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https://www.readbyqxmd.com/read/29133117/application-of-mixed-peptide-arrays-to-study-combinatorial-readout-of-chromatin-modifications
#1
Rebekka Mauser, Goran Kungulovski, David Meral, Daniel Maisch, Albert Jeltsch
The N-terminal tails of histone proteins are massively decorated with post-translational modifications (PTMs), which play important roles in the regulation of gene expression. Several highly conserved chromatin interacting proteins can bind to histone modifications in a sequence and modification specific manner employing specific reading domains. These proteins often contain several reading domains, which can cooperate in the readout of different PTMs. To gain a better insight into the combinatorial readout of PTMs, we developed a method to study the binding of double reading domains to mixed peptide arrays containing two different peptides in each spot...
November 10, 2017: Biochimie
https://www.readbyqxmd.com/read/29126969/snhg16-mir-216-5p-zeb1-signal-pathway-contributes-to-the-tumorigenesis-of-cervical-cancer-cells
#2
Hong Zhu, Yan Zeng, Chen-Chen Zhou, Weiping Ye
Long non-coding RNAs (lncRNAs) have been confirmed as crucial regulators in tumorgenesis. Small nucleolar RNA host gene 16 (SNHG16) has been recently uncovered to be a potential oncogene in several types of cancers. However, its expression level and potential role in cervical cancer remain uncertain. In our research, we assessed the expression level of SNHG16 in clinical cervical cancer tissues and cells. We made use of functional assays to determine the biological effects of SNHG16 on cell proliferation and migration of cervical cancer...
November 7, 2017: Archives of Biochemistry and Biophysics
https://www.readbyqxmd.com/read/29080175/investigating-ornithine-decarboxylase-posttranscriptional-regulation-via-a-pulldown-assay-using-biotinylated-transcripts
#3
Anh Mai, Shannon L Nowotarski
Ornithine decarboxylase (ODC) is the first rate-limiting enzyme in the polyamine biosynthetic pathway. It has been well documented that ODC is tightly regulated at the levels of transcription, posttranscriptional changes in RNA, and protein degradation during normal conditions and that these processes are dysregulated during tumorigenesis. Moreover, it has been recently shown that ODC is posttranscriptionally regulated by RNA binding proteins (RBPs) which can bind to the ODC mRNA transcript and alter its stability and translation...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29073096/staufen1-inhibits-myod-translation-to-actively-maintain-muscle-stem-cell-quiescence
#4
Antoine de Morrée, Cindy T J van Velthoven, Qiang Gan, Jayesh S Salvi, Julian D D Klein, Igor Akimenko, Marco Quarta, Stefano Biressi, Thomas A Rando
Tissue regeneration depends on the timely activation of adult stem cells. In skeletal muscle, the adult stem cells maintain a quiescent state and proliferate upon injury. We show that muscle stem cells (MuSCs) use direct translational repression to maintain the quiescent state. High-resolution single-molecule and single-cell analyses demonstrate that quiescent MuSCs express high levels of Myogenic Differentiation 1 (MyoD) transcript in vivo, whereas MyoD protein is absent. RNA pulldowns and costainings show that MyoD mRNA interacts with Staufen1, a potent regulator of mRNA localization, translation, and stability...
October 24, 2017: Proceedings of the National Academy of Sciences of the United States of America
https://www.readbyqxmd.com/read/29058444/combining-in-vitro-and-in-silico-single-molecule-force-spectroscopy-to-characterize-and-tune-cellulosomal-scaffoldin-mechanics
#5
Tobias Verdorfer, Rafael C Bernardi, Aylin Meinhold, Wolfgang Ott, Zaida Luthey-Schulten, Michael A Nash, Hermann E Gaub
Cellulosomes are polyprotein machineries that efficiently degrade cellulosic material. Crucial to their function are scaffolds consisting of highly homologous cohesin domains, which serve a dual role by coordinating a multiplicity of enzymes as well as anchoring the microbe to its substrate. Here we combined two approaches to elucidate the mechanical properties of the main scaffold ScaA of Acetivibrio cellulolyticus. A newly developed parallelized one-pot in vitro transcription-translation and protein pull-down protocol enabled high-throughput atomic force microscopy (AFM)-based single-molecule force spectroscopy (SMFS) measurements of all cohesins from ScaA with a single cantilever, thus promising improved relative force comparability...
November 3, 2017: Journal of the American Chemical Society
https://www.readbyqxmd.com/read/29058181/monitoring-changes-in-the-abundance-of-endogenously-expressed-atp-sensitive-potassium-katp-channels-in-the-plasma-membrane-using-surface-biotinylation
#6
Jing-Syuna Ruan, Pei-Chun Chen
The conductance of KATP channel activity can be regulated by gating and/or surface expression. Gating analysis is usually performed by electrophysiological recording. Analysis of surface KATP channel expression levels requires cell fractionation, protein separation, and quantification. Cell fractionation involves time-consuming high-speed centrifugation steps and skilled techniques for taking out specific layers. Moreover, contamination of intracellular membranes can confound results. Although commercial kits have been developed to make it easier for scientists, qualities of these kits vary which can give rise to variable results...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29049411/xrcc5-cooperates-with-p300-to-promote-cyclooxygenase-2-expression-and-tumor-growth-in-colon-cancers
#7
Zhifeng Zhang, Fufu Zheng, Zhenlong Yu, Jiajiao Hao, Miao Chen, Wendan Yu, Wei Guo, Yiming Chen, Wenlin Huang, Zhijun Duan, Wuguo Deng
Cyclooxygenase (COX) is the rate-limiting enzyme in prostaglandins (PGs) biosynthesis. Previous studies indicate that COX-2, one of the isoforms of COX, is highly expressed in colon cancers and plays a key role in colon cancer carcinogenesis. Thus, searching for novel transcription factors regulating COX-2 expression will facilitate drug development for colon cancer. In this study, we identified XRCC5 as a binding protein of the COX-2 gene promoter in colon cancer cells with streptavidin-agarose pulldown assay and mass spectrometry analysis, and found that XRCC5 promoted colon cancer growth through modulation of COX-2 signaling...
2017: PloS One
https://www.readbyqxmd.com/read/29029323/lncrnal20992-regulates-apoptotic-proteins-to-promote-lead-induced-neuronal-apoptosis
#8
Aruo Nan, Yangyang Jia, Xin Li, Meiling Liu, Nan Zhang, Lijian Chen, Ti Yang, Yiqin Xu, Xin Dai, Ying Cheng, Zhenzhong Liu, Yihui Ling, Yiguo Jiang
Lead is a heavy metal pollutant that is widely present in the environment and can seriously harm human health, especially the nervous system. Long noncoding RNAs (lncRNAs) play important roles in many physiological and pathological processes; however, there remains a lack of in-depth studies on the molecular mechanisms associated with lead neurotoxicity. Here, our results showed that lead exposure inhibited cell proliferation and promoted cell apoptosis. We observed that lncRNAL20992 was significantly upregulated in a lead-induced neuronal-injury cell model according to quantitative reverse transcription polymerase chain reaction...
September 25, 2017: Toxicological Sciences: An Official Journal of the Society of Toxicology
https://www.readbyqxmd.com/read/29028450/comparing-two-approaches-of-mir-34a-target-identification-biotinylated-mirna-pulldown-vs-mirna-overexpression
#9
Hassaan Mehboob Awan, Abdullah Shah, Farooq Rashid, Shuai Wei, Liang Chen, Ge Shan
microRNAs (miRNAs) are critical regulators of gene expression. For elucidating functional roles of miRNAs, it is critical to identify their direct targets. There are debates about whether pulldown of biotinylated miRNA mimics can be used to identify miRNA targets or not. Here we show that biotin-labelled miR-34a can be loaded to AGO2, and AGO2 immunoprecipitation can pulldown biotinylated miR-34a (Bio-miR pulldown). RNA-sequencing (RNA-seq) of the Bio-miR pulldown RNAs efficiently identified miR-34a mRNA targets, which could be verified with luciferase assays...
October 13, 2017: RNA Biology
https://www.readbyqxmd.com/read/29026005/low-expression-of-mir-30a-5p-induced-the-proliferation-and-invasion-of-oral-cancer-via-promoting-the-expression-of-fap
#10
Peng Ruan, Aili Tan, Zezhang Tao
The study aimed at investigating the effects of miR-30a-5p on the biological functions of oral cancer cells and figuring out the potential mechanism. We firstly verified the low expression of miR-30a-5p and high expression of FAP (Homo sapiens fibroblast activation protein alpha) in oral cancerous tissues and their negative correlation. Then, the target relationship between miR-30a-5p and FAP was validated by dual luciferase reporter assay and biotin-coupled miRNA pulldown assay. After transfection in Tca-8113 cells and SCC-15 cells, MTT, colony formation, Transwell and wound healing assays were performed to investigate how miR-30a-5p and FAP adjusted propagation, invasiveness and migration respectively...
October 12, 2017: Bioscience Reports
https://www.readbyqxmd.com/read/29020972/sumo1-modification-of-khsrp-regulates-tumorigenesis-by-preventing-the-tl-g-rich-mirna-biogenesis
#11
Haihua Yuan, Rong Deng, Xian Zhao, Ran Chen, Guofang Hou, Hailong Zhang, Yanli Wang, Ming Xu, Bin Jiang, Jianxiu Yu
BACKGROUND: MicroRNAs (miRNAs) are important regulators involved in diverse physiological and pathological processes including cancer. SUMO (small ubiquitin-like modifier) is a reversible protein modifier. We recently found that SUMOylation of TARBP2 and DGCR8 is involved in the regulation of the miRNA pathway. KHSRP is a single stranded nucleic acid binding protein with roles in transcription and mRNA decay, and it is also a component of the Drosha-DGCR8 complex promoting the miRNA biogenesis...
October 11, 2017: Molecular Cancer
https://www.readbyqxmd.com/read/29018350/folliculin-interacts-with-rab35-to-regulate-egf-induced-egfr-degradation
#12
Jianchao Zheng, Biao Duan, Shixiu Sun, Jie Cui, Jun Du, Yujie Zhang
Aims and Hypothesis: This study aims to investigate the mechanism involved in intracellular regulation of EGFR degradation induced by EGF. Methods: Phosphorylation of proteins related to EGFR signaling was examined by western blot analysis. Activation, connection between Rab35 and folliculin (FLCN) were assessed by pulldown, coimmunoprecipitation assays separately. The relationship between FLCN and cell growth was detected using gene overexpression and knock-down techniques. Results: Here, we demonstrate that interfering with FLCN, a tumor suppressor, reduces the rate of EGF-induced EGFR degradation, resulting in prolonged activation of downstream signaling...
2017: Frontiers in Pharmacology
https://www.readbyqxmd.com/read/29018139/shca-adaptor-protein-promotes-nephrin-endocytosis-and-is-upregulated-in-proteinuric-nephropathies
#13
Claire E Martin, Kelly A Petersen, Lamine Aoudjit, Manali Tilak, Vera Eremina, W Rod Hardy, Susan E Quaggin, Tomoko Takano, Nina Jones
Nephrin is a key structural component of the podocyte slit diaphragm, and proper expression of nephrin on the cell surface is critical to ensure integrity of the blood filtration barrier. Maintenance of nephrin within this unique cell junction has been proposed to require dynamic phosphorylation events and endocytic recycling, although the molecular mechanisms that control this interplay are poorly understood. Here, we investigated the possibility that the phosphotyrosine adaptor protein ShcA regulates nephrin turnover...
October 10, 2017: Journal of the American Society of Nephrology: JASN
https://www.readbyqxmd.com/read/28981715/rna-guanine-7-methyltransferase-catalyzes-the-methylation-of-cytoplasmically-recapped-rnas
#14
Jackson B Trotman, Andrew J Giltmier, Chandrama Mukherjee, Daniel R Schoenberg
Cap homeostasis is a cyclical process of decapping and recapping that impacts a portion of the mRNA transcriptome. The metastable uncapped forms of recapping targets redistribute from polysomes to non-translating mRNPs, and recapping is all that is needed for their return to the translating pool. Previous work identified a cytoplasmic capping metabolon consisting of capping enzyme (CE) and a 5'-monophosphate kinase bound to adjacent domains of Nck1. The current study identifies the canonical cap methyltransferase (RNMT) as the enzyme responsible for guanine-N7 methylation of recapped mRNAs...
October 13, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/28958078/stable-isotope-labeling-reveals-novel-insights-into-ubiquitin-mediated-protein-aggregation-with-age-calorie-restriction-and-rapamycin-treatment
#15
Nathan B Basisty, Yuxin Liu, Jason Reynolds, Pabalu P Karunadharma, Dao-Fu Dai, Jeanne Fredrickson, Richard P Beyer, Michael J MacCoss, Peter S Rabinovitch
Accumulation of protein aggregates with age was first described in aged human tissue over 150 years ago and has since been described in virtually every human tissue. Ubiquitin modifications are a canonical marker of insoluble protein aggregates; however, the composition of most age-related inclusions remains relatively unknown. To examine the landscape of age-related protein aggregation in vivo, we performed an antibody-based pulldown of ubiquitinated proteins coupled with metabolic labeling and mass spectrometry on young and old mice on calorie restriction (CR), rapamycin (RP)-supplemented, and control diets...
August 18, 2017: Journals of Gerontology. Series A, Biological Sciences and Medical Sciences
https://www.readbyqxmd.com/read/28935601/ctenopharyngodon-idella-perk-eif2ak3-decreases-cell-viability-by-phosphorylating-eif2%C3%AE-under-er-stress
#16
Tao Zhang, Dongming Li, Lijuan Wan, Xin Chen, Xiangqin Wang, Bin Zhong, Zhen Wu, Huiling Mao, Chengyu Hu
As an upstream kinase of eIF2α, protein kinase RNA-like ER (endoplasmic reticulum) kinase (PERK) is a type I transmembrane protein located in ER in eukaryotic cells. PERK is mainly composed of two domains, the intracavitary domain for BIP protein combination and the dissociative C-terminal region containing a typical serine/threonine kinase domain which promotes the phosphorylation of eIF2α. In this study, we cloned a PERK (also known as EIF2AK3) gene from grass carp (Ctenopharyngodon idella). The full-length cDNA of grass carp PERK (CiPERK) is 5192 bp including a 176 bp of 5' untranslated region, a 1719 bp of 3' untranslated region and a 3297 bp of the longest open reading frame (ORF) encoding 1098 amino acids...
September 19, 2017: Fish & Shellfish Immunology
https://www.readbyqxmd.com/read/28932319/dna-hypomethylation-of-a-transcription-factor-binding-site-within-the-promoter-of-a-gout-risk-gene-nrbp1-upregulates-its-expression-by-inhibition-of-tfap2a-binding
#17
Zaihua Zhu, Weida Meng, Peiru Liu, Xiaoxia Zhu, Yun Liu, Hejian Zou
BACKGROUND: Genome-wide association studies (GWASs) have identified dozens of loci associated with gout, but for most cases, the risk genes and the underlying molecular mechanisms contributing to these associations are unknown. This study sought to understand the molecular mechanism of a common genetic variant, rs780093, in the development of gout, both in vitro and in vivo. RESULTS: Nuclear receptor binding protein 1 (NRBP1), as a gout risk gene, and its regulatory region, 72 bp upstream of the transcription start site, designated as B1, were identified through integrative analyses of genome-wide genotype and DNA methylation data...
2017: Clinical Epigenetics
https://www.readbyqxmd.com/read/28928748/a-peptide-fc-opsonin-with-pan-amyloid-reactivity
#18
James S Foster, Angela D Williams, Sallie Macy, Tina Richey, Alan Stuckey, Daniel Craig Wooliver, Richa Koul-Tiwari, Emily B Martin, Stephen J Kennel, Jonathan S Wall
There is a continuing need for therapeutic interventions for patients with the protein misfolding disorders that result in systemic amyloidosis. Recently, specific antibodies have been employed to treat AL amyloidosis by opsonizing tissue amyloid deposits thereby inducing cell-mediated dissolution and organ improvement. To develop a pan-amyloid therapeutic agent, we have produced an Fc-fusion product incorporating a peptide, p5, which binds many if not all forms of amyloid. This protein, designated Fcp5, expressed in mammalian cells, forms the desired bivalent dimer structure and retains pan-amyloid reactivity similar to the p5 peptide as measured by immunosorbent assays, immunohistochemistry, surface plasmon resonance, and pulldown assays using radioiodinated Fcp5...
2017: Frontiers in Immunology
https://www.readbyqxmd.com/read/28912651/manual-resistance-versus-conventional-resistance-training-impact-on-strength-and-muscular-endurance-in-recreationally-trained-men
#19
Iván Chulvi-Medrano, Tamara Rial, Juan M Cortell-Tormo, Yasser Alakhdar, Caue V La Scala Teixeira, Laura Masiá-Tortosa, Sandor Dorgo
Manual resistance training (MRT) has been widely used in the field of physical therapy. It has also been used as a strength training method due to the accommodating resistance nature of this modality. The aim of the present study was to compare the effects of an 8-week MRT program on maximum strength and muscular endurance in comparison to conventional resistance training in recreationally trained men. Twenty healthy recreationally trained male subjects were recruited and divided into a MRT training group and a conventional training (CT) group...
September 2017: Journal of Sports Science & Medicine
https://www.readbyqxmd.com/read/28901216/creatine-or-vitamin-d-supplementation-in-individuals-with-a-spinal-cord-injury-undergoing-resistance-training-a-double-blinded-randomized-pilot-trial
#20
Samuel Amorim, Vitor Hugo Teixeira, Rui Corredeira, Maria Cunha, Bruno Maia, Paulo Margalho, Joana Pires
Purpose Determine whether creatine or vitamin D supplementation improves muscle strength in individuals with spinal cord injury undergoing resistance training. Methods Thirteen male and one female with spinal cord injury, from two Portuguese rehabilitation centers, were randomized to creatine (3g daily), vitamin D (25000 IU each two weeks) or placebo group in a double-blind design. All participants performed progressive resistance training during eight weeks. The outcome measures, obtained at baseline and after intervention, included: Sum of four skinfolds; Corrected arm muscle area; Seated medicine ball throw; Handgrip strength with dynamometer; Manual wheelchair slalom test and one repetition maximum for Chest press, Triceps, Pec deck and Lat pulldown...
September 13, 2017: Journal of Spinal Cord Medicine
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