Search results for: Rab5
#36472440 2022/12/06 To Up
Seneca Valley Virus Enters PK-15 Cells via Caveolae-Mediated Endocytosis and Macropinocytosis Dependent on Low-pH, Dynamin, Rab5, and Rab7.Seneca Valley virus (SVV), a new pathogen resulting in porcine vesicular disease, is prevalent in pig herds worldwide. Although an understanding of SVV biology pathogenesis is crucial for preventing and controlling this disease, the molecular mechanisms for the entry and post-internalization of SVV, which represent crucial steps in viral infection, are not well characterized. In this study, specific inhibitors, Western blotting, and immunofluorescence detection revealed that SVV entry into PK-15 cells depends on low-pH conditions and dynamin. Furthermore, results showed that caveolae-mediated endocytosis (CavME) contributes crucially to the internalization of SVV, as evidenced by cholesterol depletion, downregulation of caveolin-1 expression by small interfering RNA knockdown, and overexpression of a caveolin-1 dominant negative (caveolin-1-DN) in SVV-infected PK-15 cells. However, SVV entry into PK-15 cells did not depend on clathrin-mediated endocytosis (CME). Furthermore, treatment with specific inhibitors demonstrated that SVV entry into PK-15 cells via macropinocytosis depended on the Na+/H+ exchanger (NHE), p21-activated kinase 1 (Pak1), and actin rearrangement, but not phosphatidylinositol 3-kinase (PI3K). Electron microscopy showed that SVV particles or proteins were localized in CavME and macropinocytosis. Finally, knockdown of GTPase Rab5 and Rab7 by siRNA significantly inhibited SVV replication, as determined by measuring viral genome copy numbers, viral protein expression, and viral titers. In this study, our results demonstrated that SVV utilizes caveolae-mediated endocytosis and macropinocytosis to enter PK-15 cells, dependent on low pH, dynamin, Rab5, and Rab7. Entry of virus into cells represents the initiation of a successful infection. As an emerging pathogen of porcine vesicular disease, clarification of the process of SVV entry into cells enables us to better understand the viral life cycle and pathogenesis. In this study, patterns of SVV internalization and key factors required were explored. We demonstrated for the first time that SVV entry into PK-15 cells via caveolae-mediated endocytosis and macropinocytosis requires Rab5 and Rab7 and is independent of clathrin-mediated endocytosis, and that low-pH conditions and dynamin are involved in the process of SVV internalization. This information increases our understanding of the patterns in which all members of the family enter host cells, and provides new insights for preventing and controlling SVV infection.
Lei Hou, Xinxin Tong, Yang Pan, Ruihan Shi, Changzhe Liu, Jinshuo Guo, Yongyan Shi, Xiaoyu Yang, Yong Wang, Xufei Feng, Jianwei Zhou, Jue Liu
1388 related Products with: Seneca Valley Virus Enters PK-15 Cells via Caveolae-Mediated Endocytosis and Macropinocytosis Dependent on Low-pH, Dynamin, Rab5, and Rab7.10 mg200ul50 ug 25 mg5 x 10A5 cells/vial10 mg100 ug/vial25 mg100ug/vial15ml100ul5mg
#36459881 2022/11/22 To Up
Central nervous system specific high molecular weight ALS2/alsin homophilic complex is enriched in mouse brain synaptosomes.ALS2/alsin, the causative gene product for a number of juvenile recessive motor neuron diseases, acts as a guanine nucleotide exchange factor (GEF) for Rab5, regulating early endosome trafficking and maturation. It has been demonstrated that ALS2 forms a tetramer, and this oligomerization is essential for its GEF activity and endosomal localization in established cancer cells. However, despite that ALS2 deficiency is implicated in neurological diseases, neither the subcellular distribution of ALS2 nor the form of its complex in the central nervous system (CNS) has been investigated. In this study, we showed that ALS2 in the brain was enriched both in synaptosomal and cytosolic fractions, while those in the liver were almost exclusively present in cytosolic fraction by differential centrifugation. Gel filtration chromatography revealed that cytosolic ALS2 prepared both from the brain and liver formed a tetramer. Remarkably, synaptosomal ALS2 existed as a high-molecular weight complex in addition to a tetramer. Such complex was also observed not only in embryonic brain but also several neuronal and glial cultures, but not in fibroblast-derived cell lines. Thus, the high-molecular weight ALS2 complex represents a unique form of ALS2-homophilic oligomers in the CNS, which may play a role in the maintenance of neural function.
Kai Sato, Kyoko Suzuki-Utsunomiya, Shun Mitsui, Suzuka Ono, Kento Shimakura, Asako Otomo, Shinji Hadano
1702 related Products with: Central nervous system specific high molecular weight ALS2/alsin homophilic complex is enriched in mouse brain synaptosomes.4 Membranes/Box4 Arrays/Slide4 Membranes/Box4 Membranes/Box4 Arrays/Slide4 Membranes/Box4 Membranes/Box1 ml50 ul100 ul4 Arrays/Slide4 Membranes/Box
#36444566 2022/11/29 To Up
Functions of neuronal Synaptobrevin in the post-Golgi transport of rhodopsin in Drosophila photoreceptors.Polarized transport is essential for constructing multiple plasma membrane domains in the cell. Drosophila photoreceptors are an excellent model system to study the mechanisms of polarized transport. Rab11 is the key factor regulating the post-Golgi transport of rhodopsin 1 (Rh1), a photoreceptive protein, to the rhabdomere, a photoreceptive plasma membrane. Here, we found that neuronal Synaptobrevin (nSyb) colocalizes with Rab11 on the trans-side of Golgi stacks and post-Golgi vesicles at the rhabdomere-base, and nSyb-deficiency impairs rhabdomeric transport and induces accumulation of Rh1 and vesicles in the cytoplasm; this is similar to the effects of Rab11 loss. These results indicate that nSyb acts as a post-Golgi SNARE toward rhabdomeres. Surprisingly, in Rab11-, Rip11-, and nSyb-deficient photoreceptors, illumination enhances cytoplasmic accumulation of Rh1 colocalizing with Rab11, Rabenosyn5, nSyb, and Arrestin 1 (Arr1). Arr1 loss but not Rab5 dominant negative (Rab5DN) protein expression inhibits the light enhanced cytoplasmic Rh1 accumulation. Rab5DN inhibits the generation of Rh1 containing multi-vesicle bodies rather than Rh1 internalization. Overall, these results indicate that exocytic Rh1 mingle with endocytosed Rh1 and are then transported together to rhabdomeres.
Hitomi Yamashita, Yuka Ochi, Yumi Yamada, Shogo Sasaki, Tatsuya Tago, Takunori Satoh, Akiko K Satoh
2065 related Products with: Functions of neuronal Synaptobrevin in the post-Golgi transport of rhodopsin in Drosophila photoreceptors.11 Set100ug Lyophilized
#36389067 2022/10/31 To Up
Signaling abnormality leading to excessive/aberrant synaptic plasticity in Alzheimer's disease.
2461 related Products with: Signaling abnormality leading to excessive/aberrant synaptic plasticity in Alzheimer's disease.96 tests96 tests96 tests96 tests 50 UG8 inhibitors2 Pieces/Box2 Pieces/Box
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#36359874 2022/11/03 To Up
Fabricated AIE-Based Probe to Detect the Resistance to Anoikis of Cancer Cells Detached from Tumor Tissue.(1) Background: Resisting anoikis is a vital and necessary characteristic of malignant cancer cells, but there is no existing quantification method. Herein, a sensitive probe for assessing anoikis resistance of cancer cells detached from the extracellular matrix was developed based on the aggregation-induced emission (AIE) of AIEgens. It has been reported that detached cancer cell endocytose activated integrin clusters, and in the endosome these clusters recruit and activate phosphorylate focal adhesion kinase (pFAK) in the cytoplasm to induce signaling that supports the growth of detached cancer cells. (2) Methods: We established a lost nest cell model of cancer cells and determined their ability to resist anoikis. The colocalization of the activated integrin, pFAK, and endosomes in model cells was observed and calculated. (3) Results: The fluorescence signal intensity of the probe was significantly higher than that of the integrin antibody in the model cells and the fluorescence signal of probe signal was better overlapped with labeled pFAK by fluorescence in endosomes in model cells. (4) Conclusions: We developed a quantitative multi-parametric image analysis program to calculate fluorescent intensity of the probe and antibodies against pFAK and Rab5 in the areas of colocalization. A positive correlation of fluorescence signal intensity between the probe and pFAK on the endosome was observed. Therefore, the probe was used to quantitatively evaluate resisting anoikis of different cancer cell lines under the lost nest condition.
Ya-Nan Chang, Yuelan Liang, Jiayi Wang, Ziteng Chen, Ruyu Yan, Kui Chen, Juan Li, Jiacheng Li, Haojun Liang, Gengmei Xing
1358 related Products with: Fabricated AIE-Based Probe to Detect the Resistance to Anoikis of Cancer Cells Detached from Tumor Tissue.1 mg10
#36340038 2022/10/20 To Up
Participation of OCRL1, and APPL1, in the expression, proteolysis, phosphorylation and endosomal trafficking of megalin: Implications for Lowe Syndrome.Megalin/LRP2 is the primary multiligand receptor for the re-absorption of low molecular weight proteins in the proximal renal tubule. Its function is significantly dependent on its endosomal trafficking. Megalin recycling from endosomal compartments is altered in an X-linked disease called Lowe Syndrome (LS), caused by mutations in the gene encoding for the phosphatidylinositol 5-phosphatase OCRL1. LS patients show increased low-molecular-weight proteins with reduced levels of megalin ectodomain in the urine and accumulation of the receptor in endosomal compartments of the proximal tubule cells. To gain insight into the deregulation of megalin in the LS condition, we silenced OCRL1 in different cell lines to evaluate megalin expression finding that it is post-transcriptionally regulated. As an indication of megalin proteolysis, we detect the ectodomain of the receptor in the culture media. Remarkably, in OCRL1 silenced cells, megalin ectodomain secretion appeared significantly reduced, according to the observation in the urine of LS patients. Besides, the silencing of APPL1, a Rab5 effector associated with OCRL1 in endocytic vesicles, also reduced the presence of megalin's ectodomain in the culture media. In both silencing conditions, megalin cell surface levels were significantly decreased. Considering that GSK3ß-mediated megalin phosphorylation reduces receptor recycling, we determined that the endosomal distribution of megalin depends on its phosphorylation status and OCRL1 function. As a physiologic regulator of GSK3ß, we focused on insulin signaling that reduces kinase activity. Accordingly, megalin phosphorylation was significantly reduced by insulin in wild-type cells. Moreover, even though in cells with low activity of OCRL1 the insulin response was reduced, the phosphorylation of megalin was significantly decreased and the receptor at the cell surface increased, suggesting a protective role of insulin in a LS cellular model.
Lisette Sandoval, Luz M Fuentealba, María-Paz Marzolo
1918 related Products with: Participation of OCRL1, and APPL1, in the expression, proteolysis, phosphorylation and endosomal trafficking of megalin: Implications for Lowe Syndrome.2 Pieces/Box2 Pieces/Box1,000 tests100ul1 mg200 100 mg2 Pieces/Box
#36288306 2022/10/26 To Up
LC3-associated endocytosis and the functions of Rubicon and ATG16L1.LC3-associated endocytosis (LANDO) is a noncanonical function of the autophagy machinery, in which LC3 (microtubule-associated protein light chain) is conjugated to rab5-positive endosomes, using a portion of the canonical autophagy pathway. LANDO was initially discovered in a murine model of Alzheimer's disease as a critical regulator of amyloid-β receptor recycling in microglial cells, playing a protective role against neuronal loss and memory impairment. Recent evidence suggests an emerging role of LANDO in cytokine receptor signaling and innate immunity. Here, we discuss the regulation of two crucial effectors of LANDO, Rubicon and ATG16L1, and their impact on endocytosis, autophagy, and phagocytosis.
Joëlle Magné, Douglas R Green100ug1 g2.5 mg96 wells (1 kit)100ug200ul10 mg50 ug 100 mg 25 MG100ul25 mg
#36275033 2022/10/05 To Up
Vitamin D3 alters macrophage phenotype and endosomal trafficking markers in dairy cattle naturally infected with Mycobacterium avium subsp. paratuberculosis.Macrophages are important host defense cells in ruminant paratuberculosis (Johne's Disease; JD), a chronic enteritis caused by Mycobacterium avium subsp. paratuberculosis (MAP). Classical macrophage functions of pathogen trafficking, degradation, and antigen presentation are interrupted in mycobacterial infection. Immunologic stimulation by 25-hydroxyvitamin D3 (25(OH)D3) and 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) enhances bovine macrophage function. The present study aimed to investigate the role of vitamin D3 on macrophage phenotype and endosomal trafficking of MAP in monocyte-derived macrophages (MDMs) cultured from JD-, JD+ subclinical, and JD+ clinically infected cattle. MDMs were pre-treated 100 ng/ml 25(OH)D3 or 4 ng/ml 1,25(OH)2D3 and incubated 24 hrs with MAP at 10:1 multiplicity of infection (MOI). In vitro MAP infection upregulated pro-inflammatory (M1) CD80 and downregulated resolution/repair (M2) CD163. Vitamin D3 generally decreased CD80 and increased CD163 expression. Furthermore, early endosomal marker Rab5 was upregulated 140× across all stages of paratuberculosis infection following in vitro MAP infection; however, Rab5 was reduced in MAP-activated MDMs from JD+ subclinical and JD+ clinical cows compared to healthy controls. Rab7 expression decreased in control and clinical cows following MDM infection with MAP. Both forms of vitamin D3 reduced Rab5 expression in infected MDMs from JD- control cows, while 1,25(OH)2D3 decreased Rab7 expression in JD- and JD+ subclinical animals regardless of MAP infection in vitro. Vitamin D3 promoted phagocytosis in MDMs from JD- and JD+ clinical cows treated with either vitamin D3 analog. Results from this study show exogenous vitamin D3 influences macrophage M1/M2 polarization and Rab GTPase expression within MDM culture.
Taylor L T Wherry, Rohana P Dassanayake, John P Bannantine, Shankumar Mooyottu, Judith R Stabel
2086 related Products with: Vitamin D3 alters macrophage phenotype and endosomal trafficking markers in dairy cattle naturally infected with Mycobacterium avium subsp. paratuberculosis.5ug1 mg0.2 mg5ug5ug1 mg5ug2ug2ug2ug
#36267578 2022/10/04 To Up
RagC GTPase regulates mTOR to promote chemoresistance in senescence-like HepG2 cells.Radiotherapy and chemotherapy can arrest cancer cells in a senescence-like state, which can lead to therapy resistance and cancer relapse. mTOR is hyperactivated in senescent cells but the mechanisms remain unclear. In this study, we examine the roles of several mTOR-regulated GTPases in senescence-like liver cancer cells and the mechanisms in drug resistance. We show that although RagC, Rheb, Rab1A, Rab5 and Arf1 GTPases were required for optimal mTOR activation in proliferating HepG2 cells, only RagC and Rheb are required in the senescence-like counterparts. Consistently, the drug resistance of the senescence-like HepG2 can be reduced by knocking down RagC and Rheb but not the other GTPases. Autophagic and lysosomal activity were increased in senescence-like cells; pharmacological inhibition of autophagy-lysosome decreased mTOR activity and preferentially sensitized senescence-like HepG2 cells to chemotherapy drugs including trametinib, cisplatin, and doxorubicin. In liver cancer patients, expression of RagC and Rheb but not other GTPases examined was associated with unfavorable prognosis. Our study therefore has defined a key role of Rag-Rheb GTPase in mediating mTOR activation and drug resistance in senescence-like HepG2 cells, which could have important implications in developing second-line treatments for liver cancer patients.
Wei Jiang, Zhenglin Ou, Qin Zhu, Hongyan Zai
2951 related Products with: RagC GTPase regulates mTOR to promote chemoresistance in senescence-like HepG2 cells.96 wells10mg 1 G1mg1 mg1x10e7 cells5mg96 wells (1 kit)
#36227834 2022/10/13 To Up
The entry of unclosed autophagosomes into vacuoles and its physiological relevance.It is widely stated in the literature that closed mature autophagosomes (APs) fuse with lysosomes/vacuoles during macroautophagy/autophagy. Previously, we showed that unclosed APs accumulated as clusters outside vacuoles in Vps21/Rab5 and ESCRT mutants after a short period of nitrogen starvation. However, the fate of such unclosed APs remains unclear. In this study, we used a combination of cellular and biochemical approaches to show that unclosed double-membrane APs entered vacuoles and formed unclosed single-membrane autophagic bodies after prolonged nitrogen starvation or rapamycin treatment. Vacuolar hydrolases, vacuolar transport chaperon (VTC) proteins, Ypt7, and Vam3 were all involved in the entry of unclosed double-membrane APs into vacuoles in Vps21-mutant cells. Overexpression of the vacuolar hydrolases, Pep4 or Prb1, or depletion of most VTC proteins promoted the entry of unclosed APs into vacuoles in Vps21-mutant cells, whereas depletion of Pep4 and/or Prb1 delayed the entry into vacuoles. In contrast to the complete infertility of diploid cells of typical autophagy mutants, diploid cells of Vps21 mutant progressed through meiosis to sporulation, benefiting from the entry of unclosed APs into vacuoles after prolonged nitrogen starvation. Overall, these data represent a new observation that unclosed double-membrane APs can enter vacuoles after prolonged autophagy induction, most likely as a survival strategy.
Zulin Wu, Haiqian Xu, Pei Wang, Ling Liu, Juan Cai, Yun Chen, Xiaomin Zhao, Xia You, Junze Liu, Xiangrui Guo, Tingting Xie, Jiajie Feng, Fan Zhou, Rui Li, Zhiping Xie, Yanhong Xue, Chuanhai Fu, Yongheng Liang
2893 related Products with: The entry of unclosed autophagosomes into vacuoles and its physiological relevance.200ul125 mg500 Units2.5 mg1000 tests100ug100ug10 mg100ul
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