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| Status |
Public on Oct 17, 2013 |
| Title |
Genome-wide binding profiles of KLF3 and KLF3 mutants in MEF cells |
| Organism |
Mus musculus |
| Experiment type |
Genome binding/occupancy profiling by high throughput sequencing
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| Summary |
Transcription factors are often regarded as being comprised of a DNA-binding domain and a functional domain. The two domains are considered separable and autonomous, with the DNA-binding domain directing the factor to its target genes and the functional domain imparting transcriptional regulation. We have examined a typical Zinc Finger (ZF) transcription factor from the Krüppel-like factor (KLF) family, KLF3. This factor has an N-terminal repression domain that binds the co-repressor C-terminal binding protein (CtBP), and a DNA-binding domain composed of three classical (ZFs) at its C-terminus. We established a system to compare the genomic occupancy profile of wildtype KLF3 with two mutants affecting the N-terminal functional domain: a mutant unable to contact its cofactor CtBP and a mutant lacking the entire N-terminal domain, but retaining the ZFs intact. We used chromatin immunoprecipitation followed by sequencing (ChIP-seq) to assess binding across the genome in murine embryonic fibroblasts. Our results define the in vivo recognition site for KLF3 and the two mutants as a typical CACCC-like element. Unexpectedly, we observe that mutations in the N-terminal functional domain severely affect DNA binding. In general, both mutations reduce binding but there are also instances where binding is retained or even increased. These results provide a clear demonstration that the correct localization of transcription factors to their target genes is not solely dependent on their DNA-contact domains. This informs our understanding of how transcription factors operate and is of relevance to the design of artificial ZF proteins.
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| Overall design |
ChIP-seq was performed on the three samples, KLF3, ΔDL and DBD in duplicate (biological replicates). Input samples were used as controls.
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| Contributor(s) |
Burdach J, Funnell AP, Artuz CM, Sin MK, Tan LY, Pearson RC, Crossley M |
| Citation(s) |
24106088 |
| Submission date |
Feb 28, 2013 |
| Last update date |
May 15, 2019 |
| Contact name |
Jon Burdach |
| Organization name |
University of New South Wales
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| Department |
BABS
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| Street address |
UNSW
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| City |
Sydney |
| State/province |
NSW |
| ZIP/Postal code |
2052 |
| Country |
Australia |
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| Platforms (1) |
| GPL13112 |
Illumina HiSeq 2000 (Mus musculus) |
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| Samples (6)
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| Relations |
| BioProject |
PRJNA191577 |
| SRA |
SRP019227 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE44748_All_peaks.txt.gz |
1.4 Mb |
(ftp)(http) |
TXT |
| GSE44748_DBD-1_peaks.txt.gz |
754.0 Kb |
(ftp)(http) |
TXT |
| GSE44748_DBD-2_peaks.txt.gz |
670.5 Kb |
(ftp)(http) |
TXT |
| GSE44748_DBD_merged_peaks.txt.gz |
434.3 Kb |
(ftp)(http) |
TXT |
| GSE44748_DDL-1_peaks.txt.gz |
1.4 Mb |
(ftp)(http) |
TXT |
| GSE44748_DDL-2_peaks.txt.gz |
2.7 Mb |
(ftp)(http) |
TXT |
| GSE44748_DDL_merged_peaks.txt.gz |
1.1 Mb |
(ftp)(http) |
TXT |
| GSE44748_KLF3-1_peaks.txt.gz |
2.3 Mb |
(ftp)(http) |
TXT |
| GSE44748_KLF3-2_peaks.txt.gz |
2.1 Mb |
(ftp)(http) |
TXT |
| GSE44748_KLF3_merged_peaks.txt.gz |
1.3 Mb |
(ftp)(http) |
TXT |
| GSE44748_RAW.tar |
1.9 Gb |
(http)(custom) |
TAR (of BEDGRAPH) |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
| Processed data are available on Series record |
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