Browsing by Subject "LARGE GENE LISTS"

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  • Conesa, Ana; Madrigal, Pedro; Tarazona, Sonia; Gomez-Cabrero, David; Cervera Taboada, Alejandra; McPherson, Andrew; Szczesniak, Michal Wojciech; Gaffney, Daniel J.; Elo, Laura L.; Zhang, Xuegong; Mortazavi, Ali (2016)
    RNA-sequencing (RNA-seq) has a wide variety of applications, but no single analysis pipeline can be used in all cases. We review all of the major steps in RNA-seq data analysis, including experimental design, quality control, read alignment, quantification of gene and transcript levels, visualization, differential gene expression, alternative splicing, functional analysis, gene fusion detection and eQTL mapping. We highlight the challenges associated with each step. We discuss the analysis of small RNAs and the integration of RNA-seq with other functional genomics techniques. Finally, we discuss the outlook for novel technologies that are changing the state of the art in transcriptomics.
  • Tolkachov, Alexander; Fischer, Cornelius; Ambrosi, Thomas H.; Bothe, Melissa; Han, Chung-Ting; Muenzner, Matthias; Mathia, Susanne; Salminen, Marjo; Seifert, Georg; Thiele, Mario; Duda, Georg N.; Meijsing, Sebastiaan H.; Sauer, Sascha; Schulz, Tim J.; Schupp, Michael (2018)
    The transcription factor GATA2 is required for expansion and differentiation of hematopoietic stem cells (HSCs). In mesenchymal stem cells (MSCs), GATA2 blocks adipogenesis, but its biological relevance and underlying genomic events are unknown. We report a dual function of GATA2 in bone homeostasis. GATA2 in MSCs binds near genes involved in skeletal system development and colocalizes with motifs for FOX and HOX transcription factors, known regulators of skeletal development. Ectopic GATA2 blocks osteoblastogenesis by interfering with SMAD1/5/8 activation. MSC-specific deletion of GATA2 in mice increases the numbers and differentiation capacity of bone-derived precursors, resulting in elevated bone formation. Surprisingly, MSC-specific GATA2 deficiency impairs the trabecularization and mechanical strength of bone, involving reduced MSC expression of the osteoclast inhibitor osteoprotegerin and increased osteoclast numbers. Thus, GATA2 affects bone turnover via MSC-autonomous and indirect effects. By regulating bone trabecularization, GATA2 expression in the osteogenic lineage may contribute to the anatomical and cellular microenvironment of the HSC niche required for hematopoiesis.
  • Lohmus, Andres; Varjosalo, Markku; Mäkinen, Kristiina (2016)
    The definition of the precise molecular composition of membranous replication compartments is a key to understanding the mechanisms of virus multiplication. Here, we set out to investigate the protein composition of the potyviral replication complexes. We purified the potyviral 6K2 protein-induced membranous structures from Potato virus A (PVA)-infected Nicotiana benthamiana plants. For this purpose, the 6K2 protein, which is the main inducer of potyviral membrane rearrangements, was expressed in fusion with an N-terminal Twin-Strep-tag and Cerulean fluorescent protein (SC6K) from the infectious PVA cDNA. A non-tagged Cerulean-6K2 (C6K) virus and the SC6K protein alone in the absence of infection were used as controls. A purification scheme exploiting discontinuous sucrose gradient centrifugation followed by Strep-tag-based affinity chromatography was developed. Both (+)- and (-)-strand PVA RNA and viral protein VPg were co-purified specifically with the affinity tagged PVA-SC6K. The purified samples, which contained individual vesicles and membrane clusters, were subjected to mass spectrometry analysis. Data analysis revealed that many of the detected viral and host proteins were either significantly enriched or fully specifically present in PVA-SC6K samples when compared with the controls. Eight of eleven potyviral proteins were identified with high confidence from the purified membrane structures formed during PVA infection. Ribosomal proteins were identified from the 6K2-induced membranes only in the presence of a replicating virus, reinforcing the tight coupling between replication and translation. A substantial number of proteins associating with chloroplasts and several host proteins previously linked with potyvirus replication complexes were co-purified with PVA-derived SC6K, supporting the conclusion that the host proteins identified in this study may have relevance in PVA replication.
  • Fagerholm, Rainer; Schmidt, Marjanka K.; Khan, Sofia; Rafiq, Sajjad; Tapper, William; Aittomaki, Kristiina; Greco, Dario; Heikkinen, Tuomas; Muranen, Taru A.; Fasching, Peter A.; Janni, Wolfgang; Weinshilboum, Richard; Loehberg, Christian R.; Hopper, John L.; Southey, Melissa C.; Keeman, Renske; Lindblom, Annika; Margolin, Sara; Mannermaa, Arto; Kataja, Vesa; Chenevix-Trench, Georgia; Lambrechts, Diether; Wildiers, Hans; Chang-Claude, Jenny; Seibold, Petra; Couch, Fergus J.; Olson, Janet E.; Andrulis, Irene L.; Knight, Julia A.; Garcia-Closas, Montserrat; Figueroa, Jonine; Hooning, Maartje J.; Jager, Agnes; Shah, Mitul; Perkins, Barbara J.; Luben, Robert; Hamann, Ute; Kabisch, Maria; Czene, Kamila; Hall, Per; Easton, Douglas F.; Pharoah, Paul D. P.; Liu, Jianjun; Eccles, Diana; Blomqvist, Carl; Nevanlinna, Heli; kConFab Investigators (2015)
    We have utilized a two-stage study design to search for SNPs associated with the survival of breast cancer patients treated with adjuvant chemotherapy. Our initial GWS data set consisted of 805 Finnish breast cancer cases (360 treated with adjuvant chemotherapy). The top 39 SNPs from this stage were analyzed in three independent data sets: iCOGS (n=6720 chemotherapy-treated cases), SUCCESS-A (n=3596), and POSH (n=518). Two SNPs were successfully validated: rs6500843 (any chemotherapy; per-allele HR 1.16, 95% C.I. 1.08-1.26, p=0.0001, p((adjusted))=0.0091), and rs11155012 (anthracycline therapy; per-allele HR 1.21, 95% C.I. 1.08-1.35, p=0.0010, p((adjusted))=0.0270). The SNP rs6500843 was found to specifically interact with adjuvant chemotherapy, independently of standard prognostic markers (p((interaction))=0.0009), with the rs6500843-GG genotype corresponding to the highest hazard among chemotherapy-treated cases (HR 1.47, 95% C.I. 1.20-1.80). Upon trans-eQTL analysis of public microarray data, the rs6500843 locus was found to associate with the expression of a group of genes involved in cell cycle control, notably AURKA, the expression of which also exhibited differential prognostic value between chemotherapy-treated and untreated cases in our analysis of microarray data. Based on previously published information, we propose that the eQTL genes may be connected to the rs6500843 locus via a RBFOX1-FOXM1 -mediated regulatory pathway.