Influenza virus assembly requires the completion of viral protein and vRNP transport to the assembly site at the plasma membrane. Therefore, efficient regulation of intracellular transport of the viral proteins and vRNPs to the surface of the host cell is especially important for virus morphogenesis. Influenza A virus uses the machineries of host cells to transport its own components including ribonucleoproteins (vRNPs) and three transmembrane proteins hemagglutinin (HA), neuraminidase (NA) and matrix 2 protein (M2). It has been shown that newly synthesized vRNPs are associated with active form of Rab11 and accumulate at recycling endosomes adjacent to the microtubule organizing center (MTOC) following nuclear export. Subsequently, they are transported along the microtubule network toward the plasma membranes in cargo vesicles. The viral transmembrane proteins are translated on the rough endoplasmic reticulum and transported to the virus assembly site at the plasma membrane. It has been found that several host factors such as ARHGAP21 and GTPase Cdc42 are involved in regulation of intracellular trafficking of influenza A virus transmembrane proteins including NA. In this review, we will highlight the current knowledge about anterograde transport and its regulation of the influenza A virus transmembrane proteins and genome in the host cytoplasm.
Cytoplasm ; metabolism ; GTP Phosphohydrolases ; metabolism ; GTPase-Activating Proteins ; metabolism ; Genome, Viral ; Hemagglutinin Glycoproteins, Influenza Virus ; metabolism ; Humans ; Influenza A virus ; genetics ; pathogenicity ; physiology ; Neuraminidase ; metabolism ; Protein Transport ; Ribonucleoproteins ; metabolism ; Viral Matrix Proteins ; metabolism ; cdc42 GTP-Binding Protein ; metabolism

BACKGROUND AND OBJECTIVEMicroRNAs have emerged as post-transcriptional regulators that are critically involved in the biologic behavior of cells. This study was designed to investigate the effect of members of the microRNA-29 family on the expression of cell division cycle 42 (Cdc42) and their roles on proliferation, migration, and invasion of gastric cancer cells.

METHODSWe detected microRNA-29s and Cdc42 expression in gastric cancer cells by real-time polymerase chain reaction (PCR) and Western blot analysis. Negative controlled RNA (ncontrol), microRNA-29 family members (microRNA-29a, -29b, and -29c), and Cdc42-specific small interfering RNA (si-Cdc42) were chemically synthesized and transfected into SGC7901 and BGC823 gastric cancer cells, which have a relatively low expression of microRNA-29s and a relatively high expression of Cdc42. The expression of Cdc42 and the phosphorylation of its downstream molecular PAK1 expressions were determined by Western bolt analysis. Cell Counting Kit-8 was used to measure cell proliferation, and wound-healing and invasion assays were used to examine the abilities of migration and invasion.

RESULTSSimilar to si-Cdc42, the ectopic expression of microRNA-29 family members significantly reduced the expression of Cdc42 and its downstream molecular PAK1 phosphorylation levels. Consistently, ectopic expression of microRNA-29s inhibited proliferation and migration in gastric cancer cells. Invasive cell counts of the SGC7901, ncontrol/SGC7901, si-Cdc42/SGC7901, microRNA-29a/SGC7901, microRNA-29b/SGC7901, and microRNA-29c/SGC7901 cell groups were 84.0+/-4.2, 71.7+/-4.6, 16.3+/-3.2, 15.7+/-3.8, 16.3+/-3.0, and 16.7+/-3.1, respectively. The invasive cell counts of the BGC823, ncontrol/BGC823, si-Cdc42/BGC823, microRNA-29a/BGC823, microRNA-29b/BGC823, and microRNA-29c/BGC823 cell groups were 199.0+/-10.5, 146.3+/-9.7, 72.7+/-8.2, 86.7+/-8.5, 86.0+/-8.5, and 73.3+/-8.3, respectively (P<0.05).

CONCLUSIONSMembers of the microRNA-29 family can obviously inhibit cell proliferation, migration, and invasion of gastric cancer cells by targeting Cdc42.

Animals ; Cell Line, Tumor ; Cell Movement ; Cell Proliferation ; Gene Expression Regulation, Neoplastic ; Humans ; Mice ; MicroRNAs ; genetics ; metabolism ; NIH 3T3 Cells ; Neoplasm Invasiveness ; Phosphorylation ; Stomach Neoplasms ; genetics ; metabolism ; pathology ; Transfection ; cdc42 GTP-Binding Protein ; metabolism ; p21-Activated Kinases ; metabolism

BACKGROUNDHigh microvascular permeability plays an essential role in pathological process of multiple diseases such as septic shock, acute lung injury and acute respiratory distress syndrome, and burns. Inhibiting hyperpermeability is significant for controlling these conditions. Cdc42, as a main member of the small Rho GTPase family, plays a critical role in controlling and regulating the endothelial junctional permeability. We aimed to generate and identify endothelial specific cdc42-deficient mice by the Cre/loxp recombination approach, for examination in an animal model of the contribution of the cdc42 gene in the microvascular barrier function.

METHODSWe crossed cdc42(Flox/Flox) mice with mice expressing endothelial cell-specific Cre recombinase, and the offspring with the genotype cdc42(Flox/+)Tie2Cre(+/-) were back-crossed with the cdc42(Flox/Flox) mice. The cdc42(Flox/Flox)Tie2Cre(+/-) mice in the F2 generation were the target mice. If the cdc42 deficient mice did not survive, we would observe the cdc42 deficient mice embryos, and compare them with wild-type mice embryos.

RESULTSCdc42(flox/+)Cre(+/-) mice were mated with the cdc42(Flox/Flox) mice and among the living offspring there were no cdc42(Flox/Flox)Cre(+/-) target mice. We found the endothelial special cdc42 deficient embryos at the E7.5-E16.5 stage. We observed that cdc42 deficient embryos were much smaller, had fewer vessels and were a little more swollen compared with the wild-type embryos.

CONCLUSIONSEndothelial specific knockout of cdc42 caused embryonic lethality and the mice did not survive to birth. The target embryos were much smaller, had fewer vessels and were a little more swollen compared with the wild-type embryos. These results demonstrated that the cdc42 plays an important role in development of embryos and in development of microvessels as well as microvascular permeability.

Animals ; Embryo, Mammalian ; blood supply ; metabolism ; Endothelium, Vascular ; embryology ; metabolism ; Female ; Immunohistochemistry ; Male ; Mice ; Mice, Inbred C57BL ; Mice, Knockout ; Neovascularization, Physiologic ; genetics ; physiology ; cdc42 GTP-Binding Protein ; genetics ; metabolism

Diabetic nephropathy (DN) is a progressive kidney disease that is caused by injury to kidney glomeruli. Podocytes are glomerular epithelial cells and play critical roles in the glomerular filtration barrier. Recent studies have shown the importance of regulating the podocyte actin cytoskeleton in early DN. The phosphoinositide 3-kinase (PI3K) inhibitor, wortmannin, simultaneously regulates Rac1 and Cdc42, which destabilize the podocyte actin cytoskeleton during early DN. In this study, in order to evaluate the reno-protective effects of wortmannin in early DN by regulating Rac1 and Cdc42, streptozotocin (STZ)-induced proteinuric renal disease (SPRD) rats were treated with wortmannin. The albuminuria value of the SPRD group was 3.55 +/- 0.56 mg/day, whereas wortmannin group was 1.77 +/- 0.48 mg/day. Also, the albumin to creatinine ratio (ACR) value of the SPRD group was 53.08 +/- 10.82 mg/g, whereas wortmannin group was 20.27 +/- 6.41 mg/g. Changes in the expression level of nephrin, podocin and Rac1/Cdc42, which is related to actin cytoskeleton in podocytes, by wortmannin administration were confirmed by Western blotting. The expression levels of nephrin (79.66 +/- 0.02), podocin (87.81 +/- 0.03) and Rac1/Cdc42 (86.12 +/- 0.02) in the wortmannin group were higher than the expression levels of nephrin (55.32 +/- 0.03), podocin (53.40 +/- 0.06) and Rac1/Cdc42 (54.05 +/- 0.04) in the SPRD group. In addition, expression and localization of nephrin, podocin and desmin were confirmed by immunofluorescence. In summary, we found for the first time that wortmannin has a reno-protective effect on SPRD rats during the early DN. The beneficial effects of wortmannin in SPRD rats indicate that this compound could be used to delay the progression of the disease during the early DN stage.
Albumins/metabolism ; Androstadienes/*administration & dosage/pharmacology ; Animals ; Creatinine/blood ; Desmin/genetics/metabolism ; Diabetes Mellitus, Experimental/*drug therapy/metabolism/pathology ; Diabetic Nephropathies/*drug therapy/metabolism/pathology ; Disease Models, Animal ; Humans ; Intracellular Signaling Peptides and Proteins/genetics/metabolism ; Kidney/*pathology ; Membrane Proteins/genetics/metabolism ; Phosphatidylinositol 3-Kinases/*antagonists & inhibitors ; Podocytes/*drug effects/metabolism/pathology ; Rats ; Rats, Inbred Strains ; cdc42 GTP-Binding Protein/genetics/metabolism ; rac1 GTP-Binding Protein/genetics/metabolism

The establishment of a polarized cellular morphology is essential for a variety of processes including neural tube morphogenesis and the development of the brain. Cdc42 is a Ras-related GTPase that plays an essential role in controlling cell polarity through the regulation of the actin and microtubule cytoskeleton architecture. Previous studies have shown that Cdc42 plays an indispensable role in telencephalon development in earlier embryo developmental stage (before E12.5). However, the functions of Cdc42 in other parts of brain in later embryo developmental stage or in adult brain remain unclear. Thus, in order to address the role of Cdc42 in the whole brain in later embryo developmental stage or in adulthood, we used Cre/loxP technology to generate two lines of tissue-specific Cdc42-knock-out mice. Inactivation of Cdc42 was achieved in neuroepithelial cells by crossing Cdc42/ flox mice with Nestin-Cre mice and resulted in hydrocephalus, causing death to occur within the postnatal stage. Histological analyses of the brains from these mice showed that ependymal cell differentiation was disrupted, resulting in aqueductal stenosis. Deletion of Cdc42 in the cerebral cortex also induced obvious defects in interkinetic nuclear migration and hypoplasia. To further explore the role of Cdc42 in adult mice brain, we examined the effects of knocking-out Cdc42 in radial glial cells by crossing Cdc42/flox mice with human glial fibrillary acidic protein (GFAP)-Cre mice. Inactivation of Cdc42 in radial glial cells resulted in hydrocephalus and ependymal cell denudation. Taken together, these results highlight the importance of Cdc42 for ependymal cell differentiation and maintaining, and suggest that these functions likely contribute to the essential roles played by Cdc42 in the development of the brain.
Animals ; Brain ; metabolism ; pathology ; Cell Differentiation ; Cell Polarity ; Cerebral Cortex ; cytology ; metabolism ; Constriction, Pathologic ; Embryo, Mammalian ; metabolism ; Embryonic Development ; Ependyma ; cytology ; metabolism ; Glial Fibrillary Acidic Protein ; genetics ; metabolism ; Humans ; Hydrocephalus ; metabolism ; pathology ; Integrases ; genetics ; metabolism ; Mice ; Mice, Knockout ; cdc42 GTP-Binding Protein ; genetics ; metabolism

OBJECTIVETo investigate the changes of Cdc42 expression under estrogen stimulation, and to explore the signaling pathway of intracellular material transportation caused by estrogen.

METHODSMTT was used to test the drug sensitivity of cells. Real-time PCR was used to evaluate the expression of Cdc42 mRNA. The amount of ADM accumulated in MCF-7 cells was detected by flow cytometry. The protein levels of active-Cdc42 and Total-Cdc42 were measured by Western blot.

RESULTSIC(50) of ADM in MCF-7 cells was increased from (0.098 ± 0.011) µg/ml to (0.134 ± 0.130) µg/ml (P < 0.05) after estrogen stimulation. The amount of ADM accumulated in MCF-7 cells was reduced from 7.253 ± 0.310 to 3.233 ± 0.313 (P < 0.05). All of Cdc42 mRNA, active-Cdc42 protein and total-Cdc42 protein were increased (P < 0.05). After the treatment with siRNA, the IC(50) of ADM in siRNA group was decreased to (0.057 ± 0.017) µg/ml (P < 0.05) compared with that in the control group. The amount of accumulated ADM was significantly increased in the siRNA group, and all the expression levels of Cdc42 mRNA, active-Cdc42 protein and total-Cdc42 protein were decreased in the siRNA group (P < 0.05).

CONCLUSIONSEstrogen enhances the drug resistance in breast cancer cells. The mechanism of this effect may be via the enhancing Cdc42 expression and decreasing the accumulation of chemotherapeutic drugs in the cancer cells.

Antibiotics, Antineoplastic ; metabolism ; pharmacology ; Breast Neoplasms ; genetics ; metabolism ; pathology ; Cell Line, Tumor ; Doxorubicin ; metabolism ; pharmacology ; Drug Resistance, Multiple ; Drug Resistance, Neoplasm ; Estrogens ; pharmacology ; Female ; Humans ; Inhibitory Concentration 50 ; RNA Interference ; RNA, Messenger ; metabolism ; RNA, Small Interfering ; genetics ; Transfection ; cdc42 GTP-Binding Protein ; genetics ; metabolism