The cytoskeleton and motor proteins of human schistosomes and their roles in surface maintenance and host-parasite interactions

Schistosomes are parasitic blood flukes, responsible for significant human disease in tropical and developing nations. Here we review information on the organization of the cytoskeleton and associated motor proteins of schistosomes, with particular reference to the organization of the syncytial tegument, a unique cellular adaptation of these and other neodermatan flatworms. Extensive EST databases show that the molecular constituents of the cytoskeleton and associated molecular systems are likely to be similar to those of other eukaryotes, although there are potentially some molecules unique to schistosomes and platyhelminths. The biology of some components, particular those contributing to host-parasite interactions as well as chemotherapy and immunotherapy are discussed. Unresolved questions in relation to the structure and function of the tegument relate to dynamic organization of the syncytial layer. (C) 2004 Wiley Periodicals, Inc.

Classical cadherin adhesion molecules: coordinating cell adhesion, signaling and the cytoskeleton

Classical cadherin adhesion molecules are fundamental determinants of tissue organization in both health and disease. Recent advances in understanding the molecular and cellular basis of cadherin function have revealed that these adhesion molecules serve as molecular couplers, linking cell surface adhesion and recognition to both the actin cytoskeleton and cell signalling pathways. We will review some of these developments. to provide an overview of progress in this rapidly-developing area of cell and developmental biology.

H-ras, K-ras, and inner plasma membrane raft proteins operate in nanoclusters with differential dependence on the actin cytoskeleton

Plasma membrane compartmentalization imposes lateral segregation on membrane proteins that is important for regulating signal transduction. We use computational modeling of immunogold spatial point patterns on intact plasma membrane sheets to test different models of inner plasma membrane organization. We find compartmentalization at the nanoscale level but show that a classical raft model of preexisting stable domains into which lipid raft proteins partition is incompatible with the spatial point patterns generated by the immunogold labeling of a palmitoylated raft marker protein. Rather, approximate to 30% of the raft protein exists in cholesterol-dependent nanoclusters, with approximate to 70% distributed as monomers. The cluster/monomer ratio (number of proteins in clusters/number of proteins outside clusters) is independent of expression level. H-rasG12V and K-rasG12V proteins also operate in nanoclusters with fixed cluster/monomer ratios that are independent of expression level. Detailed calibration of the immunogold imaging protocol suggests that radii of raft and RasG12V protein nanoclusters may be as small as 11 and 6 nm, respectively, and shows that the nanoclusters contain small numbers (6.0-7.7) of proteins. Raft nanoclusters do not form if the actin cytoskeleton is disassembled. The formation of K-rasG12V but not H-rasG12V nanoclusters also is actin-dependent. K-rasG12V but not H-rasG12V signaling is abrogated by actin cytoskeleton disassembly, which shows that nanoclustering is critical for Ras function. These findings argue against stable preexisting domains on the inner plasma membrane in favor of dynamic actively regulated nanoclusters similar to those proposed for the outer plasma membrane. RasG12V nanoclusters may facilitate the assembly of essential signal transduction complexes.

PKC-alpha-mediated remodeling of the actin cytoskeleton is involved in constitutive albumin uptake by proximal tubule cells

One key role of the renal proximal tubule is the reabsorption of proteins from the glomerular filtrate by constitutive receptor-mediated endocytosis. In the opossum kidney (OK) renal proximal tubule cell line, inhibition of protein kinase C (PKC) reduces albumin uptake, although the isoforms involved and mechanisms by which this occurs have not been identified. We used pharmacological and molecular approaches to investigate the role of PKC-α in albumin endocytosis. We found that albumin uptake in OK cells was inhibited by the pan-PKC blocker bisindolylmaleimide-1 and the isoform-specific PKC blockers Go-6976 and 2',3,3',4,4'-hexahydroxy-1,1'-biphenyl-6,6'-dimethanol dimethyl ether, indicating a role for PKC-α. Overexpression of a kinase deficient PKC-α(K368R) but not wild-type PKC-α significantly reduced albumin endocytosis. Western blot analysis of fractionated cells showed an increased association of PKC-α-green fluorescent protein with the membrane fraction within 10-20 min of exposure to albumin. We used phalloidin to demonstrate that albumin induces the formation of clusters of actin at the apical surface of OK cells and that these clusters correspond to the location of albumin uptake. These clusters were not present in cells grown in the absence of albumin. In cells treated either with PKC inhibitors or overexpressing kinase-deficient PKC-α(K368R) this actin cluster formation was significantly reduced. This study identifies a role for PKC-α in constitutive albumin uptake in OK cells by mediating assembly of actin microfilaments at the apical membrane.

Expression of human neuronal protein 22, a novel cytoskeleton-associated protein, was decreased in the anterior cingulate cortex of schizophrenia

Human neuronal protein 22 (hNP22) is a novel neuron-specific protein featuring numerous motifs previously described in cytoskeleton-associating and signaling proteins. Because previous studies have supported abnormalities in neuronal cytoarchitecture and/or development in the schizophrenia brain, we examined the expression of hNP22 in the anterior cingulate cortex, the hippocampus and the prefrontal cortex of schizophrenic and normal control postmortem brains using high-sensitive immunohistochemistry. Seven schizophrenic and seven age- and sex-matched control brains were examined. The ratio of hNP22-immunopositive cells/total cells was significantly reduced in layer V (p = .020) and layer VI (p = .022) of the anterior cingulate cortex of schizophrenic brain compared with controls. In contrast, there were no significant changes observed in the hippocampus and the prefrontal cortex. These results suggest that altered expression of hNP22 may be associated with modifications in neuronal cytoarchitecture leading to dysregulation of neural signal transduction in the anterior cingulate cortex of the schizophrenia brain.

The novel cytoskeleton-associated protein Neuronal protein 22: Elevated expression in the developing rat brain

Neuronal development and process targeting is mediated by proteins of the cytoskeleton. However, the signaling pathways underlying these mechanisms are complex and have not yet been fully elucidated. Neuronal protein 22 (NP22) has been identified as a cytoskeleton-associated protein. It colocalizes with microtubules and actin, the two major components of the cytoskeleton. It contains numerous signaling motifs and induces process formation in non-neuronal cells. Expression of rat NP22 (rNP22) rises incrementally at specific time points during brain development, with the greatest elevation occurring during synaptogenesis in the rat brain. its neuronal localization is primarily at the plasma membrane of the soma in the embryonic brain and progresses into homogeneous expression in the postnatal rat brain. Data suggest that NP22 may play a role in mediating the molecular events governing development of the neuronal architecture. Furthermore, its sustained expression in postnatal brain implies a function in the maintenance of neuronal morphology.

Minimal mutation of the cytoplasmic tail inhibits the ability of E-cadherin to activate Rac but not phosphatidylinositol 3-kinase - Direct evidence of a role for cadherin-activated Rac signaling in adhesion and contact formation

Classic cadherins are adhesion-activated cell signaling receptors. In particular, homophilic cadherin ligation can directly activate Rho family GTPases and phosphatidylinositol 3-kinase (PI3-kinase), signaling molecules with the capacity to support the morphogenetic effects of these adhesion molecules during development and disease. However, the molecular basis for cadherin signaling has not been elucidated, nor is its precise contribution to cadherin function yet understood. One attractive hypothesis is that cadherin-activated signaling participates in stabilizing adhesive contacts ( Yap, A. S., and Kovacs, E. M. ( 2003) J. Cell Biol. 160, 11-16). We now report that minimal mutation of the cadherin cytoplasmic tail to uncouple binding of p120-ctn ablated the ability of E-cadherin to activate Rac. This was accompanied by profound defects in the capacity of cells to establish stable adhesive contacts, defects that were rescued by sustained Rac signaling. These data provide direct evidence for a role of cadherin-activated Rac signaling in contact formation and adhesive stabilization. In contrast, cadherin-activated PI3-kinase signaling was not affected by loss of p120-ctn binding. The molecular requirements for E-cadherin to activate Rac signaling thus appear distinct from those that stimulate PI3-kinase, and we postulate that p120-ctn may play a central role in the E-cadherin-Rac signaling pathway.

Direct cadherin-activated cell signaling: a view from the plasma membrane

Classical cadherin adhesion molecules are key determinants of cell recognition and tissue morphogenesis, with diverse effects on cell behavior. Recent developments indicate that classical cadherins are adhesion-activated signaling receptors. In particular, early-immediate Rac signaling is emerging as a mechanism to coordinate cadherin-actin integration at the plasma membrane.

Cofilin interacts with ClC-5 and regulates albumin uptake in proximal tubule cell lines

Receptor-mediated endocytosis is a constitutive high capacity pathway for the reabsorption of proteins from the glomerular filtrate by the renal proximal tubule. ClC-5 is a voltage-gated chloride channel found in the proximal tubule where it has been shown to be essential for protein uptake, based on evidence from patients with Dent's disease and studies in ClC-5 knockout mice. To further delineate the role of ClC-5 in albumin uptake, we performed a yeast two-hybrid screen with the C-terminal tail of ClC-5 to identify any interactions of the channel with proteins involved in endocytosis. We found that the C-terminal tail of ClC-5 bound the actin depolymerizing protein, cofilin, a result that was confirmed by GST-fusion pulldown assays. In cultured proximal tubule cells, cofilin was distributed in nuclear, cytoplasmic, and microsomal fractions and co-localized with ClC-5. Phosphorylation of cofilin by overexpressing LIM kinase 1 resulted in a stabilization of the actin cytoskeleton. Phosphorylation of cofilin in two proximal tubule cell models (porcine renal proximal tubule and opossum kidney) was also accompanied by a pronounced inhibition of albumin uptake. This study identifies a novel interaction between the C-terminal tail of ClC-5 and cofilin, an actin-associated protein that is crucial in the regulation of albumin uptake by the proximal tubule.

Isolation and characterization of a Cotesia rubecula bracovirus gene expressed in the lepidopteran Pieris rapae

Polydnaviruses are endogenous particles that are crucial for the survival of endoparasitoid wasps, providing active suppression of the immune function of the lepidopteran host in which wasp larvae develop. The Cotesia rubecula bracovirus (CrBV) is unique in that only four gene products are detected in larval host (Pieris rapae) tissues and expression of CrBV genes is transient, occurring between 4 and 12 h post-parasitization. Two of the four genes, CrV1 and CrV3, have been characterized. CrV1 is a secreted glycoprotein that has been implicated in depolymerization of the actin cytoskeleton of host haemocytes, leading to haemocyte inactivation; CrV3 is a multimeric C-type lectin that shares homology with insect immune lectins. Here, a third CrBV-specific gene is described, CrV2, which is expressed in larval P. rapae tissues. CrV2, which is transcribed in haemocytes and fat body cells, has an ORF of 963 bp that produces a glycoprotein of approximately 40 kDa. CrV2 is secreted into haemolymph and appears to be internalized by host haemocytes. CrV2 has a coiled-coil region predicted at its C-terminus, which may be involved in the formation of putative CrV2 trimers that are detected in haemolymph of parasitized host larvae.

Lectin-induced haemocyte inactivation in insects

Most multimeric lectins are adhesion molecules, promoting attachment and spreading on surface glycodeterminants. In addition, some lectins have counter-adhesion properties, detaching already spread cells which then acquire round or spindle-formed cell shapes. Since lectin-mediated adhesion and detachment is observed in haemocyte-like Drosophila cells, which have haemomucin as the major lectin-binding glycoprotein, the two opposite cell behaviours may be the result of lectin-mediated receptor rearrangements on the cell surface. To investigate oligomeric lectins as a possible extracellular driving force affecting cell shape changes, we examined lectin-mediated reactions in lepidopteran haemocytes after cytochalasin D-treatment and observed that while cell-spreading was dependent on F-actin, lectin-uptake was less dependent on F-actin. We propose a model of cell shape changes involving a dynamic balance between adhesion and uptake reactions. (C) 2004 Elsevier Ltd. All rights reserved.

Evolution of polydnaviruses as insect immune suppressors

Polydnaviruses (PDVs) are endogenous particles that are used by some endoparasitic hymenoptera to disrupt host immunity and development. Recent analyses of encapsidated PDV genes have increased the number of known PDV gene families, which are often closely related to insect genes. Several PDV proteins inactivate host haemocytes by damaging their actin cytoskeleton. These proteins share no significant sequence homology and occur in polyphyletic PDV genera, possibly indicating that convergent evolution has produced functionally similar immune-suppressive molecules causing a haemocyte phenotype characterised by damaged cytoskeleton and inactivation. These phenomena provide further insights into the immune-suppressive activity of PDVs and raise interesting questions about PDV evolution, a topic that has puzzled researchers ever since the discovery of PDVs.

The LFA-1-associated molecule PTA-1 (CD226) on T cells forms a dynamic molecular complex with protein 4.1G and human discs large

Clustering of the T cell integrin, LFA-1, at specialized regions of intercellular contact initiates integrin-mediated adhesion and downstream signaling, events that are necessary for a successful immunological response. But how clustering is achieved and sustained is not known. Here we establish that an LFA-1-associated molecule, PTA-1, is localized to membrane rafts and binds the carboxyl-terminal domain of isoforms of the actin-binding protein 4.1G. Protein 4.1 is known to associate with the membrane-associated guanylate kinase homologue, human discs large. We show that the carboxyl-terminal peptide of PTA-1 also can bind human discs large and that the presence or absence of this peptide greatly influences binding between PTA-1 and different isoforms of 4.1G. T cell stimulation with phorbol ester or PTA-1 cross-linking induces PTA-1 and 4.1G to associate tightly with the cytoskeleton, and the PTA-1 from such activated cells now can bind to the amino-terminal region of 4.1G. We propose that these dynamic associations provide the structural basis for a regulated molecular adhesive complex that serves to cluster and transport LFA-1 and associated molecules.