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Series GSE12239 Query DataSets for GSE12239
Status Public on Feb 28, 2010
Title RIP-chip analysis of Tino/hMEX-3D mRNPs
Organism Homo sapiens
Experiment type Other
Summary Tino is an A+U-Rich Element (ARE) binding protein first identified through its ability to bind to bcl-2 mRNA and to contribute to its degradation. It has recently been recognized as a shorter form of the human Mex-3D protein (hMex-3D), one of the four members of the family of Mex-3 RNA-binding phosphoproteins. In C. elegans, ceMex-3 is a translational regulator that plays a key role in early embryonic development and in the maintenance of worm germ line totipotency. To examine the potential functional conservation between ceMex-3 and hMex3, we have used complementary microarray-based approaches to identify mRNAs directly bound to Tino/hMex-3D. Computational analysis of these target mRNAs resulted in the identification of an U-rich, 34- to 39-nucleotide long, consensus, forming loops of variable sizes. Remarkably, more than half of Tino/hMex-3D targets also contain the consensus for Quaking, which is the human ortholog of GLD-1, a regulator of nematode gametogenesis. All together, our results suggest that Tino/hMex-3D belongs to a regulatory circuit of mRNA trans-acting factors involved in cell fate and differentiation.

Keywords: RIP-chip and (recombinant)RIP-Chip analysis of Tino/hMEX-3D mRNPs
Overall design To identify the transcripts directly bound in vivo by Tino/hMex-3D, we have used two complementary strategies. First, using Tino-his transfected HEK293 cells, we have analysed the RNAs immunoprecipitated by Tino/hMex-3D (RNA-IP complexes). Second, we have developed an in vitro nitrocellulose RNA-protein binding assay using a truncated form of Tino, TinoΔRING-his. First for the in vivo approach, we have transiently transfected HEK293 cells with a His-tagged Tino expressing construct (see Materials) and isolated Tino target mRNAs by immunoprecipitation (IP) assays carried out under conditions preserving mRNA-protein complex integrity. Second, for the in vitro approach, cRNAs were prepared from a human placenta cDNA library and incubated with TinoΔRING-his. This deleted form of Tino-his protein conserves its mRNA binding activity. mRNAs directly bound to TinoΔRING-his protein were subsequently isolated using a nitrocellulose RNA-protein binding assay.

A supplementary file of ordered genes based on z-ratios, derived from experimental and negative control samples for each Affymetrix probe, is appended below.
Contributor(s) Minardi S, Venturini E, Lulli M, Donnini M, Lapucci A
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Submission date Jul 24, 2008
Last update date Dec 06, 2018
Contact name martino donnini
Phone 00390554598243
Fax 00394598900
Organization name University of Florence
Department Pathology and Oncology
Lab Sergio Capaccioli
Street address viale GB Morgagni
City Firenze
State/province Firenze
ZIP/Postal code 50134
Country Italy
Platforms (1)
GPL571 [HG-U133A_2] Affymetrix Human Genome U133A 2.0 Array
Samples (4)
GSM307601 Tino-his HEK293 overespressing
GSM307602 mock HEK293 overexpressing
GSM307603 TinoDeltaRING-his Filter Binding
BioProject PRJNA113699

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
GSE12239_RAW.tar 9.2 Mb (http)(custom) TAR (of CEL)
GSE12239_z_ratio_data.txt 1.2 Mb (ftp)(http) TXT
Processed data included within Sample table
Processed data are available on Series record

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