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Subcellular RNA

Vanesa Mongelli, Maria-Carla Saleh
Like every other organism on Earth, insects are infected with viruses, and they rely on RNA interference (RNAi) mechanisms to circumvent viral infections. A remarkable characteristic of RNAi is that it is both broadly acting, because it is triggered by double-stranded RNA molecules derived from virtually any virus, and extremely specific, because it targets only the particular viral sequence that initiated the process. Reviews covering the different facets of the RNAi antiviral immune response in insects have been published elsewhere...
September 29, 2016: Annual Review of Virology
Dipen Rajgor, Jonathan G Hanley, Catherine M Shanahan
Nesprins are highly conserved spectrin-repeat containing scaffold proteins pre-dominantly known to function at the nuclear envelope (NE). However, nesprin isoforms are emerging with localizations and scaffolding functions at sites away from the NE, suggesting their functions are more diverse than originally thought. In this study, we combined nesprin-1 coimmunoprecipitations with mass spectrometry to identify novel nesprin-1 binding partners for isoforms that localize to subcellular compartments beyond the NE...
October 12, 2016: Molecular Biology of the Cell
Yan Xu, Jie Chen, Lan Xiao, Hee Kyoung Chung, Yuan Zhang, Joseph C Robinson, Jaladanki N Rao, Jian-Ying Wang
The RNA-binding protein HuR is crucial for normal intestinal mucosal regeneration by modulating the stability and translation of target mRNAs, but the exact mechanism underlying HuR trafficking between the cytoplasm and nucleus remains largely unknown. Here we report a novel function of transcription factor JunD in the regulation of HuR subcellular localization through the control of importin-α1 expression in intestinal epithelial cells (IECs). Ectopically expressed JunD specifically inhibited importin-α1 at the transcription level, and this repression is mediated via interaction with CREB-binding site that was located at the proximal region of importin-α1 promoter...
October 12, 2016: American Journal of Physiology. Cell Physiology
Jun Zhang, Kathleen L McCann, Chen Qiu, Lauren E Gonzalez, Susan J Baserga, Traci M Tanaka Hall
Numerous factors direct eukaryotic ribosome biogenesis, and defects in a single ribosome assembly factor may be lethal or produce tissue-specific human ribosomopathies. Pre-ribosomal RNAs (pre-rRNAs) must be processed stepwise and at the correct subcellular locations to produce the mature rRNAs. Nop9 is a conserved small ribosomal subunit biogenesis factor, essential in yeast. Here we report a 2.1-Å crystal structure of Nop9 and a small-angle X-ray-scattering model of a Nop9:RNA complex that reveals a 'C'-shaped fold formed from 11 Pumilio repeats...
October 11, 2016: Nature Communications
Jyan-Chyun Jang
Tandem CCCH zinc finger (TZF) proteins are evolutionarily conserved regulators of gene expression at the post-transcriptional level. TZFs target AU-rich RNA elements at 3' un-translated region and recruit catabolic machineries to trigger mRNA degradation. The plant TZF families are over-represented by a class of proteins with a unique TZF domain preceded by an arginine-rich motif (RR-TZF). RR-TZF proteins are mainly involved in hormone- and environmental cues-mediated plant growth and stress responses. Numerous reports have suggested that RR-TZF proteins control seed germination, plant size, flowering time, and biotic and abiotic stress responses via regulation of gene expression...
November 2016: Plant Science: An International Journal of Experimental Plant Biology
M-L Liu, K J Williams, V P Werth
During apoptosis or activation, cells can release a subcellular structure, called a membrane microvesicle (also known as microparticle) into the extracellular environment. Microvesicles bud-off as a portion of cell membrane with its associated proteins and lipids surrounding a cytosolic core that contains intracellular proteins, lipids, and nucleic acids (DNA, RNA, siRNA, microRNA, lncRNA). Biologically active molecules on the microvesicle surface and encapsulated within can act on recipient cells as a novel mode of intercellular communication...
2016: Advances in Clinical Chemistry
Ruslan Aphasizhev, Takuma Suematsu, Liye Zhang, Inna Aphasizheva
RNA uridylation is a significant transcriptome-shaping factor in protists, fungi, metazoans, and plants. The 3' U-additions are catalyzed by terminal uridyltransferases (TUTases), a diverse group of enzymes that along with non-canonical poly(A) polymerases form a distinct group in the superfamily of DNA polymerase β-like nucleotidyl transferases. Within and across studied organisms and subcellular compartments, TUTases differ in nucleotide triphosphate selectivity, interacting partners, and RNA targets. A general premise linking RNA uridylation to 3'-5' degradation received support from several studies of small RNAs and mRNA turnover...
October 7, 2016: RNA Biology
Thekkinghat Anantharaman Arvind, Pundi N Rangarajan
Mouse Apolipoprotein L9 (ApoL9) is an understudied cytoplasmic, interferon-inducible protein. The details of its intracellular localization and normal cellular functions are unclear. We report here that ApoL9 localizes to small puncta diffusely distributed in the cytoplasm, as well as to larger granules of varying size and number that are similar to aggresome-like induced structures (ALIS) and contain the autophagy receptor Sqstm1/p62, the autophagosome marker Lc3, and ubiquitin. Transfection of B16F10 mouse melanoma cells stably expressing ApoL9 (B16F10(L9)) with certain liposome-based transfection reagents causes dramatic disturbances in its subcellular distribution...
October 28, 2016: Biochemical and Biophysical Research Communications
Lucia Mendoza-Viveros, Pascale Bouchard-Cannon, Sara Hegazi, Arthur H Cheng, Stephen Pastore, Hai-Ying Mary Cheng
Circadian timekeeping is a ubiquitous mechanism that enables organisms to maintain temporal coordination between internal biological processes and time of the local environment. The molecular basis of circadian rhythms lies in a set of transcription-translation feedback loops (TTFLs) that drives the rhythmic transcription of core clock genes, whose level and phase of expression serve as the marker of circadian time. However, it has become increasingly evident that additional regulatory mechanisms impinge upon the TTFLs to govern the properties and behavior of the circadian clock...
September 29, 2016: Cellular and Molecular Life Sciences: CMLS
Stefanie J Kellermann, Andrea Rentmeister
Sequence-specific and programmable binding of proteins to RNA bears potential to detect and manipulate target RNAs. Applications include analysis of subcellular RNA localization or post-transcriptional regulation but require sequence-specificity to be readily adjustable to any target RNA. The Pumilio homology domain binds an eight nucleotide target sequence in a predictable manner allowing for rational design of variants with new specificities. We describe a high-throughput system for screening Pumilio variants based on fluorescenceactivated cell sorting of E...
September 28, 2016: Biological Chemistry
Z B Ruan, X L Fu, W Li, J Ye, R Z Wang, Y G Yin, L Zhu
Objective: To investigate the effects of Notch1, 2, 3 genes silencing by siRNA on Notch signaling pathway (Delta-like 4(DLL4), Jagged 1(JAG1)) and nuclear factor-κB (NF-κB) signaling pathway (IκBα, P52) of macrophages derived from patients with coronary artery disease (CAD), thus to explore the potential genetic treatment perspectives for CAD. Methods: Peripheral blood mononuclear cells of CAD patients were isolated by density gradient centrifugation and transformed by phorbol-12-myristate-13-acetate (PMA) to macrophages...
September 24, 2016: Zhonghua Xin Xue Guan Bing za Zhi
Cindy Meyer, Aitor Garzia, Thomas Tuschl
Fluorescence in situ hybridization (FISH) and immunofluorescence (IF) are sensitive techniques for detecting nucleic acids and proteins in cultured cells. However, these techniques are rarely applied together, and standard protocols are not readily compatible for sequential application on the same specimen. Here, we provide a user-friendly step-by-step protocol to perform multicolor RNA-FISH in combination with IF to simultaneously detect the subcellular localization of distinct RNAs and proteins in cultured cells...
September 21, 2016: Methods: a Companion to Methods in Enzymology
Thomas Conrad, Ulf Andersson Ørom
In eukaryotic cells, the synthesis, processing, and functions of RNA molecules are confined to distinct subcellular compartments. Biochemical fractionation of cells prior to RNA isolation thus enables the analysis of distinct steps in the lifetime of individual RNA molecules that would be masked in bulk RNA preparations from whole cells. Here, we describe a simple two-step differential centrifugation protocol for the isolation of cytoplasmic, nucleoplasmic, and chromatin-associated RNA that can be used in downstream applications such as qPCR or deep sequencing...
2017: Methods in Molecular Biology
Puxian Fang, Liurong Fang, Xiaorong Liu, Yingying Hong, Yongle Wang, Nan Dong, Panpan Ma, Jing Bi, Dang Wang, Shaobo Xiao
Porcine deltacoronavirus (PDCoV) is an emerging swine enteric coronavirus. Accessory proteins are genus-specific for coronavirus, and two putative accessory proteins, NS6 and NS7, are predicted to be encoded by PDCoV; however, this remains to be confirmed experimentally. Here, we identified the leader-body junction sites of NS6 subgenomic RNA (sgRNA) and found that the actual transcription regulatory sequence (TRS) utilized by NS6 is non-canonical and is located upstream of the predicted TRS. Using the purified NS6 from an Escherichia coli expression system, we obtained two anti-NS6 monoclonal antibodies that could detect the predicted NS6 in cells infected with PDCoV or transfected with NS6-expressing plasmids...
September 20, 2016: Virology
Lorey K Smith, Daniel W Thomas, Kaylene J Simpson, Patrick O Humbert
Correct subcellular localization of proteins is a requirement for appropriate function. This is especially true in epithelial cells, which rely on the precise localization of a diverse array of epithelial polarity and cellular adhesion proteins. Loss of cell polarity and adhesion is a hallmark of cancer, and mislocalization of core polarity proteins, such as Scribble, is observed in a range of human epithelial tumors and is prognostic of poor survival. Despite this, little is known about how Scribble membrane localization is regulated...
October 2016: Assay and Drug Development Technologies
Huan-Lei Wu, Sen-Mao Li, Jia Hu, Xiao Yu, Hua Xu, Zhong Chen, Zhang-Qun Ye
BACKGROUND: The recently identified phenomenon of double-stranded RNA (dsRNA)-mediated gene activation (RNAa) has been studied extensively, as it is present in humans, mice, and Caenorhabditis elegans, suggesting that dsRNA-mediated RNAa is an evolutionarily conserved mechanism. Previous studies have shown that dsP21-322 can induce tumor suppressor gene p21 expression in several human cancer cells. Nonetheless, the role of dsRNAs in the activation of gene expression, including their target molecules and associated key factors, remains poorly understood...
2016: Journal of Experimental & Clinical Cancer Research: CR
Jiafeng Li, Jinsheng Zhao, Yue Li
Overexpression of the nucleoporin NUP88 has been observed in a large number of tumors and has been experimentally proven to promote tumorigenesis. However, the mechanism underlying the tumor-promoting activity of overexpressed NUP88 is not clear. To investigate the potential pathways that drive tumorigenesis under NUP88 overexpressed condition, we applied a proteomic approach to identify NUP88-associated proteins at a subcellular compartment level. Gene ontology analysis revealed significant associations between NUP88 interactome and biological processes that are related to nuclear transport, RNA processing, cell cycle progression, metabolic regulation, and viral infection...
September 16, 2016: Genes, Chromosomes & Cancer
Seyed Jalal Kiani, Tahereh Taheri, Sima Rafati, Katayoun Samimi-Rad
RNA-binding proteins play critical roles in the regulation of gene expression. Among several families of RNA-binding proteins, PUF (Pumilio and FBF) proteins have been the subject of extensive investigations, as they can bind RNA in a sequence-specific manner and they are evolutionarily conserved among a wide range of organisms. The outstanding feature of these proteins is a highly conserved RNA-binding domain, which is known as the Pumilio-homology domain (PUM-HD) that mostly consists of eight tandem repeats...
September 14, 2016: Current Protein & Peptide Science
Anne Plochowietz, Ian Farrell, Zeev Smilansky, Barry S Cooperman, Achillefs N Kapanidis
Transfer RNA (tRNA) links messenger RNA nucleotide sequence with amino acid sequence during protein synthesis. Despite the importance of tRNA for translation, its subcellular distribution and diffusion properties in live cells are poorly understood. Here, we provide the first direct report on tRNA diffusion localization in live bacteria. We internalized tRNA labeled with organic fluorophores into live bacteria, applied single-molecule fluorescence imaging with single-particle tracking and localized and tracked single tRNA molecules over seconds...
September 12, 2016: Nucleic Acids Research
Moon Y F Tay, Kate Smith, Ivan H W Ng, Kitti W K Chan, Yongqian Zhao, Eng Eong Ooi, Julien Lescar, Dahai Luo, David A Jans, Jade K Forwood, Subhash G Vasudevan
Dengue virus NS5 is the most highly conserved amongst the viral non-structural proteins and is responsible for capping, methylation and replication of the flavivirus RNA genome. Interactions of NS5 with host proteins also modulate host immune responses. Although replication occurs in the cytoplasm, an unusual characteristic of DENV2 NS5 is that it localizes to the nucleus during infection with no clear role in replication or pathogenesis. We examined NS5 of DENV1 and 2, which exhibit the most prominent difference in nuclear localization, employing a combination of functional and structural analyses...
September 2016: PLoS Pathogens
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