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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Jan 21, 2011 |
| Title |
An ENU-induced point mutation in the mouse Btaf1 gene causes post-gastrulation embryonic lethality and protein instability |
| Organism |
Mus musculus |
| Experiment type |
Expression profiling by array
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| Summary |
The mouse Btaf1 gene, an ortholog of yeast MOT1, encodes a highly conserved general transcription factor. The function of this SNF-2 like ATPase has been studied mainly in yeast and human cells, which has revealed that it binds directly to TBP, forming the B-TFIID complex. This complex binds to core promoters of RNA polymerase II- transcribed genes and, of crucial importance, BTAF1-TBP interactions have been shown to affect the kinetics of TBP-promoter interactions. Here we report the isolation of a mouse line carrying a Btaf1 allele containing an ENU-induced point mutation that causes a substitution mutation in the BTAF1 ATPase domain. Embryos homozygous for this loss-of-function mutation appear to be morphologically normal until early somite stages, but die between embryonic day 9 and 10.5 displaying growth arrest and edema. Analyses in vitro suggest that the altered protein is less stable and, independent from this, functionally impaired in releasing of TBP from chromatin, but not in binding to TBP.
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| Overall design |
A microarray screen for genes with altered expression in the Btaf V1330M mutant as compared to WT embryos was performed with RNA from whole embryos. We compared mRNA transcripts from pools of Btaf1-/- and Btaf1+/+ embryos. Where possible, we used 3-5-somite embryos, a stage we considered to be immediately prior to the emergence of a perceptible phenotype. The experiments were done in duplo, i.e. two sets of comparisons were made: in one case stage matching (i.e. number of somites counted) was the highest priority, in the other case the pools were designed such that a maximum number of litter mates were compared while a difference of up to five somites was allowed. cDNAs were synthesized and Cy-3/Cy-5 labeled cRNAs were generated by using a Low RNA Input Fluorescent Linear Amplification Kit (Agilent Technologies, Santa Clara, CA). The labeled cRNAs were hybridized on 4X44K Agilent Whole Mouse Genome Dual Color Microarrays (G4122F). Two dye-swap experiments were performed, resulting in four individual arrays. Microarray signal and background information were retrieved with Feature Extraction (V9.5.3, Agilent technologies). All data analyses were performed by using ArrayAssist (5.5.1, Stratagene Inc, La Jolla, CA) and Microsoft Excel (Microsoft Corporation, Redmont, WA). Genetic background: C57BL/6J were mutagenized, and crossed against FVB/NJ mice for positional mapping. The background of the mice used in the experiment is hybrid C57BL/6J *FVB/NJ with a percentage FVB between 50 and 90%.
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| Contributor(s) |
van Gurp L, Wansleeben C, Meijlink F |
| Citation(s) |
21419221 |
| Submission date |
Jan 20, 2011 |
| Last update date |
May 10, 2018 |
| Contact name |
Frits Meijlink |
| E-mail(s) |
f.meijlink@hubrecht.eu
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| Phone |
+31 30 2121894
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| Organization name |
Hubrecht Institute
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| Street address |
Uppsalalaan 8
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| City |
Utrecht |
| ZIP/Postal code |
3584CT |
| Country |
Netherlands |
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| Platforms (1) |
| GPL4134 |
Agilent-014868 Whole Mouse Genome Microarray 4x44K G4122F (Feature Number version) |
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| Samples (4)
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| GSM658205 |
Btaf1_mutant vs Wt stage matched |
| GSM658206 |
Btaf1_mutant vs Wt stage matched dye swap |
| GSM658207 |
Btaf1_mutant vs Wt age matched |
| GSM658208 |
Btaf1_mutant vs Wt age matched dye swap |
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| Relations |
| BioProject |
PRJNA136329 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE26743_RAW.tar |
69.3 Mb |
(http)(custom) |
TAR (of TXT) |
| Processed data included within Sample table |
| Processed data provided as supplementary file |
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