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Pull down assay

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https://www.readbyqxmd.com/read/28102638/farnesoid-x-receptor-ablation-sensitizes-mice-to-hepatitis-b-virus-x-protein-induced-hepatocarcinogenesis
#1
Yongdong Niu, Meishu Xu, Betty L Slagle, Haihua Huang, Song Li, Grace L Guo, Ganggang Shi, Wenxin Qin, Wen Xie
: Chronic hepatitis B virus infection is a major risk factor for hepatocellular carcinoma (HCC). Hepatitis B virus X protein (HBx) is a hepatitis B virus protein that has multiple cellular functions, but its role in HCC pathogenesis has been controversial. Farnesoid X receptor (FXR) is a nuclear receptor with activities in anti-inflammation and inhibition of hepatocarcinogenesis. However, whether or how FXR can impact hepatitis B virus/HBx-induced hepatocarcinogenesis remains unclear...
November 5, 2016: Hepatology: Official Journal of the American Association for the Study of Liver Diseases
https://www.readbyqxmd.com/read/28102235/clumping-factor-a-of-staphylococcus-aureus-interacts-with-annexina2-on-mammary-epithelial-cells
#2
Shoaib Ashraf, Jing Cheng, Xin Zhao
Staphylococcus aureus is one of major pathogens that can cause a series of diseases in different hosts. In the bovine, it mainly causes subclinical and contagious mastitis, but its mechanisms of infection are not fully understood. Considering the fact that virulence factors play key roles in interactions between the bacterium and host cells, this study aimed to identify if a binding partner of S. aureus clumping factor A (ClfA) exists on the bovine mammary epithelial cells. The ClfA protein was used as a bait to pull down lysates of cultured bovine mammary epithelial cells (MAC-T cells)...
January 19, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28100698/cdc45-induced-loading-of-human-rpa-onto-single-stranded-dna
#3
Anna Szambowska, Ingrid Tessmer, Piotr Prus, Bernhard Schlott, Helmut Pospiech, Frank Grosse
Cell division cycle protein 45 (Cdc45) is an essential component of the eukaryotic replicative DNA helicase. We found that human Cdc45 forms a complex with the single-stranded DNA (ssDNA) binding protein RPA. Moreover, it actively loads RPA onto nascent ssDNA. Pull-down assays and surface plasmon resonance studies revealed that Cdc45-bound RPA complexed with ssDNA in the 8-10 nucleotide binding mode, but dissociated when RPA covered a 30-mer. Real-time analysis of RPA-ssDNA binding demonstrated that Cdc45 catalytically loaded RPA onto ssDNA...
January 18, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/28092041/in-solution-sh2-domain-binding-assay-based-on-proximity-ligation
#4
Kazuya Machida
Protein-protein interactions mediated by SH2 domains confer specificity in tyrosine kinase pathways. Traditional assays for assessing interactions between an SH2 domain and its interacting protein such as far-Western and pull-down are inherently low throughput. We developed SH2-PLA, an in-solution SH2 domain binding assay, that takes advantage of the speed and sensitivity of proximity ligation and real-time PCR. SH2-PLA allows for rapid assessment of SH2 domain binding to a target protein using only a few microliters of cell lysate, thereby making it an attractive new tool to study tyrosine kinase signaling...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28092040/binding-assays-using-recombinant-sh2-domains-far-western-pull-down-and-fluorescence-polarization
#5
Kazuya Machida, Bernard Liu
Recognition of phosphotyrosine-containing sequences by SH2 domains confers specificity in tyrosine kinase pathways. By assessing interactions between isolated SH2 domains and their binding proteins, it is possible to gain insight into otherwise inaccessible complex cellular systems. Far-Western, pull-down, and fluorescence polarization (FP) have been frequently used for characterization of phosphotyrosine signaling. Here, we outline standard protocols for these established assays using recombinant SH2 domain, emphasizing the importance of appropriate sample preparation and assay controls...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28081465/biological-function-and-mechanism-of-lncrna-meg3-in-tenon-s-capsule-fibroblasts-proliferation-by-meg3-nrf2-protein-interaction
#6
Yuan Wang, Jing Wang, Li-Juan Wei, Dong-Mei Zhu, Jin-Song Zhang
OBJECTIVE: The proliferation of Tenon's capsule fibroblasts after glaucoma filtration surgery is a harmful element for operation failure. However, the underlying mechanism remains unclear. In this study, we evaluated the role of long non-coding RNA (lncRNA)-MEG3 in the proliferation of Glaucoma Tenon's capsule fibroblasts (GTFs) with a model of transforming growth factor-β (TGF-β)-induced proliferation which has been used in previous studies to explore the mechanism underlying glaucoma surgery failure...
January 9, 2017: Biomedicine & Pharmacotherapy, Biomédecine & Pharmacothérapie
https://www.readbyqxmd.com/read/28079129/circrna_000203-enhances-the-expression-of-fibrosis-associated-genes-by-derepressing-targets-of-mir-26b-5p-col1a2-and-ctgf-in-cardiac-fibroblasts
#7
Chun-Mei Tang, Ming Zhang, Lei Huang, Zhi-Qin Hu, Jie-Ning Zhu, Zhen Xiao, Zhuo Zhang, Qiu-Xiong Lin, Xi-Long Zheng, Min -Yang, Shu-Lin Wu, Jian-Ding Cheng, Zhi-Xin Shan
Circular RNAs (circRNAs) participate in regulating gene expression in diverse biological and pathological processes. The present study aimed to investigate the mechanism underlying the modulation of circRNA_000203 on expressions of fibrosis-associated genes in cardiac fibroblasts. CircRNA_000203 was shown upregulated in the diabetic mouse myocardium and in Ang-II-induced mouse cardiac fibroblasts. Enforced-expression of circRNA_000203 could increase expressions of Col1a2, Col3a1 and α-SMA in mouse cardiac fibroblasts...
January 12, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28067229/tubules-of-plant-reoviruses-exploit-tropomodulin-to-regulate-actin-based-tubule-motility-in-insect-vector
#8
Qian Chen, Linghua Zhang, Yanshuang Zhang, Qianzhuo Mao, Taiyun Wei
Plant reoviruses are known to exploit virion-packaging tubules formed by virus-encoding non-structural proteins for viral spread in insect vectors. Tubules are propelled by actin-based tubule motility (ABTM) to overcome membrane or tissue barriers in insect vectors. To further understand which insect factors mediate ABTM, we utilized yeast two-hybrid and bimolecular fluorescence complementation assays to test interactions between tubule protein Pns10 of rice dwarf virus (RDV), a plant reovirus, and proteins of its insect vector, the leafhopper Nephotettix cincticeps...
January 9, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28061540/metallothionein-2a-an-interactive-protein-linking-phosphorylated-fadd-to-nf-%C3%AE%C2%BAb-pathway-leads-to-colorectal-cancer-formation
#9
Faiz M M T Marikar, Guanghui Jin, Wang Sheng, Dingyuan Ma, Zichun Hua
BACKGROUND: The rapid increase in the incidence rate of colorectal cancer has led to the search and identification of biomarkers that can predict risk for and future behavior of this malignancy and management. To study the biological role of the phosphorylated Fas associated death domain (pFADD) gene in colorectal cancer, we performed a GAL4-based yeast two-hybrid screening of a human heart cDNA library. METHODS: A series of two yeast hybrid method was used to identification of protein-protein interaction...
December 2016: Chinese Clinical Oncology
https://www.readbyqxmd.com/read/28053002/jab1-is-a-t2-dependent-coactivator-or-a-t3-dependent-corepressor-of-trb1-mediated-gene-regulation
#10
Gabriela Hernández-Puga, Arturo Mendoza, Alfonso León-Del-Río, Aurea Orozco
Thyroid hormones (THs) induce pleiotropic effects in vertebrates, mainly through the activation or repression of gene expression. These mechanisms involve thyroid hormone binding to thyroid hormone receptors, an event that is followed by the sequential recruitment of coactivator or corepressor proteins, which in turn modify the rate of transcription. In the present study we looked for specific coregulators recruited by the long isoform of the teleostean thyroid hormone receptor beta 1 (L-Trb1) when bound to the bioactive TH 3,5-T2 (T2)...
January 4, 2017: Journal of Endocrinology
https://www.readbyqxmd.com/read/28049590/inhibitory-potential-of-flavonoids-on-ptdins-3-4-5-p3-binding-with-the-phosphoinositide-dependent-kinase-1-pleckstrin-homology-domain
#11
Yelin Kang, Bong-Gyu Kim, Sunghoon Kim, Youngshim Lee, Youngdae Yoon
Many membrane-associated proteins are involved in various signaling pathways, including the phosphoinositide 3-kinase (PI3K) pathway, which has key roles in diverse cellular processes. Disruption of the activities of these proteins is involved in the development of disease in humans, making these proteins promising targets for drug development. In most cases, the catalytic domain is targeted; however, it is also possible to target membrane associations in order to regulate protein activity. In this study, we established a novel method to study protein-lipid interactions and screened for flavonoid-derived antagonists of PtdIns(3,4,5)P3 binding with the phosphoinositide-dependent kinase 1 (PDK1) pleckstrin homology (PH) domain...
December 23, 2016: Bioorganic & Medicinal Chemistry Letters
https://www.readbyqxmd.com/read/28039930/interacting-protein-partners-of-arabidopsis-rna-binding-protein-atrbp45b
#12
Meenakumari Muthuramalingam, Yixing Wang, Yongfang Li, Ramamurthy Mahalingam
• RNA binding proteins (RBPs), important players in post-transcriptional gene regulation usually exist in ribonuclear complexes. However, even in model systems like Arabidopsis characterization of RBP associated proteins are limited. In this study, we investigated the interacting proteins of the Arabidopsis AtRBP45b, which is involved in stress signaling. • In vivo localization of AtRBP45b was conducted using 35S-GFP. FLAG-tagged AtRBP45b under the control of 35S promoter in the Atrbp45b-1 mutant background was used to pull down AtRBP45b interacting proteins...
December 31, 2016: Plant Biology
https://www.readbyqxmd.com/read/28036027/the-phosphorylation-of-pdx-1-by-protein-kinase-ck2-is-crucial-for-its-stability
#13
Sabrina Klein, Rui Meng, Mathias Montenarh, Claudia Götz
The homeodomain protein PDX-1 is a critical regulator of pancreatic development and insulin production in pancreatic β-cells. We have recently shown that PDX-1 is a substrate of protein kinase CK2; a multifunctional protein kinase which is implicated in the regulation of various cellular aspects, such as differentiation, proliferation, and survival. The CK2 phosphorylation site of PDX-1 is located within the binding region of the E3 ubiquitin ligase adaptor protein PCIF1. To study the interaction between PDX-1 and PCIF1 we used immunofluorescence analysis, co-immunoprecipitation, GST-pull-down studies, and proximity ligation assay (PLA)...
December 28, 2016: Pharmaceuticals
https://www.readbyqxmd.com/read/28034779/a-mitochondrial-membrane-protein-is-a-target-for-rice-ragged-stunt-virus-in-its-insect-vector
#14
Hai-Jian Huang, Cheng-Wen Liu, Xiang Zhou, Chuan-Xi Zhang, Yan-Yuan Bao
Rice ragged stunt virus (RRSV; Reoviridae) is exclusively transmitted by the brown planthopper Nilaparvata lugens in a persistent-propagative manner. It is understood that RNA viral proliferation is associated with the intracellular membranes of the insect host cells. However, the molecular mechanisms of the interaction between the RRSV proliferation and the intracellular membranes remain essentially unknown. It will be of great interest to determine whether RRSV protein(s) directly interact with intracellular membrane components of its host cells...
February 2, 2017: Virus Research
https://www.readbyqxmd.com/read/28032027/valosin-containing-protein-is-a-target-of-5-l-fuligocandin%C3%A2-b-and-enhances-trail-resistance-in-cancer-cells
#15
Midori A Arai, Shota Taguchi, Kazuhiro Komatsuzaki, Kento Uchiyama, Ayaka Masuda, Mana Sampei, Mamoru Satoh, Sayaka Kado, Masami Ishibashi
Fuligocandin B (2) is a novel natural product that can overcome TRAIL resistance. We synthesized enatiomerically pure fuligocandin B (2) and its derivative 5'-I fuligocandin B (4), and found that the latter had an improved biological activity against the human gastric cancer cell line, AGS. We attached a biotin linker and photoactivatable aryl diazirine group to 5'-I fuligocandin B (4), and employed a pull-down assay to identify valosin-containing protein (VCP/p97), an AAA ATPase, as a 5'-I fuligocandin B (4) target protein...
December 2016: ChemistryOpen
https://www.readbyqxmd.com/read/28031320/tetraspanin-cd63-controls-basolateral-sorting-of-organic-cation-transporter-2-in-renal-proximal-tubules
#16
Ulf Schulze, Sabine Brast, Alexander Grabner, Christian Albiker, Beatrice Snieder, Svenja Holle, Eberhard Schlatter, Rita Schröter, Hermann Pavenstädt, Edwin Herrmann, Carsten Lambert, Gilles A Spoden, Luise Florin, Paul Saftig, Giuliano Ciarimboli
CD63 is a ubiquitously expressed member of the tetraspanin superfamily. Using a mating-based split-ubiquitin-yeast 2-hybrid system, pull-down experiments, total internal reflection fluorescence microscopy, Förster resonance energy transfer, and biotinylation assays, we found that CD63 interacts with human organic cation transporter 2 (hOCT2), which transports endogenous and exogenous substrates, such as neurotransmitters and drugs in several epithelial cells. CD63 overexpression affects cellular localization of hOCT2 expressed in human embryonic kidney (HEK)293 cells...
December 28, 2016: FASEB Journal: Official Publication of the Federation of American Societies for Experimental Biology
https://www.readbyqxmd.com/read/28031293/the-roles-of-ush1-proteins-and-pdz-domain-containing-ush-proteins-in-ush2-complex-integrity-in-cochlear-hair-cells
#17
Junhuang Zou, Qian Chen, Ali Almishaal, Pranav Dinesh Mathur, Tihua Zheng, Cong Tian, Qing Y Zheng, Jun Yang
Usher syndrome (USH) is the most common cause of inherited deaf-blindness, manifested as USH1, USH2 and USH3 clinical types. The protein products of USH2 causative and modifier genes, USH2A, ADGRV1, WHRN and PDZD7, interact to assemble a multiprotein complex at the ankle link region of the mechanosensitive stereociliary bundle in hair cells. Defects in this complex cause stereociliary bundle disorganization and hearing loss. The four USH2 proteins also interact in vitro with USH1 proteins including myosin VIIa, USH1G (SANS), CIB2 and harmonin...
December 28, 2016: Human Molecular Genetics
https://www.readbyqxmd.com/read/28028166/potato-tuber-induction-is-regulated-by-interactions-between-components-of-a-tuberigen-complex
#18
Chin-Jit Teo, Kenta Takahashi, Kanae Shimizu, Ko Shimamoto, Ken-Ichiro Taoka
Photoperiod-regulated flowering and potato tuber formation involve leaf-produced mobile signals, florigen and tuberigen, respectively. The major protein component of florigen has been identified as the FLOWERING LOCUS T (FT) protein. In rice, an FT-like protein, Heading date 3a (Hd3a), induces flowering by making the florigen activation complex (FAC) through interactions with 14-3-3 and OsFD1, a rice FD-like protein. In potato, StSP6A, an FT-like protein, was identified as a major component of tuberigen. However, the molecular mechanism of how StSP6A triggers tuber formation remains elusive...
December 27, 2016: Plant & Cell Physiology
https://www.readbyqxmd.com/read/28027319/scavenger-receptor-c-mediates-phagocytosis-of-white-spot-syndrome-virus-and-restricts-virus-proliferation-in-shrimp
#19
Ming-Chong Yang, Xiu-Zhen Shi, Hui-Ting Yang, Jie-Jie Sun, Ling Xu, Xian-Wei Wang, Xiao-Fan Zhao, Jin-Xing Wang
Scavenger receptors are an important class of pattern recognition receptors that play several important roles in host defense against pathogens. The class C scavenger receptors (SRCs) have only been identified in a few invertebrates, and their role in the immune response against viruses is seldom studied. In this study, we firstly identified an SRC from kuruma shrimp, Marsupenaeus japonicus, designated MjSRC, which was significantly upregulated after white spot syndrome virus (WSSV) challenge at the mRNA and protein levels in hemocytes...
December 2016: PLoS Pathogens
https://www.readbyqxmd.com/read/28026967/identification-of-molecular-targets-for-4-5-dichloro-2-n-octyl-4-isothiazolin-3-one-dcoit-in-teleosts-new-insight-into-mechanism-of-toxicity
#20
Lianguo Chen, Doris W T Au, Chenyan Hu, Drew R Peterson, Bingsheng Zhou, Pei-Yuan Qian
Environmental pollutants are capable of concomitantly inducing diverse toxic effects. However, it is largely unknown which effects are directly induced and which effects are secondary, thus calling for definitive identification of the initiating molecular event for a pollutant to elucidate the mechanism of toxicity. In present study, affinity pull-down assays were used to identify target proteins for 4,5-dichloro-2-n-octyl-4-isothiazolin-3-one (DCOIT), a costal pollutant of emerging concern, in various tissues (e...
December 27, 2016: Environmental Science & Technology
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