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| Status |
Public on Dec 20, 2017 |
| Title |
ChIP-seq analysis of HN1L protein in SUM159 triple-negative breast cancer cells |
| Organism |
Homo sapiens |
| Experiment type |
Genome binding/occupancy profiling by high throughput sequencing
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| Summary |
Purpose: The goal of this study is to identify the role of HN1L protein as a transcription factor or co-factor in regulating TNBC cells.
Methods: Due to the unavailability of a ChIP-grade HN1L antibody, we overexpressed FLAG-tagged HN1L in SUM159 cells and performed ChIP using anti-FLAG antibodies. ChIP DNA was prepared into libraries and sequenced by the Epigenomics Core of Weill Cornell Medical College using SR50 lane.
Results: ChIP-Seq analysis began with mapping the sequenced reads to the genome. We utilized the Burrows-Wheeler Aligner (BWA) MEM algorithm to align the sequence reads against the human genome GRCh37/hg19 Assembly. We next used the Hypergoemetric Optimization of Motif Enrichment (HOMER) suite of tools to find and annotate peaks, and identify enriched motifs. HN1L showed 2,249 binding peaks from 10k bp upstream to 5k bp downstream of transcription start site (TSS).
Conclusions: among the binding peaks 35 genes overlapped with a previously published BCSC gene signature, 10 genes overlapped with the HN1L knockdown gene signature and 8 genes are well established CSC transcription factors. When applying overlapped genes in the STRING10 pathway analysis database , a protein interaction network centered with STAT3 was obtained. STAT3 and FGFR2 peaks were validated by qPCR. Model-based Analysis of ChIP-Seq (MACS) was then used to confirm the peaks found by Hypergeometric Optimization of Motif EnRichment (HOMER) . Besides the binding peaks found in HOMER, another peak was called by MACS within LEPR , and was also validated by qPCR . These findings indicate that HN1L may act as a transcription factor through binding to enhancer regions of STAT3 and other STAT3 regulators.
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| Overall design |
Gene profile directly bound and transcriptionly regulated by HN1L protein were generated by Illumina HiSeq2000 system with each library sequenced in a 50 bases single-read run.
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| Contributor(s) |
Chang JC, Liu Y, Choi DS, Benz S, Polley A |
| Citation(s) |
29249663 |
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| Submission date |
Oct 20, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Jenny Chang |
| Organization name |
Houston Methodist Hospital
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| Department |
Houston Methodist Cancer Center
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| Street address |
6670 Bertner Ave.
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| City |
Houston |
| State/province |
TX |
| ZIP/Postal code |
77030 |
| Country |
USA |
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| Platforms (1) |
| GPL11154 |
Illumina HiSeq 2000 (Homo sapiens) |
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| Samples (2) |
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| Relations |
| BioProject |
PRJNA415189 |
| SRA |
SRP120608 |
