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Series GSE117384 Query DataSets for GSE117384
Status Public on Oct 01, 2018
Title Comparison of PRPF4 knockdown in HEK293T cells compared to scrambled shRNA control
Organism Homo sapiens
Experiment type Expression profiling by array
Summary HEK293T cells transduced with two shRNAs from MISSION library (TRCN0000364755 and TRCN0000074769) using lentiviral delivery system. HEK293T cells transduced with scrambled shRNA, gifted from Dr. Mauricio Reginato. Sequence is 5′-CCTAAGGTTAAGTCGCCCTCGCTCTAGCGAGGGCGACTTAACCTT-3′. Primary contributions to creation of cell lines, preparation of RNA/cDNA for microarray, and validation of results were from Tara L Davis, S. RaElle Jackson, Beth Adams, Anh Trinh, Nicholas Fox, Jessica Kopenhaver, Alyson Hurlock, and Achraf Jardaly, all Drexel University College of Medicine, Philadephia PA, 19102. Hetty Rodriguez and John Tobias, affiliated with the Molecular Profiling Facility and Genomic Analysis Core Bioinformatics Group at the University of Pennsylvania, Philadelphia PA, performed Bioanalyzer and microarray expreriments and initial data processing.
pre-mRNA Processing Factor 4 (PRPF4) is a scaffolding protein, consisting of two N-terminal intrinsically disordered motifs and a C-terminal WD-40 domain. PRPF4 associates with the human spliceosome, the complex and dynamic machinery that removes intronic sequence from pre-messenger RNA (pre-mRNA). Although PRPF4 affects the ability of the spliceosome to assemble and catalyze splicing in vitro through de-stabilization of the tri-snRNP complex, little is known about what PRPF4 does to regulate transcription and splicing in vivo. To understand the function of PRPF4 in the nucleus, we knocked down PRPF4 in human cells. We characterized a set of alternative splicing and transcriptional events that are PRPF4-responsive. We used these splicing and transcriptional bioassays to show that PRPF4-responsive events are largely specific. The development of a bioassay for PRPF4 function can be used to answer fundamental questions about the role of spliceosomal proteins in regulating splicing and other nuclear functions.
Overall design 3 replicates of PRPF4 knockdown cell lines in HEK293T cells, stably integrated, selected using puro, shRNAs TRCN0000364755 sequence 5'-CCGGCCACGAACTGTGTAGACATTGCTCGAGCAATGTCTACACAGTTCGTGGTTTTTG-3' and TRCN0000074769 sequence 5'-CCGGGCTCTCTCTAAGGAGCTGTTTCTCGAGAAACAGCTCCTTAGAGAGAGCTTTTTG-3'. 3 replicates of SCR control cell lines in HEK293T cells, stably integrated, selected using puro, sequence 5′-CCTAAGGTTAAGTCGCCCTCGCTCTAGCGAGGGCGACTTAACCTT-3′. Total RNA purified from ~1 million cells, cDNA converted using random hexamer primers, quantity measured using NanoDrop, quality quantified using BioAnaylzer). Affymatrix HTA2.0 array
Contributor(s) Davis TL, Jackson SR, Adams B, Trinh A, Fox N, Kopenhaver J, Hurlock A, Jardaly A, Rodriguez H, Tobias J
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NIH grant(s)
Grant ID Grant title Affiliation Name
R00 GM094293 Functional and structural characterization of spliceosomal cyclophilins DREXEL UNIVERSITY Tara L Davis
Submission date Jul 19, 2018
Last update date Oct 29, 2018
Contact name Tara Davis
Organization name Drexel University College of Medicine
Department Biochemistry and Molecular Biology
Lab Lab 10127
Street address 245 N. 15th St. MS 497
City Philadelphia
State/province PA
ZIP/Postal code 19102
Country USA
Platforms (2)
GPL17585 [HTA-2_0] Affymetrix Human Transcriptome Array 2.0 [probe set (exon) version]
GPL17586 [HTA-2_0] Affymetrix Human Transcriptome Array 2.0 [transcript (gene) version]
Samples (12)
GSM3293639 HEK293T_755+769PRPF4_gene_rep1
GSM3293640 HEK293T_755+769PRPF4_gene_rep2
GSM3293641 HEK293T_755+769PRPF4_gene_rep3
BioProject PRJNA481964

Download family Format
SOFT formatted family file(s) SOFTHelp
MINiML formatted family file(s) MINiMLHelp
Series Matrix File(s) TXTHelp

Supplementary file Size Download File type/resource
GSE117384_RAW.tar 392.5 Mb (http)(custom) TAR (of CEL, CHP)
Processed data included within Sample table
Processed data provided as supplementary file

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