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| Status |
Public on Jun 01, 2008 |
| Title |
Transcription elongation factor ELL2 influences splicing versus poly(A) site choice in the Ig heavy chain gene |
| Organism |
Mus musculus |
| Experiment type |
Expression profiling by array
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| Summary |
Processing of Immunoglobulin heavy chain (IgH) mRNA is a paradigm for competition between splicing and polyadenylation. In plasma cells pre-mRNA is polyadenylated mainly at the promoter-proximal secretory site while B-cells utilize a cryptic 5’ splice site in the last secretory-specific exon; these are mutually exclusive events for all IgH pre-mRNAs. Transcription elongation factor ELL2, more abundant in plasma cells relative to B-cells, was down-modulated by overexpression of heterogenous ribonucleoprotein F, a condition which reduced production of secretory IgH mRNA. Transfection of B-cells with ELL2 and the IgH reporter showed an accelerated use of the secretory poly(A) site, positioned in competition with the splice to M1; a small interfering RNA to ELL2 reduced expression of IgH secretory mRNA. Co-transcription factors ELL1 and PC4 were ineffective at driving secretory-poly(A) site use. ELL2 had little effect on poly(A) site choice with reporters containing tandem-linked poly(A) sites. Shorter forms of ELL2 protein result from both internal initiation at M186 and protein processing. An alternative splicing reporter driven by IgH or non-Ig promoters revealed that ELL2 and its M186 initiated form were able to accelerate exon skipping. Therefore, ELL2 influences IgH pre-mRNA processing through facilitating skipping of the alternative splice to the membrane form.
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| Overall design |
AxJ plasma cells were stably transfected to overexpress hnRNP-F, -H or empty vector. Clones showing high overexpression levels of F or H by western blot were selected. The IgH sec to mb ratios of these clones were determined. A global gene expression analysis was performed on mRNA from two clones from hnRNP-F, which demonstrated a lower sec:mb ratio, and one from each of the controls: overexpression of hnRNP-H, or transfection with empty vector, or A20 B-cells, using Affymetrix gene micro array technology.
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| Contributor(s) |
Martincic K, Alkan SA, Cheatle A, Milcarek C |
| Citation(s) |
19749764 |
| Submission date |
May 28, 2008 |
| Last update date |
Feb 18, 2018 |
| Contact name |
kathleen martincic |
| E-mail(s) |
kathee@pitt.edu
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| Organization name |
University of Pittsburgh
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| Department |
Immunology
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| Lab |
Dr. Christine Milcarek
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| Street address |
200 Lothrop Street
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| City |
Pittsburgh |
| State/province |
PA |
| ZIP/Postal code |
15261 |
| Country |
USA |
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| Platforms (1) |
| GPL81 |
[MG_U74Av2] Affymetrix Murine Genome U74A Version 2 Array |
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| Samples (5)
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| Relations |
| BioProject |
PRJNA106219 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE11586_RAW.tar |
9.8 Mb |
(http)(custom) |
TAR (of CEL, CHP) |
| Processed data included within Sample table |
| Processed data provided as supplementary file |
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