Nuclear factor I revealed as family of promoter binding transcription activators

Abstract

BackgroundMultiplex experimental assays coupled to computational predictions are being increasingly employed for the simultaneous analysis of many specimens at the genome scale, which quickly generates very large amounts of data. However, inferring valuable biological information from the comparisons of very large genomic datasets still represents an enormous challenge.ResultsAs a study model, we chose the NFI/CTF family of mammalian transcription factors and we compared the results obtained from a genome-wide study of its binding sites with chromatin structure assays, gene expression microarray data, and in silico binding site predictions. We found that NFI/CTF family members preferentially bind their DNA target sites when they are located around transcription start sites when compared to control datasets generated from the random subsampling of the complete set of NFI binding sites. NFI proteins preferably associate with the upstream regions of genes that are highly expressed and that are enriched in active chromatin modifications such as H3K4me3 and H3K36me3. We postulate that this is a causal association and that NFI proteins mainly act as activators of transcription. This was documented for one member of the family (NFI-C), which revealed as a more potent gene activator than repressor in global gene expression analysis. Interestingly, we also discovered the association of NFI with the tri-methylation of lysine 9 of histone H3, a chromatin marker previously associated with the protection against silencing of telomeric genes by NFI.ConclusionTaken together, we illustrate approaches that can be taken to analyze large genomic data, and provide evidence that NFI family members may act in conjunction with specific chromatin modifications to activate gene expression.

Characterization of the neural stem cell gene regulatory network identifies OLIG2 as a multifunctional regulator of self-renewal

“…All antibodies used had all been previously used for ChIP and validated for their specificity. Immunoprecipitations were with mouse anti-ASCL1 (BD Pharmingen) (Castro et al 2006), rabbit anti TCF3 (Santa Cruz, sc-349) (Lin et al 2010), rabbit anti-OLIG2 (Millipore, AB9610) (Mazzoni et al 2011), rabbit anti-MAX (Santa Cruz, sc-197) (Rahl et al 2010), goat anti-NFI (Santa Cruz, sc-30918) (Pjanic et al 2011;Martynoga et al 2013), goat anti-SOX2 (Santa Cruz sc-17320) (Chen et al 2008), rabbit anti-SOX9 (Millipore, AB5535) (Mead et al 2013), and goat anti-SOX21 (R&D systems, AF3538) (Matsuda et al 2012). Primers used for ChIP-PCR are listed in Supplemental Table S5.…”

An Nfic-hedgehog signaling cascade regulates tooth root development

“…The NF-I transcription factors bind to the dyad symmetric consensus sequence TTGGCNNNNNGCCAA and to individual half-sites with reduced affinity (Gronostajski et al, 1985;Meisterernst et al, 1988). Previous studies have found that NF-I transcription factors preferentially bind DNA target sites that are located close to transcription start sites and mainly act as activators of transcription of downstream target genes (Pjanic et al, 2011). In this study, we provide in vivo evidence that Hh signaling participates in root development using lineage tracing and cell ablation of Gli1 + cells.…”

“…5F,G). Previous studies have demonstrated that NF-I transcription factors preferentially bind their DNA target sites around transcription start sites and that they mainly act as activators of transcription (Pjanic et al, 2011). Therefore, we searched for NF-I binding sites within 2.5 kb upstream of the transcription start site of Hhip using two different web servers.…”

“…Nuclei preparation and chromatin digestion were performed according to the manufacturer's protocols. Chromatin fragments were incubated with anti-NF-I antibody (Santa Cruz, sc-5567x; 1:50) (Pjanic et al, 2011) or IgG control antibody. After DNA purification, the following PCR primers were used: NF-I binding site (BS) 1, 5′-gctaggccaaatgtctctgc-3′ and 5′-ggtggaggatgaagctggta-3′; BS 2, 5′-gtctatgcctgccagagagg-3′ and 5′-aggaatctttgcctggcttt-3′.…”

Epigenomic enhancer annotation reveals a key role for NFIX in neural stem cell quiescence

“…S4A-C). To examine NFI protein binding to the NS cell genome, we performed a ChIP-seq analysis with an antibody that specifically recognizes the four NFI factors, which are closely related in sequence (Mason et al 2009;Pjanic et al 2011). NFI factors bound to 25,807 high-confidence sites in quiescent NS cells (Fig.…”