Zinc-finger transcription factors are associated with guanine quadruplex motifs in human, chimpanzee, mouse and rat promoters genome-wide

Abstract

Function of non-B DNA structures are poorly understood though several bioinformatics studies predict role of the G-quadruplex DNA structure in transcription. Earlier, using transcriptome profiling we found evidence of widespread G-quadruplex-mediated gene regulation. Herein, we asked whether potential G-quadruplex (PG4) motifs associate with transcription factors (TF). This was analyzed using 220 position weight matrices [designated as transcription factor binding sites (TFBS)], representing 187 unique TF, in >75,000 genes in human, chimpanzee, mouse and rat. Results show binding sites of nine TFs, including that of AP-2, SP1, MAZ and VDR, occurred significantly within 100 bases of the PG4 motif (P < 1.24E-10). PG4-TFBS combinations were conserved in 'orthologously' related promoters across all four organisms and were associated with >850 genes in each genome. Remarkably, seven of the nine TFs were zinc-finger binding proteins indicating a novel characteristic of PG4 motifs. To test these findings, transcriptome profiles from human cell lines treated with G-quadruplex-specific molecules were used; 66 genes were significantly differentially expressed across both cell-types, which also harbored conserved PG4 motifs along with one/more of the nine TFBS. In addition, genes regulated by PG4-TFBS combinations were found to be co-regulated in human tissues, further emphasizing the regulatory significance of the associations.

Figure 1.

Flowchart summarizing the approach adopted in this study. Schema of the strategy followed to test genome wide association of TF with PG4 motif(s).

Figure 2.

PG4 motif positional conservation across orthologously related promoters. Scheme showing identification of conserved PG4 motif within ±200 bases in orthologously related promoters (±2 kb of TSS) of human, chimpanzee, mouse and rat and search for associated TFBS. H represents a human gene and C, M and R represent their orthologous in chimpanzee, mouse and rat, respectively.

Figure 3.

Strategy followed for genome wide comparative analysis to identify enriched presence of TFBS within promoters harboring conserved PG4 motifs in human, chimpanzee, mouse and rat.

Figure 4.

Zinc finger TFBS are closely associated with PG4 motifs. Distribution of TFBS with respect to PG4 motif (PG4–TFBS inter-distance) in promoters of human, chimpanzee, mouse and rat that harbor conserved PG4 motifs. Pseudocolor represents percentage frequency of PG4–TFBS inter-distance values in bins of 100 bases relative to nearest PG4 motif. Asterisk represents additional zinc finger TF found in the study.

Figure 5.

Genes harboring conserved PG4–TFBS associations are differentially expressed in presence of G-quadruplex binding ligand. Left panel: expression profile of genes with conserved PG4 motif that have significant differential expression in both cell lines on treatment with ligand. Pseudocolor representing their relative expression values in HeLaS3 and A549 cells. Right panel: association of PG4 motif with TFBS in promoters of differentially expressed genes shown in left panel; black box represents PG4 motif and associated pseudocolor shows number of TFBS within 100-base windows relative to TSS.