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Hayase Mizukami, Jun-Dal Kim, Saori Tabara, Weizhe Lu, Chulwon Kwon, Misaki Nakashima, Akiyoshi Fukamizu
Males and females share the same genetic code, but gene expression profile often displays differences between two sexes. Mouse embryonic fibroblasts (MEFs) have been used to experiment as a useful tool to test gene function. They have also been characterized by gender-based differences in expressed genes such as Y-linked Sry or X-linked Hprt. However, there is no report on sex differences in global gene expression. Here, using the next-generation RNA sequencing, we compared the comprehensive transcriptome of MEFs derived from two sexes...
December 12, 2018: Journal of Biochemistry
Jik Nijssen, Julio Aguila, Rein Hoogstraaten, Nigel Kee, Eva Hedlund
Spinal motor axons traverse large distances to innervate target muscles, thus requiring local control of cellular events for proper functioning. To interrogate axon-specific processes we developed Axon-seq, a refined method incorporating microfluidics, RNA sequencing (RNA-seq), and bioinformatic quality control. We show that the axonal transcriptome is distinct from that of somas and contains fewer genes. We identified 3,500-5,000 transcripts in mouse and human stem cell-derived spinal motor axons, most of which are required for oxidative energy production and ribogenesis...
December 11, 2018: Stem Cell Reports
Mati Mann, Arnav Mehta, Carl G de Boer, Monika S Kowalczyk, Kevin Lee, Pearce Haldeman, Noga Rogel, Abigail R Knecht, Daneyal Farouq, Aviv Regev, David Baltimore
Long-term hematopoietic stem cells (LT-HSCs) maintain hematopoietic output throughout an animal's lifespan. However, with age, the balance is disrupted, and LT-HSCs produce a myeloid-biased output, resulting in poor immune responses to infectious challenge and the development of myeloid leukemias. Here, we show that young and aged LT-HSCs respond differently to inflammatory stress, such that aged LT-HSCs produce a cell-intrinsic, myeloid-biased expression program. Using single-cell RNA sequencing (scRNA-seq), we identify a myeloid-biased subset within the LT-HSC population (mLT-HSCs) that is prevalent among aged LT-HSCs...
December 11, 2018: Cell Reports
Rosa Martin-Mateos, Thiago M De Assuncao, Juan Pablo Arab, Nidhi Jalan-Sakrikar, Usman Yaqoob, Thomas Greuter, Vikas K Verma, Angela J Mathison, Sheng Cao, Gwen Lomberk, Philippe Mathurin, Raul Urrutia, Robert C Huebert, Vijay H Shah
BACKGROUND & AIMS: Transdifferentiation of hepatic stellate cells (HSCs) into myofibroblasts is a key event in the pathogenesis of liver fibrosis. Transforming growth factor β (TGF-β) and platelet-derived growth factor (PDGF) are canonical HSC activators after liver injury. The aim of this study was to analyze the epigenetic modulators that differentially control TGF-β and PDGF signaling pathways. METHODS: We performed a transcriptomic comparison of HSCs treated with TGF-β or PDGF-BB using RNA sequencing...
September 15, 2018: Cellular and Molecular Gastroenterology and Hepatology
Rui Dong, Xu-Kai Ma, Ling-Ling Chen, Li Yang
Circular RNAs (circRNAs) derived from back-spliced exons were sporadically identified about 25 years ago, and have been recently re-discovered genome-wide across different species. Interestingly, one gene locus can generate multiple circRNAs through alternative back-splicing and/or alternative splicing, thus expanding our understanding on the diversity and complexity of transcriptomes. Precise annotation of circRNAs with their alternative back-splicing and alternative splicing events is the basis for the functional characterization of different categories of circRNAs...
2019: Methods in Molecular Biology
Lee E Vandivier, Zachary D Anderson, Brian D Gregory
Ribonucleotides can be decorated with over 100 types of covalent chemical modifications. These modifications change the structure, function, and catalytic activity of RNAs, forming a layer of posttranscriptional regulation termed the epitranscriptome. Recent advances in high-throughput mapping have demonstrated these modifications are abundant and mark nearly all classes of RNAs, including messenger RNAs. Here, we outline one such technique called high-throughput annotation of modified ribonucleotides (HAMR)...
2019: Methods in Molecular Biology
Mingming Chen, Shiqi Mao, Xiaotian Wu, Zhao Ma, Yantao Yang, Christopher J Krueger, Antony K Chen
Over the past decade, emerging evidence has indicated that long intergenic noncoding RNAs (lincRNAs), a class of RNA transcripts greater than 200 nt in length, function as key regulators of gene expression in cellular physiology and pathogenesis. Greater understanding of lincRNA activities, particularly in the context of subcellular localization and dynamic regulation at the single-molecule level, is expected to provide in-depth understanding of molecular mechanisms that regulate cell behavior and disease evolution...
2019: Methods in Molecular Biology
Jenna J Guthmiller, Haley L Dugan, Karlynn E Neu, Linda Yu-Ling Lan, Patrick C Wilson
In the age of personalized medicine, an efficient method to generate monoclonal antibodies (mAbs) is essential for biomedical and immunotherapeutic research. Numerous aspects of basic B-cell biology can be studied at the monoclonal level, including B-cell development, antibody responses to infection or vaccination, and autoimmune responses. Single-cell B-cell receptor cloning allows for the rapid generation of antigen-specific mAbs in a matter of several weeks. In this chapter, we provide an efficient method to generate mAbs from peripheral blood plasmablasts and memory B cells induced by infection and vaccination...
2019: Methods in Molecular Biology
Stefan C Weise, Ganeshkumar Arumugam, Alejandro Villarreal, Pavankumar Videm, Stefanie Heidrich, Nils Nebel, Verónica I Dumit, Farahnaz Sananbenesi, Viktoria Reimann, Madeline Craske, Oliver Schilling, Wolfgang R Hess, Andre Fischer, Rolf Backofen, Tanja Vogel
Rett syndrome is a complex neurodevelopmental disorder that is mainly caused by mutations in MECP2. However, mutations in FOXG1 cause a less frequent form of atypical Rett syndrome, called FOXG1 syndrome. FOXG1 is a key transcription factor crucial for forebrain development, where it maintains the balance between progenitor proliferation and neuronal differentiation. Using genome-wide small RNA sequencing and quantitative proteomics, we identified that FOXG1 affects the biogenesis of miR200b/a/429 and interacts with the ATP-dependent RNA helicase, DDX5/p68...
December 11, 2018: Molecular Neurobiology
Jussa-Pekka Virtanen, Riikka Keto-Timonen, Kaisa Jaakkola, Noora Salin, Hannu Korkeala
Yersinia pseudotuberculosis is a bacterium that not only survives, but also thrives, proliferates, and remains infective at cold-storage temperatures, making it an adept foodborne pathogen. We analyzed the differences in gene expression between Y. pseudotuberculosis IP32953 grown at 3 and 28°C to investigate which genes were significantly more expressed at low temperature at different phases of growth. We isolated and sequenced the RNA from six distinct corresponding growth points at both temperatures to also outline the expression patterns of the differentially expressed genes...
2018: Frontiers in Cellular and Infection Microbiology
Yong Zhou, Chengzhi Gu, Jia Li, Lianhai Zhu, Guoxiang Huang, Jie Dai, Huaiyu Huang
Purpose: Parkinson's disease (PD) is a common neurodegenerative movement disorder, but the pathogenesis remains elusive. This study was aimed to explore key genes and long noncoding RNAs (lncRNAs) associated with PD. Materials and methods: Three patients with PD and three normal controls were enrolled in the present study from July 12, 2017, to August 29, 2017. RNA sequencing and bioinformatics analysis were performed to obtain differentially expressed micro RNAs (DEmRNAs) and lncRNAs (DElncRNAs) between patients with PD and normal controls...
2018: Neuropsychiatric Disease and Treatment
Anjana Bhardwaj, Harpreet Singh, Celestine Marie Trinidad, Constance T Albarracin, Kelly K Hunt, Isabelle Bedrosian
BACKGROUND: Prevention of triple-negative breast cancer (TNBC) is hampered by lack of knowledge about the drivers of tumorigenesis. METHODS: To identify molecular markers and their downstream networks that can potentially be targeted for TNBC prevention, we analyzed small RNA and RNA sequencing of a cell line model that represent early stages of TNBC development. We have identified direct gene targets of isomiRNA-140-3p and by using cell-based and in vivo model systems we have demonstrated the utility of targeting downstream pathways for prevention of TNBC...
December 11, 2018: Breast Cancer Research: BCR
Abigail F Groff, A Rasim Barutcu, Jordan P Lewandowski, John L Rinn
BACKGROUND: Recently, it has become clear that some promoters function as long-range regulators of gene expression. However, direct and quantitative assessment of enhancer activity at long intergenic noncoding RNA (lincRNA) or mRNA gene bodies has not been performed. To unbiasedly assess the enhancer capacity across lincRNA and mRNA loci, we performed a massively parallel reporter assay (MPRA) on six lincRNA loci and their closest protein-coding neighbors. RESULTS: For both gene classes, we find significantly more MPRA activity in promoter regions than in gene bodies...
December 11, 2018: Genome Biology
Qianqian Dong, Luye Shi, Yangwei Li, Mengwan Jiang, Hong Sun, Baishi Wang, Han Cheng, Yifeng Zhang, Tian Shao, Yuhua Shi, Zhenlong Wang
BACKGROUND: Subterranean rodents have evolved many features to adapt to their hypoxic environment. The brain is an organ that is particularly vulnerable to damage caused by exposure to hypoxic conditions. To investigate the mechanisms of adaption to a hypoxic underground environment, we carried out a cross-species brain transcriptome analysis by RNA sequencing and identified genes that are differentially expressed between the subterranean vole Lasiopodomys mandarinus and the closely related above-ground species Lasiopodomys brandtii under chronic hypoxia [10...
December 11, 2018: BMC Genomics
Antonio Neme, Sabine Seuter, Marjo Malinen, Tarja Nurmi, Tomi-Pekka Tuomainen, Jyrki K Virtanen, Carsten Carlberg
In the vitamin D intervention study VitDbol (NCT02063334) blood samples were drawn directly before an oral bolus (2000 µg vitamin D3 ) and 24 h later. The focus of phase II of VitDbol was the transcriptome-wide analysis of the effects of vitamin D gene expression in human peripheral blood mononuclear cells (PBMCs). All five participants responded in an individual fashion to the bolus by increases in serum levels of the vitamin D metabolites 25-hydroxyvitamin D3 (25(OH)D3 ) and 1α,25-dihydroxyvitamin D3 (1,25(OH)2 D3 )...
December 8, 2018: Journal of Steroid Biochemistry and Molecular Biology
Jianting Shi, Chenyi Xue, Wen Liu, Hanrui Zhang
Macrophages play important roles in many diseases. We describe a protocol and the associated resources for the differentiation of human induced pluripotent stem cell-derived macrophages (IPSDM) and their applications in understanding human macrophage physiology and relevant diseases. The protocol uses an embryoid body-based approach with a combination of serum-free condition for hematopoiesis specification, followed by adherent culture with serum and M-CSF for myeloid expansion and macrophage maturation. The protocol produced an almost pure culture of CD45+ /CD18+ macrophages yielding up to 2 × 107 cells per 6-well plate of iPSCs within 24 days, demonstrating high efficiency, purity, and scalability...
December 10, 2018: Current Protocols in Stem Cell Biology
Qingyao Zhu, Hongjie Yang, Peng Cheng, Qian Han
Some lncRNAs can encode small nucleolar RNAs (snoRNAs), called small nucleolar RNA host genes (SNHGs), which exert diverse regulatory effects on cellular processes. In this study, using RNA-seq and survival data in the Cancer Genome Atlas (TCGA)-Liver Hepatocellular Carcinoma (LIHC), we examined the expression profile of some SNHG genes and explored their prognostic value in hepatocellular carcinoma (HCC). Level-3 RNA-sequencing data, the clinicopathological and survival data of patients with primary HCC were downloaded from the UCSC Xena browser (https://xenabrowser...
December 8, 2018: BioFactors
Julien J Karrich, Mónica Romera-Hernández, Natalie Papazian, Sharon Veenbergen, Ferry Cornelissen, Patricia Aparicio-Domingo, Frances H Stenhouse, C Diana Peddie, Remco M Hoogenboezem, Chelsea W J den Hollander, Terri Gaskell, Tanya Medley, Louis Boon, C Claire Blackburn, David R Withers, Janneke N Samsom, Tom Cupedo
Under homeostatic conditions, dendritic cells (DCs) continuously patrol the intestinal lamina propria. Upon antigen encounter, DCs initiate C-C motif chemokine receptor 7 (CCR7) expression and migrate into lymph nodes to direct T cell activation and differentiation. The mechanistic underpinnings of DC migration from the tissues to lymph nodes have been largely elucidated, contributing greatly to our understanding of DC functionality and intestinal immunity. In contrast, the molecular mechanisms allowing DCs to efficiently migrate through the complex extracellular matrix of the intestinal lamina propria prior to antigen encounter are still incompletely understood...
December 7, 2018: European Journal of Immunology
Houqing Zeng, Xin Zhang, Ming Ding, Xiajun Zhang, Yiyong Zhu
Zinc (Zn) deficiency is a widespread agricultural problem in arable soils of the whole world. However, the molecular mechanisms underlying Zn deficiency response are largely unknown. Here, we analyzed the transcriptomic profilings of soybean leaves and roots in response to Zn deficiency through Illumina's high-throughput RNA sequencing in order to understand the molecular basis of Zn deficiency response in the plants. A total of 614 and 1011 gene loci were found to be differentially expressed in leaves and roots, respectively, and 88 loci were commonly found in both leaves and roots...
December 10, 2018: Physiologia Plantarum
Yong Jung, Yasser El-Manzalawy, Drena Dobbs, Vasant Honavar
RNA-protein interactions play essential roles in regulating gene expression. While some RNA-protein interactions are "specific", i.e., the RNA-binding proteins preferentially bind to particular RNA sequence or structural motifs, others are "non-RNA specific." Deciphering the protein-RNA recognition code is essential for comprehending the functional implications of these interactions and for developing new therapies for many diseases. Because of the high cost of experimental determination of protein-RNA interfaces, there is a need for computational methods to identify RNA-binding residues in proteins...
December 11, 2018: Proteins
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