roles of fasudil on Th17, Treg, and hepatic stellate cells were analyzed. Results: Fasudil therapy markedly reduced the granuloma size and collagen deposit in livers from mice infected with S. japonicum. However, fasudil therapy did not affect the worm burden in infected mice. The underlying cellular and molecular mechanisms were investigated. Fasudil suppressed the activation and induced the apoptosis of CD4+ T cells. Fasudil inhibited the differentiation and effector cytokine secretion of Th17 cells, whereas it upregulated Treg cells in vitro. It also restrained the in vivo interleukin (IL)-4 and IL-17 levels in infected mice. Fasudil directly induced the apoptosis of hepatic stellate cells and downregulated the expressions of hepatic fibrogenic genes, such as collagen type I (Col-I), Col-III, and transforming growth factor-1 (TGF-β1). These effects may contribute to its anti-pathogenic roles in schistosomiasis. Conclusions: Fasudil inhibits hepatic granuloma formation and fibrosis with downregulation of Th17 cells. Fasudil might serve as a novel therapeutic agent for hepatic fibrosis due to schistosome infections and perhaps other disorders. [ABSTRACT FROM AUTHOR]